The document discusses chromatographic strategies for intravenous immunoglobulin (IVIG) purification using anion exchange chromatography. It describes a case study where a Fractogel EMD TMAE (M) resin was able to efficiently separate and purify IgG from a caprylic acid-treated human plasma fraction (worst-case scenario) in a single step in negative mode. Optimization studies showed the resin was robust across a pH range of 5.7-6.3, allowing selective binding of contaminants like IgA and IgM while IgG passed through. Purification trials over 10 cycles demonstrated consistent 94% IgG recovery and 84% removal of IgA and IgM contaminants.
A Turn-Key Flow-Through-Mode Purification Process to improve Quality and Safe...Merck Life Sciences
In this webinar, you will learn:
Intensified plasma Immunoglobulin purifications
Scalable process development with latest technologies
Improved safety and quality of plasma IgG meeting required quality attributes
Detailed description:
Plasma-derived immunoglobulins (IgG) are essential medicines that are in worldwide shortage. How to develop optimized processing steps for robust and efficient manufacturing has been a constant goal, to make the most out of the precious plasma raw material.
In this study, we present a worse-case equivalent of plasma intermediate, explore various process steps along the fractionation flow, including flow-through-mode chromatography, affinity chromatography, virus inactivation steps and removal of solvent/detergent, single-pass TFF (SPTFF), clarification, and aseptic filtration, to establish a robust, easy-to-operate, readily scalable plasma IgG process with over 99% purity, depletion of IgA, isoagglutinin, and thrombogenic markers, meeting the commonly required 20% concentration for subcutaneous IgG infusion. Such solutions would be appropriate for various IgG intermediates which help to improve the global supply of immunoglobulins.
Extractables profiles for chromatography resins - adapted approach of upcomin...Merck Life Sciences
Watch the webinar here: https://bit.ly/36JaZpx
In biopharmaceutical industry there is a trend towards comprehensive risk assessments of drug manufacturing processes. Extractables studies for chromatography resins based on the adapted requirements of the upcoming USP <665> support risk evaluation for your specific chromatography steps.
In this webinar, you will learn about:
- Study design for extractables profiles of chromatography resins
- The new category Emprove® Chromatography
- Communication of extractables data as part of Emprove® Dossiers
Description:
Detailed information on any component or material in contact with the drug substance/ product is required to conduct a compreshensive risk assessment of a biopharmaceutical manufacturing process. No explicit guidelines providing required testing procedures for chromatography steps are in place yet. In the upcoming USP <665> chapter chromatography steps are in focus as well as any other plastic or polymeric component and can as such assessed as to the described criteria. To support our chromatography resin users an adapted extractables study approach was developed. The webinar will demonstrate our study design and the communication of the extractables profiles within our Emprove® Program.
Overcoming challenges of host cell DNA removal in vaccine manufacturingDr. Priyabrata Pattnaik
Regulatory agencies require residual host cell DNA in vaccines to be extremely low, typically below 10 pg/dose. Various methods are used to remove DNA during vaccine manufacturing, including nuclease treatment, adsorptive depth filtration, chromatography, and tangential flow filtration. Nuclease treatment with Benzonase is widely used to digest DNA but the nuclease then needs to be removed using techniques like anion exchange chromatography, gel filtration, or ultrafiltration with diafiltration to achieve over 99% clearance.
Process Development for Cell Therapy and Viral Gene TherapyMerck Life Sciences
Today’s viral vector manufacturing processes remain challenging. Process development is a critical enabler to bring safe, effective, sustainable products to market to address patient needs. When done properly, it can reduce the timeline of the project and the cost of producing the therapeutic product.
The webinar discusses our strategies for developing lentivirus and adeno associated virus (AAV) and the impact these early decisions can have on commercial readiness.
Watch the interactive webinar now: https://bit.ly/2VplwQq
Process development considerations for quality and safety of vaccinesDr. Priyabrata Pattnaik
The document discusses several factors that can impact vaccine quality and safety during development, including:
1) Bioburden control is important to control contamination during manufacturing and avoid issues in later stages. Key areas are raw materials, equipment cleaning, and open processing steps.
2) Operating conditions for tangential flow filtration, such as pressure and flow rates, can cause product aggregation or degradation if not optimized.
3) Residual DNA from cell substrates must be removed through processes like nuclease treatment to very low levels due to potential safety concerns.
4) Excipient quality can impact drug product safety, so their selection and control is a critical quality attribute during development.
Validation of Tangential Flow Filtration in Biotech ProcessesMerck Life Sciences
Watch the presentation of this webinar here: https://bit.ly/3hUKfd7
The objective of validation of a unit operation is to demonstrate with a high degree of confidence that the process performs consistently. The present seminar will focus on the validation of the unit operation of TFF and will provide an overview of the regulatory landscape, the validation master plan, approaches to membrane re-use, cleaning validation, and best practices.
In this webinar, you will learn:
• Validation of TFF
• Validation master plan
• Membrane reuse and cleaning
• TFF scale down models
Speaker: Dr. Subhasis Banerjee,
Principal Technical Application Expert, Bioprocessing APAC
Rapid replication competent adenovirus (rRCA) detection: Accelerate your lot ...Merck Life Sciences
Watch the presentation of this webinar here: https://bit.ly/3MJ4u9V
Testing for presence of replication competent adenovirus (RCA) is a key component to ensure patient safety and a requirement for all biologicals manufactured using adenoviral vectors. For many adenoviral-based products, the RCA assay is a rate-limiting assay for lot release.
Join this webinar to learn about a rapid RCA detection assay currently in development, which combines a 7-day culture assay with a highly sensitive molecular endpoint specific for RCA. The method can detect presence of as little as 1 RCA in adenoviral vector material at an approximate concentration of 5x107 - 2x108 vector particles (VP)/mL, making it a suitable method to meet regulatory requirements while accelerating your lot release timelines.
In this webinar, you will learn about:
• Regulatory framework for adenoviral vector products
• Considerations for lot release testing of adenoviral-based therapies
• Advantages of a rapid method for RCA testing on production lot material
Presented by:
Axel Fun, Ph.D.,
Principal Scientist
Alberto Santana, MBA,
Product Manager, Biologics Biosafety Testing
Releasing Your AAV Therapy with Confidence: Regulatory Considerations and Key...Merck Life Sciences
Watch the presentation of this webinar here: https://bit.ly/3icKkbZ
Ensuring the safety and quality of your AAV vector is of the utmost importance. Join this webinar for a high-level overview of the regulatory requirements for AAV testing throughout the manufacturing process, as well as a more detailed look at rcAAV and infectious titer assays.
Adeno-associated virus (AAV) vectors possess a number of advantages for use in human therapy including: high titer preparations, low immunogenicity, capacity to infect a wide range of cell types, and replication deficiency. Even with these advantages, there are biosafety concerns to consider when using AAV vectors.
This webinar will discuss key regulatory considerations across the manufacturing process, from the helper/packaging plasmids through to lot release testing. We will highlight critical assays that are required and delve into specifics on replication competent AAV testing and infectious titer determination by TCID50.
In this webinar, you will learn:
• Critical biosafety considerations for AAV vectors based on the latest regulatory guidance
• How replication competent AAV testing fits into your bulk and final release testing package
• The benefits of routine and platform assays over custom assay development
Presented by:
Steven McDade, Senior Technical Specialist, Field Technology Management
Alfonso Lavorgna, Ph.D., Operations Manager, Virology Services
A Turn-Key Flow-Through-Mode Purification Process to improve Quality and Safe...Merck Life Sciences
In this webinar, you will learn:
Intensified plasma Immunoglobulin purifications
Scalable process development with latest technologies
Improved safety and quality of plasma IgG meeting required quality attributes
Detailed description:
Plasma-derived immunoglobulins (IgG) are essential medicines that are in worldwide shortage. How to develop optimized processing steps for robust and efficient manufacturing has been a constant goal, to make the most out of the precious plasma raw material.
In this study, we present a worse-case equivalent of plasma intermediate, explore various process steps along the fractionation flow, including flow-through-mode chromatography, affinity chromatography, virus inactivation steps and removal of solvent/detergent, single-pass TFF (SPTFF), clarification, and aseptic filtration, to establish a robust, easy-to-operate, readily scalable plasma IgG process with over 99% purity, depletion of IgA, isoagglutinin, and thrombogenic markers, meeting the commonly required 20% concentration for subcutaneous IgG infusion. Such solutions would be appropriate for various IgG intermediates which help to improve the global supply of immunoglobulins.
Extractables profiles for chromatography resins - adapted approach of upcomin...Merck Life Sciences
Watch the webinar here: https://bit.ly/36JaZpx
In biopharmaceutical industry there is a trend towards comprehensive risk assessments of drug manufacturing processes. Extractables studies for chromatography resins based on the adapted requirements of the upcoming USP <665> support risk evaluation for your specific chromatography steps.
In this webinar, you will learn about:
- Study design for extractables profiles of chromatography resins
- The new category Emprove® Chromatography
- Communication of extractables data as part of Emprove® Dossiers
Description:
Detailed information on any component or material in contact with the drug substance/ product is required to conduct a compreshensive risk assessment of a biopharmaceutical manufacturing process. No explicit guidelines providing required testing procedures for chromatography steps are in place yet. In the upcoming USP <665> chapter chromatography steps are in focus as well as any other plastic or polymeric component and can as such assessed as to the described criteria. To support our chromatography resin users an adapted extractables study approach was developed. The webinar will demonstrate our study design and the communication of the extractables profiles within our Emprove® Program.
Overcoming challenges of host cell DNA removal in vaccine manufacturingDr. Priyabrata Pattnaik
Regulatory agencies require residual host cell DNA in vaccines to be extremely low, typically below 10 pg/dose. Various methods are used to remove DNA during vaccine manufacturing, including nuclease treatment, adsorptive depth filtration, chromatography, and tangential flow filtration. Nuclease treatment with Benzonase is widely used to digest DNA but the nuclease then needs to be removed using techniques like anion exchange chromatography, gel filtration, or ultrafiltration with diafiltration to achieve over 99% clearance.
Process Development for Cell Therapy and Viral Gene TherapyMerck Life Sciences
Today’s viral vector manufacturing processes remain challenging. Process development is a critical enabler to bring safe, effective, sustainable products to market to address patient needs. When done properly, it can reduce the timeline of the project and the cost of producing the therapeutic product.
The webinar discusses our strategies for developing lentivirus and adeno associated virus (AAV) and the impact these early decisions can have on commercial readiness.
Watch the interactive webinar now: https://bit.ly/2VplwQq
Process development considerations for quality and safety of vaccinesDr. Priyabrata Pattnaik
The document discusses several factors that can impact vaccine quality and safety during development, including:
1) Bioburden control is important to control contamination during manufacturing and avoid issues in later stages. Key areas are raw materials, equipment cleaning, and open processing steps.
2) Operating conditions for tangential flow filtration, such as pressure and flow rates, can cause product aggregation or degradation if not optimized.
3) Residual DNA from cell substrates must be removed through processes like nuclease treatment to very low levels due to potential safety concerns.
4) Excipient quality can impact drug product safety, so their selection and control is a critical quality attribute during development.
Validation of Tangential Flow Filtration in Biotech ProcessesMerck Life Sciences
Watch the presentation of this webinar here: https://bit.ly/3hUKfd7
The objective of validation of a unit operation is to demonstrate with a high degree of confidence that the process performs consistently. The present seminar will focus on the validation of the unit operation of TFF and will provide an overview of the regulatory landscape, the validation master plan, approaches to membrane re-use, cleaning validation, and best practices.
In this webinar, you will learn:
• Validation of TFF
• Validation master plan
• Membrane reuse and cleaning
• TFF scale down models
Speaker: Dr. Subhasis Banerjee,
Principal Technical Application Expert, Bioprocessing APAC
Rapid replication competent adenovirus (rRCA) detection: Accelerate your lot ...Merck Life Sciences
Watch the presentation of this webinar here: https://bit.ly/3MJ4u9V
Testing for presence of replication competent adenovirus (RCA) is a key component to ensure patient safety and a requirement for all biologicals manufactured using adenoviral vectors. For many adenoviral-based products, the RCA assay is a rate-limiting assay for lot release.
Join this webinar to learn about a rapid RCA detection assay currently in development, which combines a 7-day culture assay with a highly sensitive molecular endpoint specific for RCA. The method can detect presence of as little as 1 RCA in adenoviral vector material at an approximate concentration of 5x107 - 2x108 vector particles (VP)/mL, making it a suitable method to meet regulatory requirements while accelerating your lot release timelines.
In this webinar, you will learn about:
• Regulatory framework for adenoviral vector products
• Considerations for lot release testing of adenoviral-based therapies
• Advantages of a rapid method for RCA testing on production lot material
Presented by:
Axel Fun, Ph.D.,
Principal Scientist
Alberto Santana, MBA,
Product Manager, Biologics Biosafety Testing
Releasing Your AAV Therapy with Confidence: Regulatory Considerations and Key...Merck Life Sciences
Watch the presentation of this webinar here: https://bit.ly/3icKkbZ
Ensuring the safety and quality of your AAV vector is of the utmost importance. Join this webinar for a high-level overview of the regulatory requirements for AAV testing throughout the manufacturing process, as well as a more detailed look at rcAAV and infectious titer assays.
Adeno-associated virus (AAV) vectors possess a number of advantages for use in human therapy including: high titer preparations, low immunogenicity, capacity to infect a wide range of cell types, and replication deficiency. Even with these advantages, there are biosafety concerns to consider when using AAV vectors.
This webinar will discuss key regulatory considerations across the manufacturing process, from the helper/packaging plasmids through to lot release testing. We will highlight critical assays that are required and delve into specifics on replication competent AAV testing and infectious titer determination by TCID50.
In this webinar, you will learn:
• Critical biosafety considerations for AAV vectors based on the latest regulatory guidance
• How replication competent AAV testing fits into your bulk and final release testing package
• The benefits of routine and platform assays over custom assay development
Presented by:
Steven McDade, Senior Technical Specialist, Field Technology Management
Alfonso Lavorgna, Ph.D., Operations Manager, Virology Services
Process development guidance for AAV and lentivirus manufacturing based on co...MilliporeSigma
Access the interactive recording here: https://bit.ly/37nl3Ex
Webinar summary:
An efficient production platform is essential for successful commercial implementation of gene therapy programs. AAV and Lentivirus manufacturing process are often developed with compressed timelines, reduced process optimization and low product yields which can have significant effect on costs.
In this webinar, you will learn:
* How manufacturing costs are examined for adeno-associated virus and lentivirus production with several different for each vector
* That key process characteristics like production titer, production of empty viral particles, downstream product recovery, and the batching strategy can effect the overall manufacturing cost
* How holistic evaluation is an important tool during process development to help prioritize different approaches to improve viral vector production processes
Abstract:
An efficient production platform is essential for successful commercial implementation of gene therapy programs. Viral vector manufacturing processes are often developed under timelines which are considerably shorter than those for more mature biopharmaceuticals. Consequently, the level of process optimization is reduced and challenges related to low product yields are common. These factors, as well as the small batch sizes common for these processes, can have significant effect on manufacturing costs.
This presentation provides an introduction to tangential flow filtration and reviews the following:
- TFF process basics and terminology
- TFF membrane technology
- TFF hardware, devices and systems
- Growing applications and the future
To learn more about this topic or collaborate with our technical experts, schedule an in-person or remote visit at our M Lab™ Collaboration Centers: www.merckmillipore.com/mlab
Optimization of Tangential Flow Filtration Applications and Scale Up Consider...MilliporeSigma
This presentation provides an introduction to tangential flow filtration applications for AAV and lentivirus and will review:
• Basics of tangential flow filtration (TFF)
• TFF AAV and lentivirus process overview
• Operating parameters optimization: flux-controlled microfiltration
• Scale up considerations
To learn more about this topic or collaborate with our technical experts, schedule a remote visit at our M Lab™ Collaboration Centers: www.emdmillipore.com/remotevisit
Commercializing antibody-drug conjugates: a CMO’s journeyMerck Life Sciences
Watch the webinar here: https://bit.ly/2YLDzTE
This webinar will take you through the story of a CMO preparing for the manufacture of a Commercial Antibody Drug Conjugate (ADC).
Join us to learn about how we grew as a CMO to develop a Commercial ADC program. We will walk through the full timeline from development, process risk assessment and control strategy development and validation, finishing off with preparation for a Commercial ADC Pre-Approval Inspection Audit.
In this webinar you will learn how to:
• Properly structure your development work
• Perform a thorough process risk assessment
• Prepare for pre-approval inspection
This presentation explores bioprocessing filtration best practices, including design and scale up methods. You will learn:
• What is filtration?
• Filter capacity and fouling models
• Filter sizing approaches
• Scale up considerations
To learn more about this topic or collaborate with our technical experts, schedule a remote visit at our M Lab™ Collaboration Centers: www.merckmillipore.com/remotevisit
Selection, sizing, and operation of bioprocess filtration trains for optimal ...Merck Life Sciences
To increase filter lifetime and improve the economics of filtering bioprocess streams, a prefilter is often installed upstream of a final sterilizing-grade filter. However, determining the economic optimum prefilter and final filter configuration can be challenging. Numerous prefilter options are available, the prefilter to final filter area ratio must be determined, and operating conditions must be selected that will both satisfy the filtration requirements and provide for an economical process that minimizes the filtration system footprint.
One approach towards achieving an optimal filtration system design is to test the bioprocess fluid with several filter configuration combinations and at a range of operating conditions. However, this can be a daunting task and even impractical given the high cost and limited availability of valuable bioprocess fluids. A better approach is to run a limited filtration trial and use a mathematical model that can accurately predict the behavior of the prefilter and final filter under different conditions.
In this webinar we describe a filtration model and test methodology to rapidly and efficiently design an optimal dual-stage filtration process. The model and methodology were applied to Milligard® PES filters, a new class of autoclavable and gamma sterilizable PES membrane prefilters that are designed to protect microfiltration and nanofiltration final filters in bioprocess streams. We show how a model fit to the data from one set of filtration conditions can be used to predict filtration performance at other prefilter to final filter area ratios and operating conditions, and to determine the economic optimum filtration configuration.
In this webinar, you will learn:
- How filters for microfiltration of biological fluids work.
- The effect of operating conditions on filtration performance.
- How to design an optimal series filtration (prefilter and final filter) process.
Process development guidance for AAV and lentivirus manufacturing based on co...Merck Life Sciences
Access the interactive recording here: https://bit.ly/37nl3Ex
Webinar summary:
An efficient production platform is essential for successful commercial implementation of gene therapy programs. AAV and Lentivirus manufacturing process are often developed with compressed timelines, reduced process optimization and low product yields which can have significant effect on costs.
In this webinar, you will learn:
* How manufacturing costs are examined for adeno-associated virus and lentivirus production with several different for each vector
* That key process characteristics like production titer, production of empty viral particles, downstream product recovery, and the batching strategy can effect the overall manufacturing cost
* How holistic evaluation is an important tool during process development to help prioritize different approaches to improve viral vector production processes
Abstract:
An efficient production platform is essential for successful commercial implementation of gene therapy programs. Viral vector manufacturing processes are often developed under timelines which are considerably shorter than those for more mature biopharmaceuticals. Consequently, the level of process optimization is reduced and challenges related to low product yields are common. These factors, as well as the small batch sizes common for these processes, can have significant effect on manufacturing costs.
Introduction to Tangential Flow Filtration (TFF)MilliporeSigma
This presentation provides an introduction to tangential flow filtration and reviews the following:
- TFF process basics and terminology
- TFF membrane technology
- TFF hardware, devices and systems
- Growing applications and the future
To learn more about this topic or collaborate with our technical experts, schedule an in-person or remote visit at our M Lab™ Collaboration Centers: www.emdmillipore.com/mlab
Does your cell line have a secret? Avoid surprises with characterizationMerck Life Sciences
Watch the recording of this webinar here: https://bit.ly/2Y05bV4
The first step to avoiding an unpleasant and costly contamination event is characterization of your cell banks.
Regardless of the biotech product, careful characterization of the cell banks used in its production is the first step in mitigating the risk of a contamination event. In fact, cell line characterization is an important component of the overall viral safety strategy for the product. We will describe the testing necessary to ensure cell banks are free from infectious and other adverse agents and that meets current regulatory expectations. Different levels of testing are performed for master, working, and end of production cell banks, and the differences in testing for each of these types of banks will be discussed.
In this webinar, you will learn:
• The types of tests that are needed to fully characterize your cell banks
• The best tests to use for your particular cell line
• Reasons why a viral contaminant may be missed
Find your filter. What’s best for your process? MilliporeSigma
Selecting the right aseptic filter for your process can be complicated: today’s biomanufacturer has many filter choices each offering distinct benefits. Understanding the specific needs for individual operations, in terms of flux, capacity, bioburden reduction or sterilizing performance, gamma or thermal compatibility and single or multi-use will inform decisions that have implications for the life of the process. This webinar will provide general customer guidance and explain the benefits and disadvantages of different options to help guide customers to the most appropriate filter for their operation.
In this webinar, you will learn:
- How filter design impacts performance
- Important criteria for filter selection
- New choices and options to maximize productivity for biomanufacturers
This document discusses biosafety testing for cell and gene therapies performed by BioReliance, a testing services division of Merck KGaA. It outlines the comprehensive testing performed at various stages of development, including testing of cell banks, viral vectors, and final drug products. Testing evaluates important product attributes like identity, purity, potency and residuals to ensure safety and quality. A wide range of assays are used to characterize products and identify potential contaminants.
Parvovirus Filtration Best Practices - 25 Years of Hands-On ExperienceMerck Life Sciences
In this webinar, you will learn:
- how to measure filter performance and capacity,
- how to optimize filter virus removal capability,
- and avoid potential pit-falls
Detailed description:
This webinar will cover all aspects of parvovirus filtration best practices: process development/ optimization, pilot scale-up, and validation and explain the important connections between these activities. The rationale for the recommended best practices will be explained by discussing the underlying mechanisms that control filter performance.
Filter Validation and Regulatory aspectsbala murugan
The document discusses various aspects of filter validation including:
1) Common materials used in filters include regenerated cellulose, nylon, polyamides, and polyether sulfones.
2) Porosity refers to the total empty space in a filter and affects its performance.
3) Validation tests include bacterial challenge tests, bubble point tests, and diffusion tests to evaluate a filter's effectiveness.
4) Bubble point tests determine a filter's pore size while diffusion tests measure gas flow rates through a filter.
In this webinar, you will learn:
How Single-Use helps to simplify ADC manufacturing
Safety and Integrity of the complete process
Flexibility and adaptability to changing needs
How to speed up the synthesis of high-potent payloads and improve the bioavailability of the ADC
Detailed description:
The development of an ADC is a long journey that can be speeded-up with the adoption of proper manufacturing tools and new linker-payload solutions. Single-Use equipment applied to the GMP manufacturing of ADC means operator safety, decreased risk of contamination, scalability, reproducibility, flexibility, small footprint, lower cost, and saved time through efficiency (easy setup and cleanup, no need for cleaning validation). The adoption of payload intermediates such as DolCoreTM and MayCoreTM can speed your development project up, while innovative linkers like ChetoSensarTM increase its chances of success. This webinar explains how these technologies improve the ADC production process end-to-end, ultimately enhancing patient safety.
Membrane Chromatography Solutions for Single-Use, Intensified mAb PurificationMerck Life Sciences
Participate in the interactive webinar: http://bit.ly/NatrixChromMRK
Improve productivity, flexibility, and economics of mAb purification process with intensified, single-use membrane chromatography.
Explore our webinar library: www.merckmillipore.com/webinars
Complete single-use ADC technology from development through scale-up MilliporeSigma
This webinar will talk about the benefits of single-use technologies for the manufacturing of antibody-drug conjugates and present a successful corresponding case study.
With an expected high annual growth rate of the global Antibody-drug Conjugate (ADC) market, it is essential that CMO’s have robust manufacturing platforms to ensure successful transfer to GMP production.
Single-Use Technologies provide many advantages, including improved safety, lower costs and greater flexibility. This webinar will outline the advantages of a Single Use Platform and give a case study on how it can be used to manufacture ADC projects.
In this webinar, you will learn:
● How single-use technologies can provide benefits for ADC manufacturing
● Why a solid manufacturing platform is crucial for a successful transfer to GMP production
● How a case study demonstrates the advantages of single-use equipment in a scale up to GMP production
How to reach High Plasma Protein Concentration with Single-Pass TFFMilliporeSigma
This webinar will discuss our collaboration with Takeda on the development of a single-pass TFF system as an alternative to traditional TFF for concentrating a plasma-derived IgG solution.
Single-Pass Tangential Flow Filtration (SPTFF) is a technology that requires only one pass through the filter assembly to achieve the desired concentration with no recirculation of product.
SPTFF can offer many advantages in downstream processing, such as:
• Increased capacity and reduced process time
• Increased yield and product recovery
• Optimized processing of highly shear-sensitive products
• Reduced foam formation
• Reduced cost of goods
This presentation will cover our collaboration with Takeda, formerly Shire, for the development of a specific SPTFF system as an alternative to traditional TFF for concentrating a plasma-derived Immunoglobulin G (IgG) solution from 10% to 20%. Due to promising results, plans are underway to replace the currently used batch TFF process with a SPTFF step.
In this webinar, we will discuss:
- A comparison of traditional TFF versus SPTFF
- Design of Experiments (DOE) approach toward initial process development work and determination of the optimal parameters
- Process run results, including final product yield and product quality
Keeping the (Adventitious) Virus Out of the (Adeno-Associated) VirusMilliporeSigma
This document discusses approaches for viral safety in AAV processes. It begins with background on the growing gene therapy market and evolving regulatory landscape. It then outlines a risk mitigation strategy using a tripod approach of prevent, remove, and detect. Specific techniques discussed for prevent include using virus-resistant cell lines and pretreated raw materials. Techniques discussed for remove include virus filtration, chromatography, and the use of Benzonase endonuclease to digest DNA. The document emphasizes a layered, multi-pronged approach for complete biosafety assurance in AAV processes.
Selection, sizing, and operation of bioprocess filtration trains for optimal ...MilliporeSigma
This document discusses the selection, sizing, and operation of bioprocess filtration trains for optimal performance. It covers background on membrane types used in biopharma processing including ultrafiltration, virus retentive, and microfiltration membranes. It then discusses filtration mechanisms, series filtration, and modeling approaches to predict filter performance and sizing including classical fouling models and combined blocking and adsorption models. Experimental data is presented to validate the predictive capability of the combined blocking and adsorption fouling model for sizing filters in a series configuration.
A Turn-Key Flow-Through-Mode Purification Process to improve Quality and Safe...MilliporeSigma
In this webinar, you will learn:
Intensified plasma Immunoglobulin purifications
Scalable process development with latest technologies
Improved safety and quality of plasma IgG meeting required quality attributes
Detailed description:
Plasma-derived immunoglobulins (IgG) are essential medicines that are in worldwide shortage. How to develop optimized processing steps for robust and efficient manufacturing has been a constant goal, to make the most out of the precious plasma raw material.
In this study, we present a worse-case equivalent of plasma intermediate, explore various process steps along the fractionation flow, including flow-through-mode chromatography, affinity chromatography, virus inactivation steps and removal of solvent/detergent, single-pass TFF (SPTFF), clarification, and aseptic filtration, to establish a robust, easy-to-operate, readily scalable plasma IgG process with over 99% purity, depletion of IgA, isoagglutinin, and thrombogenic markers, meeting the commonly required 20% concentration for subcutaneous IgG infusion. Such solutions would be appropriate for various IgG intermediates which help to improve the global supply of immunoglobulins.
This presentation reviews current trends in bioprocessing purification and includes key considerations for continuous processing and connected polishing for monoclonal antibodies. Topics include:
• Market trends and the evolution of next-generation processes
• Intensified capture processing
• Continuous virus inactivation
• Connected flow-through polishing
To learn more about this topic or collaborate with our technical experts, schedule an in-person or remote visit at our M Lab™ Collaboration Centers: www.merckmillipore.com/mlab
Process development guidance for AAV and lentivirus manufacturing based on co...MilliporeSigma
Access the interactive recording here: https://bit.ly/37nl3Ex
Webinar summary:
An efficient production platform is essential for successful commercial implementation of gene therapy programs. AAV and Lentivirus manufacturing process are often developed with compressed timelines, reduced process optimization and low product yields which can have significant effect on costs.
In this webinar, you will learn:
* How manufacturing costs are examined for adeno-associated virus and lentivirus production with several different for each vector
* That key process characteristics like production titer, production of empty viral particles, downstream product recovery, and the batching strategy can effect the overall manufacturing cost
* How holistic evaluation is an important tool during process development to help prioritize different approaches to improve viral vector production processes
Abstract:
An efficient production platform is essential for successful commercial implementation of gene therapy programs. Viral vector manufacturing processes are often developed under timelines which are considerably shorter than those for more mature biopharmaceuticals. Consequently, the level of process optimization is reduced and challenges related to low product yields are common. These factors, as well as the small batch sizes common for these processes, can have significant effect on manufacturing costs.
This presentation provides an introduction to tangential flow filtration and reviews the following:
- TFF process basics and terminology
- TFF membrane technology
- TFF hardware, devices and systems
- Growing applications and the future
To learn more about this topic or collaborate with our technical experts, schedule an in-person or remote visit at our M Lab™ Collaboration Centers: www.merckmillipore.com/mlab
Optimization of Tangential Flow Filtration Applications and Scale Up Consider...MilliporeSigma
This presentation provides an introduction to tangential flow filtration applications for AAV and lentivirus and will review:
• Basics of tangential flow filtration (TFF)
• TFF AAV and lentivirus process overview
• Operating parameters optimization: flux-controlled microfiltration
• Scale up considerations
To learn more about this topic or collaborate with our technical experts, schedule a remote visit at our M Lab™ Collaboration Centers: www.emdmillipore.com/remotevisit
Commercializing antibody-drug conjugates: a CMO’s journeyMerck Life Sciences
Watch the webinar here: https://bit.ly/2YLDzTE
This webinar will take you through the story of a CMO preparing for the manufacture of a Commercial Antibody Drug Conjugate (ADC).
Join us to learn about how we grew as a CMO to develop a Commercial ADC program. We will walk through the full timeline from development, process risk assessment and control strategy development and validation, finishing off with preparation for a Commercial ADC Pre-Approval Inspection Audit.
In this webinar you will learn how to:
• Properly structure your development work
• Perform a thorough process risk assessment
• Prepare for pre-approval inspection
This presentation explores bioprocessing filtration best practices, including design and scale up methods. You will learn:
• What is filtration?
• Filter capacity and fouling models
• Filter sizing approaches
• Scale up considerations
To learn more about this topic or collaborate with our technical experts, schedule a remote visit at our M Lab™ Collaboration Centers: www.merckmillipore.com/remotevisit
Selection, sizing, and operation of bioprocess filtration trains for optimal ...Merck Life Sciences
To increase filter lifetime and improve the economics of filtering bioprocess streams, a prefilter is often installed upstream of a final sterilizing-grade filter. However, determining the economic optimum prefilter and final filter configuration can be challenging. Numerous prefilter options are available, the prefilter to final filter area ratio must be determined, and operating conditions must be selected that will both satisfy the filtration requirements and provide for an economical process that minimizes the filtration system footprint.
One approach towards achieving an optimal filtration system design is to test the bioprocess fluid with several filter configuration combinations and at a range of operating conditions. However, this can be a daunting task and even impractical given the high cost and limited availability of valuable bioprocess fluids. A better approach is to run a limited filtration trial and use a mathematical model that can accurately predict the behavior of the prefilter and final filter under different conditions.
In this webinar we describe a filtration model and test methodology to rapidly and efficiently design an optimal dual-stage filtration process. The model and methodology were applied to Milligard® PES filters, a new class of autoclavable and gamma sterilizable PES membrane prefilters that are designed to protect microfiltration and nanofiltration final filters in bioprocess streams. We show how a model fit to the data from one set of filtration conditions can be used to predict filtration performance at other prefilter to final filter area ratios and operating conditions, and to determine the economic optimum filtration configuration.
In this webinar, you will learn:
- How filters for microfiltration of biological fluids work.
- The effect of operating conditions on filtration performance.
- How to design an optimal series filtration (prefilter and final filter) process.
Process development guidance for AAV and lentivirus manufacturing based on co...Merck Life Sciences
Access the interactive recording here: https://bit.ly/37nl3Ex
Webinar summary:
An efficient production platform is essential for successful commercial implementation of gene therapy programs. AAV and Lentivirus manufacturing process are often developed with compressed timelines, reduced process optimization and low product yields which can have significant effect on costs.
In this webinar, you will learn:
* How manufacturing costs are examined for adeno-associated virus and lentivirus production with several different for each vector
* That key process characteristics like production titer, production of empty viral particles, downstream product recovery, and the batching strategy can effect the overall manufacturing cost
* How holistic evaluation is an important tool during process development to help prioritize different approaches to improve viral vector production processes
Abstract:
An efficient production platform is essential for successful commercial implementation of gene therapy programs. Viral vector manufacturing processes are often developed under timelines which are considerably shorter than those for more mature biopharmaceuticals. Consequently, the level of process optimization is reduced and challenges related to low product yields are common. These factors, as well as the small batch sizes common for these processes, can have significant effect on manufacturing costs.
Introduction to Tangential Flow Filtration (TFF)MilliporeSigma
This presentation provides an introduction to tangential flow filtration and reviews the following:
- TFF process basics and terminology
- TFF membrane technology
- TFF hardware, devices and systems
- Growing applications and the future
To learn more about this topic or collaborate with our technical experts, schedule an in-person or remote visit at our M Lab™ Collaboration Centers: www.emdmillipore.com/mlab
Does your cell line have a secret? Avoid surprises with characterizationMerck Life Sciences
Watch the recording of this webinar here: https://bit.ly/2Y05bV4
The first step to avoiding an unpleasant and costly contamination event is characterization of your cell banks.
Regardless of the biotech product, careful characterization of the cell banks used in its production is the first step in mitigating the risk of a contamination event. In fact, cell line characterization is an important component of the overall viral safety strategy for the product. We will describe the testing necessary to ensure cell banks are free from infectious and other adverse agents and that meets current regulatory expectations. Different levels of testing are performed for master, working, and end of production cell banks, and the differences in testing for each of these types of banks will be discussed.
In this webinar, you will learn:
• The types of tests that are needed to fully characterize your cell banks
• The best tests to use for your particular cell line
• Reasons why a viral contaminant may be missed
Find your filter. What’s best for your process? MilliporeSigma
Selecting the right aseptic filter for your process can be complicated: today’s biomanufacturer has many filter choices each offering distinct benefits. Understanding the specific needs for individual operations, in terms of flux, capacity, bioburden reduction or sterilizing performance, gamma or thermal compatibility and single or multi-use will inform decisions that have implications for the life of the process. This webinar will provide general customer guidance and explain the benefits and disadvantages of different options to help guide customers to the most appropriate filter for their operation.
In this webinar, you will learn:
- How filter design impacts performance
- Important criteria for filter selection
- New choices and options to maximize productivity for biomanufacturers
This document discusses biosafety testing for cell and gene therapies performed by BioReliance, a testing services division of Merck KGaA. It outlines the comprehensive testing performed at various stages of development, including testing of cell banks, viral vectors, and final drug products. Testing evaluates important product attributes like identity, purity, potency and residuals to ensure safety and quality. A wide range of assays are used to characterize products and identify potential contaminants.
Parvovirus Filtration Best Practices - 25 Years of Hands-On ExperienceMerck Life Sciences
In this webinar, you will learn:
- how to measure filter performance and capacity,
- how to optimize filter virus removal capability,
- and avoid potential pit-falls
Detailed description:
This webinar will cover all aspects of parvovirus filtration best practices: process development/ optimization, pilot scale-up, and validation and explain the important connections between these activities. The rationale for the recommended best practices will be explained by discussing the underlying mechanisms that control filter performance.
Filter Validation and Regulatory aspectsbala murugan
The document discusses various aspects of filter validation including:
1) Common materials used in filters include regenerated cellulose, nylon, polyamides, and polyether sulfones.
2) Porosity refers to the total empty space in a filter and affects its performance.
3) Validation tests include bacterial challenge tests, bubble point tests, and diffusion tests to evaluate a filter's effectiveness.
4) Bubble point tests determine a filter's pore size while diffusion tests measure gas flow rates through a filter.
In this webinar, you will learn:
How Single-Use helps to simplify ADC manufacturing
Safety and Integrity of the complete process
Flexibility and adaptability to changing needs
How to speed up the synthesis of high-potent payloads and improve the bioavailability of the ADC
Detailed description:
The development of an ADC is a long journey that can be speeded-up with the adoption of proper manufacturing tools and new linker-payload solutions. Single-Use equipment applied to the GMP manufacturing of ADC means operator safety, decreased risk of contamination, scalability, reproducibility, flexibility, small footprint, lower cost, and saved time through efficiency (easy setup and cleanup, no need for cleaning validation). The adoption of payload intermediates such as DolCoreTM and MayCoreTM can speed your development project up, while innovative linkers like ChetoSensarTM increase its chances of success. This webinar explains how these technologies improve the ADC production process end-to-end, ultimately enhancing patient safety.
Membrane Chromatography Solutions for Single-Use, Intensified mAb PurificationMerck Life Sciences
Participate in the interactive webinar: http://bit.ly/NatrixChromMRK
Improve productivity, flexibility, and economics of mAb purification process with intensified, single-use membrane chromatography.
Explore our webinar library: www.merckmillipore.com/webinars
Complete single-use ADC technology from development through scale-up MilliporeSigma
This webinar will talk about the benefits of single-use technologies for the manufacturing of antibody-drug conjugates and present a successful corresponding case study.
With an expected high annual growth rate of the global Antibody-drug Conjugate (ADC) market, it is essential that CMO’s have robust manufacturing platforms to ensure successful transfer to GMP production.
Single-Use Technologies provide many advantages, including improved safety, lower costs and greater flexibility. This webinar will outline the advantages of a Single Use Platform and give a case study on how it can be used to manufacture ADC projects.
In this webinar, you will learn:
● How single-use technologies can provide benefits for ADC manufacturing
● Why a solid manufacturing platform is crucial for a successful transfer to GMP production
● How a case study demonstrates the advantages of single-use equipment in a scale up to GMP production
How to reach High Plasma Protein Concentration with Single-Pass TFFMilliporeSigma
This webinar will discuss our collaboration with Takeda on the development of a single-pass TFF system as an alternative to traditional TFF for concentrating a plasma-derived IgG solution.
Single-Pass Tangential Flow Filtration (SPTFF) is a technology that requires only one pass through the filter assembly to achieve the desired concentration with no recirculation of product.
SPTFF can offer many advantages in downstream processing, such as:
• Increased capacity and reduced process time
• Increased yield and product recovery
• Optimized processing of highly shear-sensitive products
• Reduced foam formation
• Reduced cost of goods
This presentation will cover our collaboration with Takeda, formerly Shire, for the development of a specific SPTFF system as an alternative to traditional TFF for concentrating a plasma-derived Immunoglobulin G (IgG) solution from 10% to 20%. Due to promising results, plans are underway to replace the currently used batch TFF process with a SPTFF step.
In this webinar, we will discuss:
- A comparison of traditional TFF versus SPTFF
- Design of Experiments (DOE) approach toward initial process development work and determination of the optimal parameters
- Process run results, including final product yield and product quality
Keeping the (Adventitious) Virus Out of the (Adeno-Associated) VirusMilliporeSigma
This document discusses approaches for viral safety in AAV processes. It begins with background on the growing gene therapy market and evolving regulatory landscape. It then outlines a risk mitigation strategy using a tripod approach of prevent, remove, and detect. Specific techniques discussed for prevent include using virus-resistant cell lines and pretreated raw materials. Techniques discussed for remove include virus filtration, chromatography, and the use of Benzonase endonuclease to digest DNA. The document emphasizes a layered, multi-pronged approach for complete biosafety assurance in AAV processes.
Selection, sizing, and operation of bioprocess filtration trains for optimal ...MilliporeSigma
This document discusses the selection, sizing, and operation of bioprocess filtration trains for optimal performance. It covers background on membrane types used in biopharma processing including ultrafiltration, virus retentive, and microfiltration membranes. It then discusses filtration mechanisms, series filtration, and modeling approaches to predict filter performance and sizing including classical fouling models and combined blocking and adsorption models. Experimental data is presented to validate the predictive capability of the combined blocking and adsorption fouling model for sizing filters in a series configuration.
A Turn-Key Flow-Through-Mode Purification Process to improve Quality and Safe...MilliporeSigma
In this webinar, you will learn:
Intensified plasma Immunoglobulin purifications
Scalable process development with latest technologies
Improved safety and quality of plasma IgG meeting required quality attributes
Detailed description:
Plasma-derived immunoglobulins (IgG) are essential medicines that are in worldwide shortage. How to develop optimized processing steps for robust and efficient manufacturing has been a constant goal, to make the most out of the precious plasma raw material.
In this study, we present a worse-case equivalent of plasma intermediate, explore various process steps along the fractionation flow, including flow-through-mode chromatography, affinity chromatography, virus inactivation steps and removal of solvent/detergent, single-pass TFF (SPTFF), clarification, and aseptic filtration, to establish a robust, easy-to-operate, readily scalable plasma IgG process with over 99% purity, depletion of IgA, isoagglutinin, and thrombogenic markers, meeting the commonly required 20% concentration for subcutaneous IgG infusion. Such solutions would be appropriate for various IgG intermediates which help to improve the global supply of immunoglobulins.
This presentation reviews current trends in bioprocessing purification and includes key considerations for continuous processing and connected polishing for monoclonal antibodies. Topics include:
• Market trends and the evolution of next-generation processes
• Intensified capture processing
• Continuous virus inactivation
• Connected flow-through polishing
To learn more about this topic or collaborate with our technical experts, schedule an in-person or remote visit at our M Lab™ Collaboration Centers: www.merckmillipore.com/mlab
Technology Trends in Bioprocessing PurificationMilliporeSigma
This presentation reviews current trends in bioprocessing purification and includes key considerations for continuous processing and connected polishing for monoclonal antibodies. Topics include:
• Market trends and the evolution of next-generation processes
• Intensified capture processing
• Continuous virus inactivation
• Connected flow-through polishing
To learn more about this topic or collaborate with our technical experts, schedule an in-person or remote visit at our M Lab™ Collaboration Centers: www.emdmillipore.com/mlab
Validation of anti niv igm capture elisa version#1krishgen
NiV is a negative-sense, non-segmented RNA virus that was first isolated from cerebrospinal fluid of human patients and classified in the family Paramyxoviridae under the new genus
Henipavirus. Its genome encodes six structural proteins: the nucleocapsid (N) protein,
phosphoprotein (P), matrix (M) protein, fusion (F) protein, glycoprotein (G), and large (L)
protein.
Nipah virus glycoprotein G has a globular head domain formed of a six-bladed beta sheet propeller, connected via a flexible stalk domain to a transmembrane anchor. The G binds to the cellular receptors ephrin B2 are ephrin B3, mediating viral attachment. Following attachment Nipah Virus glycoprotein G undergoes a conformational change that leads to triggering of glycoprotein F which leads to membrane fusion (Biering et al, 2012).
The Nipah virus glycoprotein G is a recombinant protein expressed in mammalian HEK293 cells. It is presented as a fusion protein with a mouse Fc tag linked to the C-terminus of glycoprotein G, amino acids 71-602.
We established preliminary specifications defining acceptable ranges for the parameters indicated herein below for our Anti Nipah Virus IgM Capture ELISA kit. These parameters were tracked day-to-day, run-to-run, and operator-to-operator, over a schedule defined inhouse.
Recommended assay characteristics included absorbance of a zero concentration standard; factors which describe the calibration for each standard and statistical description of the calibration curve such as coefficient of correlation, slope and/or intercept; and recovery of results on control samples. It is important to be able to relate the specifications for a parameter to expected reliability of the result. Our in-house standard defined was r=0.990.
The present article deals with design of antibacterial and antifungal pH-responsive hydrogels based on Quaternary Ammonium Functionalized-Tragacanth Gum (QTG) biopolymer as drug delivery systems. The antimicrobial effects of the graft-copolymer hydrogels QTG/ polyacrylic acid (QTG-AA) and QTG/polyacrylamide (QTG-AM) were investigated against fi ve standard microorganisms including Candida albicans, Escherichia coli, Bacillus subtilis, Staphylococcus aureus, and Pseudomonas aeruginosa. The results of the in-vitro release of quercetin as a drug from the functionalized copolymer hydrogels exhibited dependence on the pH, immersion time, medium, and temperature. All copolymer hydrogels demonstrated antibacterial and antifungal properties and moreover, QTG-AM copolymers presented higher antimicrobial activity than QTG-AA copolymers.
Webinar: Evaluating Viral Clearance for Continuous ProcessesMerck Life Sciences
Participate in the interactive webinar now: http://bit.ly/ViralClearanceWebinar
Is viral clearance a hurdle to implementing continuous processing? We’ll share virus spiking alternatives that may pave the way for effectively evaluating viral clearance by chromatography steps in a continuous process.
Explore our webinar library: www.merckmillipore.com/webinars
Webinar: Evaluating Viral Clearance for Continuous ProcessesMilliporeSigma
Participate in the interactive webinar now: http://bit.ly/ViralClearanceWebinar
Is viral clearance a hurdle to implementing continuous processing? We’ll share virus spiking alternatives that may pave the way for effectively evaluating viral clearance by chromatography steps in a continuous process.
Explore our webinar library: www.emdmillipore.com/webinars
Optimization of Corynebacterium glutamicum Immobilization on Alginate and In...IJMER
The parameters of the immobilized process of Corynebacterium glutamicum VTCC – B – 0632
on alginate were identified by Plackett-Burman matrix, and the experiments were designed by response
surface methodology having the central composite designs (RSM-CCD). The maximum yield of cell
immobilization on alginate carrier reached at 92.6%. Optimal parameters were the cell density of 89.3
million cells/mL in the 4% sterile alginate with ratio 1:1. This mixture went through the syringe system of
the 2M CaCl2 solution at 200C with the shaking speed of 75 rpm until the gels get in shape. Then, these
gels were soaked in the CaCl2 liquor and shaken for 41 minutes (150 rpm). At last, the particle size of
final products was 4mm and the average cell density was 14.75 million cells/gram. This immobile product
is maintained under the suitable condition in the CaCl2 liquor (w/v), pH=7. The cell survival percentage
after 72 hours were 98% when it was stored in 4
0C, 0.5% CaCl2 and pH of 7
High-performance anion-exchange chromatography with pulsed amperometric detection is valuable for oligosaccharide analysis with the value derived from the high-resolution separation followed by sensitive detection of native oligosaccharides. In this presentation the application of HPAE-PAD to oligosaccharides released from glycoproteins is demonstrated.
PCUBE--Protein production platform for mAb generation. Part IIIcarlociatto
An efficient three-step process can purify IgM monoclonal antibodies. The first step uses anion exchange chromatography at pH 8.0 to capture the IgM. Alternatively, PEG precipitation followed by ceramic hydroxyapatite chromatography can be used. The second step uses size exclusion chromatography to further purify. The final step employs cation exchange chromatography at pH 6.0 to polish the purified IgM monoclonal antibody product.
1) The document describes the preparation and certification of a curry paste reference material containing benzoic acid for use in food testing laboratories.
2) Tests were conducted to evaluate homogeneity, stability, and characterize the material. The material demonstrated homogeneity and stability when stored at -22C for 2 years.
3) Based on the test results, the certified value for benzoic acid in the reference material is 378 mg/kg with an expanded uncertainty of 15.6 mg/kg at a 95% confidence level.
A Novel Approach to Internal Standardization in LC/MS/MS Analysis; Sensitive ...MicroConstants
This presentation was prepared by Bruce Babson, Research Fellow at MicroConstants, Inc. in San Diego, California, for the CACO-PBS Mini-Symposium on Bioanalytical and Analytical Applications and Problem Investigation Case Studies. Bruce is one of twelve presenters at the August 10, 2012 event in Foster City, California.
Fast-track solutions to address challenges with Host Cell Proteins in early d...Merck Life Sciences
Watch this webinar here: https://bit.ly/3fFRXDb
This webinar illustrates a customer case study about the challenges related to the removal of Host Cell Proteins from bioreactor harvest, the selected fast-track approach and outcome.
Our customer had one month to reduce the level of HCPs in the bioreactor harvest prior to a production run to supply drug substance for a scheduled Phase 1 clinical trial. The high level of HCPs (1,000,000 in the harvest and 700 ppm at the end of purification) unfavorably impacted the planned clarification process and subsequent downstream steps.
The goal was to reduce the level of HCPs to maximum of 300 ppm at the end of process purification and ensure clarification of the entire 2000L harvest.
In this webinar, you will learn about:
- Challenges with Host Cell Proteins
- Fast-Track Approach using caprylic acid precipitation followed by filtration using Clarisolve® filters
- The impact of Design of Experiment
Fast-track solutions to address challenges with Host Cell Proteins in early d...MilliporeSigma
This document discusses a case study where caprylic acid precipitation and depth filtration were used to address high levels of host cell proteins (HCPs) in a biotherapeutics process. A design of experiments approach was used to optimize the caprylic acid concentration and temperature for precipitation. Depth filtration with a 40MS filter was shown to improve recovery while significantly reducing HCP levels. The modified process was successfully implemented at 2000L scale within one month to meet HCP specifications for an upcoming clinical trial.
This document summarizes a research article that studied the transport of the antibiotic tetracycline (Tet) in Escherichia coli. The key findings are:
1. Tet is transported into E. coli cells by three transporters: TetA, TetB, and TetH.
2. TetA is a primary transporter that uses ATP hydrolysis to actively transport Tet into cells. TetB and TetH are secondary transporters that harness gradients to transport Tet.
3. Experiments showed TetA is the major transporter and is responsible for most Tet uptake. TetB and TetH play lesser but still significant roles in Tet transport.
4. Understanding the roles and interactions of these
Quality Criteria Establishment for Dissolution of Ascorbic Acid from Sustaine...CrimsonpublishersNTNF
Quality Criteria Establishment for Dissolution of Ascorbic Acid from Sustained Release Pellets by Mostafa Essam Ahmed Mostafa Eissa in Nutrition and food science open access journal
Protein Purification and Analysis with PhyNexus and Caliper LifeSciencesChris Suh
PhyTip columns perform high throughput protein separation on Caliper liquid handling robotics and proteins analyzed by Caliper LabChip GXII microfluidic device
Application of Validated High-performance Liquid Chromatography Method for De...BRNSS Publication Hub
A novel and simple reversed-phase liquid chromatographic method has been established for the determination of saxagliptin and metformin HCl Saxagliptin and metformin HCl is used to control Type 2 diabetes. The proposed work was performed on Young Lin (S.K) isocratic System UV Detector. Saxagliptin and metformin HCl is used to control Type 2 diabetes. The proposed work was performed on Young Lin (S.K) isocratic System UV Detector C18 column (150 mm × 4.6 mm). A mixture of potassium phosphate, mobile phase in this method with flow rate of 0.7 mL/min (UV detection at 203 nm) and the method was validated as per the ICH guidelines. Forced degradation studies were performed by exposing the drug saxagliptin and metformin HCl to acidic, alkaline, oxidation, and thermal stress degradations. The proposed reversed-phase-high-performance liquid chromatography method was found to be robust and specific, and this method is suitable for the assay of pharmaceutical dosage forms as well as kinetic studies.
Similar to Chromatography: Chromatographic strategies for IVIG purification – Part 2 (20)
The Viscosity Reduction Platform: Viscosity-reducing excipients for improveme...Merck Life Sciences
Protein viscosity is a major challenge in preparing highly concentrated protein formulations suitable for subcutaneous injection. Recently, the Viscosity Reduction Platform (VRP) was introduced and its technical key features and benefits for formulations were discussed. However, highly viscous solutions do not only pose a challenge when administering a drug to a patient, they can also impose technical limitations in the manufacturing process.
This white paper evaluates the effect of the excipients in the Viscosity Reduction Platform on ultrafiltration processes used to produce a highly concentrated formulation of a monoclonal antibody (mAb). Two filtration methods are demonstrated in this work.
Find more information about the Viscosity Reduction Platform on our website: https://www.sigmaaldrich.com/products/pharma-and-biopharma-manufacturing/formulation/viscosity-reduction-platform
Use of Excipients in Downstream Processing to Improve Protein PurificationMerck Life Sciences
Excipients are used to improve the stability of protein-based therapeutics by protecting the protein against a range of stress conditions such as temperature changes, pH changes, or agitation. Similar stresses are applied to proteins during downstream purification. Shifts in pH during Protein A chromatography, subsequent incubations at low pH for virus inactivation, and changes in conductivity in ion exchange chromatography can lead to aggregation, fragmentation, or other chemical modifications of the therapeutic protein. Given the potential impact on the protein’s structural integrity, there is a need for approaches to reduce the risk presented by the conditions during downstream processing. For example, integration of a solution to prevent aggregation of proteins would be a more efficient strategy than implementing steps to remove multimeric forms.
This white paper highlights the results from a recent paper by Stange et. al., in which protein stabilizing excipients such as polyols, sugars, and polyethylene glycol (PEG4000) were used as buffer system additives. Effect of the excipients on elution patterns, stabilization of the monomer antibody, host-cell protein removal, virus inactivation rates and binding capacity of cation exchange chromatography were explored.
Exploring the protein stabilizing capability of surfactants against agitation...Merck Life Sciences
Agitation of therapeutic protein solutions during manufacturing, shipping and handling is one of the major initiators for protein aggregation and particle formation during the life history of a protein drug. Adsorption of protein molecules to liquid-air interfaces leads to the formation of highly concentrated protein surface films. The rupture of these protein films due to various mechanical processes can then result in the appearance of protein aggregates and particles in the bulk solution phase.
One technique to stabilize proteins against stress induced by liquid-air interfaces is the use of non-ionic surfactants. About 91% of antibody formulations commercially available in 2021 contained a surfactant. Polysorbate 20 and 80, composed of a hydrophilic polyoxyethylene sorbitan and hydrophobic fatty acid esters, made up the largest part being employed in 87% of said formulations.
Despite their frequent use in parenteral drug products, concerns have been raised for decades about the application of polysorbates as surfactants in biopharmaceutical formulations. Autoxidation of polysorbate, caused by residual peroxides in polysorbates, can damage the proteins and can further drive the oxidative degradation of polysorbate. Chemical and enzymatic hydrolysis of polysorbate may lead to the formation of free fatty acid particles, which may become visible; and both mechanisms eventually lead to the reduction in polysorbate concentration. Therefore, the purpose of the current study was to compare various molecules for their capabilities to reduced agitation-induced protein aggregation and particle formation; and furthermore, investigate their underlying protein stabilizing mechanisms.
The Viscosity Reduction Platform: Viscosity Reducing Excipients for Protein F...Merck Life Sciences
Protein viscosity is one of the major obstacles in preparing highly concentrated protein formulations suitable for subcutaneous injection.
This whitepaper examines how combining an amino acid with a second viscosity-reducing excipient circumvents adverse effects on protein stability and improves viscosity-reducing capacity.
To find more information about the Viscosity Reduction Platform, please visit our website: https://sigmaaldrich.com/products/pharma-and-biopharma-manufacturing/formulation/viscosity-reduction-platform
Characterization of monoclonal antibodies and Antibody drug conjugates by Sur...Merck Life Sciences
Watch the presentation of this webinar: https://bit.ly/3Pjpjvr
Highlights of this webinar:
- Surface plasmon resonance as a powerful tool for biologic characterization including mAbs and ADCs.
- SPR allows rapid binding analysis in real time without using labels for SARS-CoV-2 receptor binding domain mutations.
- Kinetic data is indicative of possible neutralizing activity allowed assessment of neutralizing ability of therapeutic monoclonal antibodies.
- The application can provide preliminarily efficacy information and facilitated mAbs/ACDs candidate selection process
Detailed description:
Characterization of therapeutic monoclonal antibodies (mAbs) or Antibody drug conjugates (ADCs) is challenging due to their ability to bind to a variety of proteins via their Fc and Fab domains, giving rise to diverse biological functions associated with each domain. The Fc domain of mAbs interacts with Fc receptors with varying affinities, which can influence biological processes such as Complement-dependent cytotoxicity (CDC) and Antibody-dependent cellular cytotoxicity (ADCC), transcytosis, phagocytosis, and/or serum half-life.
An important characteristic of an antibody is its Fc effector function. Antibodies can be engineered to obtain desired binding of the Fc region to Fc receptors expressed on effector cells. Hence, it is crucial to evaluate the binding interaction of mAbs/ADC with Fc receptors in the early phase of drug development to understand the potential biological activity of the product in vivo.
Surface Plasmon Resonance (SPR) is a powerful technique to establish binding kinetics in real-time, label free, and high sensitivity with low sample consumption. Along with target antigen binding, it is crucial to evaluate the binding interaction of antibodies and ADCs with Fc receptors. Our SPR case studies investigated the impact on binding kinetics of ADCs with different linkers and the binding interactions of SARS-CoV-2 spike protein variants and evaluated the neutralizing ability of therapeutic mAbs. SPR characterisation can be facilitated in all stages of the product life cycle to ensure the quality and safety of mAbs and ADCs.
The Role of BioPhorum Extractables Data in the Effective Adoption of Single-U...Merck Life Sciences
Regulatory expectation does require patient safety evaluations with supporting data for manufacturing components that directly come into contact with drug manufacturing process streams. Readily available extractables data can help manufacturers using singleuse technology to accelerate product qualifications, risk assessments and process optimization
This white paper guides you on how to save time and resources with supplier-provided single-use system extractables data and gives you an overview about the overall strategy for Extractables & Leachables. At the end you will find a case study.
Find more information about filters and single-use components on our website: https://www.sigmaaldrich.com/DE/en/services/product-services/emprove-program/emprove-filter-and-single-use-component-portfolio
Watch the recording of this presentation here: https://bit.ly/3zTOpe4
Detailed description:
SARS-CoV-2 showed us that technology supports us during our inspection activity even if on-site visits are not possible. Travel restrictions of various kinds will remain a risk in the future. The use of new technologies has shown that inspections and audits can be carried out despite these restrictions. We will focus on what possibilities the new technologies offer and take a look at the future of inspections and audits.
In this webinar, you will learn:
• Regulatory overview of remote audits
• The technologies needed to support the audit process
• What types of inspections are possible with the use of these technologies
• How audits may look in the future
Presented by:
Daniel Buescher, Product Manager - Digital Solutions
Moving your Gene Therapy from R&D to IND: How to navigate the Regulatory Land...Merck Life Sciences
Watch the recording of this presentation here: https://bit.ly/3SqOsoP
Novel therapies, including cell and gene therapies, continue to be central to innovation in healthcare and represent the fastest growing area of therapeutic medicine. As a consequence, the number of gene therapies undergoing clinical trials has increased significantly in the last five years.
Manufacturing processes for these novel therapeutics are very complex with a high risk of contamination. Regulatory agencies world-wide have responded by issuing guidance to outline their expectations for development and manufacture of cell and gene therapies. Currently, regulatory guidance is not harmonized globally and can often lead to confusion within industry and increased risk of non-compliance.
In this webinar, we'll answer:
• Which regulatory guidelines do you need to comply for your INDs?
• When do you start implementing GMPs and validated assays?
• How do you get your QC testing strategy ‘right the first time’?
• How do you ensure testing is not your rate limiting step for the IND submission?
Presented by:
Manjula Aysola, Senior Regulatory Consultant
Dr. Alison Armstrong, Sr. Director, Technical and Scientific Solutions
Identity testing by NGS as a means of risk mitigation for viral gene therapiesMerck Life Sciences
Watch the presentation of this webinar here: https://bit.ly/3RijkHC
Detailed description:
Imagine you’ve just completed a manufacturing run for your viral vector. Identity testing is performed to confirm the vector sequence. But when the results come back the data reveals unexpected sequence variants! With an appropriate risk mitigation testing strategy, this situation can be prevented.
The situation described above is not hypothetical, and happens more that you think, costing valuable time and resources.
Investigatory testing has shown that sequence variants present in starting materials (e.g. plasmids) are likely to make their way to the final product. Adequate identification of low-level variants with an appropriately sensitive method is critical in ensuring the quality of the final product. A risk-based testing strategy, in the context of identity, for viral vector manufacturing will be presented, focusing on key testing points. NGS assays for identity and variant detection will be highlighted due to their extremely sensitive nature compared to traditional approaches.
In this webinar, we'll explore:
• Regulatory requirements for identity testing
• NGS applications for identity testing as compared to traditional methods
• A case study on the impact of not establishing a proper risk-based testing strategy
Presented by: Bradley Hasson, Director of Lab Operations for NGS Services
Latest advancements of melt based 3D printing technologies for oral drug deli...Merck Life Sciences
Watch the presentation of this webinar here: https://bit.ly/3A2WcH4
The application of polymer excipients in 3D printing manufacturing is usually limited due to the concerns of filament strength, high processing temperature and large scale manufacturing.
Latest technology developments are targeting a direct melt deposition to simplify the process and enable a constant and efficient process. Two different processing approaches will be presented:
The advanced melt drop deposition, where individual three dimensional geometries can be created by depostition of polymer droplets and the MED® 3D printing technology which allows by precise layer-by-layer deposition to produce objects with well-designed geometric structures.
In this webinar, you will learn:
• Latest advancements of melt based 3D printing approaches
• Application examples for the individual technologies
• Deep dive in the MED® 3D printing technology to design dedicated drug release profiles
Presented by:
Dr. Thomas Kipping, Head of Drug Carriers
Dr. Xianghao Zuo, Deputy Director of R&D, Triastek
CAR-T Manufacturing Innovations that Work - Automating Low Volume Processes a...Merck Life Sciences
Watch the presentation of this webinar here: https://bit.ly/3NDNIKe
Automated, fit-for-purpose tools are essential in CAR-T processing to support sustainable manufacturing of clinical and market-approved cell therapy products. This webinar will discuss how the ekko™ Acoustic Cell Processing System uses acoustic technology as a touchless approach to manipulate cells, enabling a modular tool across the CAR-T manufacturing workflow. Typical performance of templated ekko™ System processes for DMSO washout of leukapheresis material, low volume and high cell concentrate for electroporation preparation, and harvest of expanded T cells will be reviewed.
This webinar will also give an early glimpse at the ekko™ Select System for unmatched T cell selection.
In this webinar, you will:
• Uncover how the ekko™ System supports the broad industrialization of cell therapy, with particular focus on how to achieve low volume, high concentrate cell product for critical transduction and transfection steps
• Discover how ekko™ System for wash and concentrate processes throughout the cell therapy workflow achieve high cell recovery, viability, and effective residual removal
• Preview to ekko™ Select, our cell therapy selection platform, to achieve unmatched ease-of-use with direct processing from leukopaks reducing the need for preparation steps
Presented by:
Benjamin Ross-Johnsrud, Acoustic Technology Expert
Robert Scott, Mechanical Engineer III
Viral safety of biologics: What's changing with the ICH Q5A revision?Merck Life Sciences
Watch the presentation of this webinar here: https://bit.ly/3t7X9tg
How does the ICH Q5A revision impact viral safety strategies for biologics?
Biologics continue to grow at a fast pace. Manufactured using cell lines of human or animal origin, these are at risk of viral contamination making safety strategies critical. A comprehensive risk mitigation strategy using multiple orthogonal measures is a regulatory expectation. ICH Q5A, the globally-harmonized guideline outlines the expectations. ICH Q5A is currently being revised to address recent scientific advancements including novel therapeutic modalities, new manufacturing paradigms, updates in viral clearance applications, and alternate detection technologies. We’ll discuss the expected changes and potential impact on viral safety strategies with case studies and examples.
In this webinar, you will learn about:
• The Importance of virus testing in biologics products
• Regulatory landscape, expectations for the Q5A revision
• What's new and changing
• Examples of alternate testing schedules, impact on viral clearance
Presented by:
Manjula Aysola, Senior Regulatory Consultant
Alison Armstrong, PhD, Sr. Director, Technical and Scientific Solutions
Improve Operational Efficiency by Over 30% with Product, Process, & Systems A...Merck Life Sciences
Watch the presentation of this webinar here: https://bit.ly/3adaxWh
When implementing new automation systems, organizations must consider things like deployment time, user adoption, and costs.
They must also consider the cost of doing nothing – that is, what competitive advantage is lost in standing still? What time and quality is lost in repetitive, manual tasks rather than an automated, digital workflow? What operational efficiencies are lost?
In this webinar we examine how a product, process, and system agnostic automation platform can be deployed faster than traditional system specific software while bringing greater operational efficiencies (in many cases over 30% improvement).
To remain competitive in the market, biopharma manufacturers must adopt automation and digital technologies, but most plants still have island of automation consisting of independently functioning, standalone unit operations. This results in operational inefficiency, regulatory concerns, and a poor understanding of the process and product life cycle.
Taking the first, right step must include considering risks, costs, timelines, and technology alternatives. Traditional automation approaches tied to specific systems, processes, and products are, by their nature, limited; while an agnostic platform will address current biomanufacturing business challenges and ensure future readiness. With the right platform, a phased automation implementation can yield operational efficiency gains of up to 30% and improved product quality and regulatory compliance.
In this webinar, let's explore:
• Challenges of automation and digital technology adoption
• What a product, process, and system agnostic platform entails
• Applications and benefits of a process orchestration platform
• Ensuring future readiness with process orchestration
Presented by:
Braj Nandan Thakur, Global Product Manager - Automation
Insights from a Global Collaboration Accelerating Vaccine Development with an...Merck Life Sciences
Watch the presentation of this webinar here: https://bit.ly/3Nbb5ug
Get insights and best practices from a multinational team establishing a platform for vaccine production. See how a long-term collaboration on a bench-scale process used to produce a Virus Like Particle (VLP) vaccine for SARS-CoV-2 was successfully converted to a robust GMP-compatible, scalable process.
The COVID-19 pandemic further emphasized the need for collaboration in the development of urgently needed vaccines and therapeutics. In this webinar, we take you behind the scenes of our collaboration with Technovax and Innovative Biotech in which a scalable VLP vaccine platform was optimized for use in a production facility in Nigeria in response to the need for local production of SARS-CoV-2 vaccines. The flexibility and robustness of the platform will enable its rapid deployment to support the West African pandemic readiness program. Initial development of the VLP process began in late 2019 and by March 2020, was already adapted for production of a SARS-CoV-2 vaccine.
In this webinar, you will learn:
• About building a priceless collaborative network with integrated solutions
• Virus-Like Particle Vaccines
• Process Development Overview and Challenges
• Pre-clinical Results and Next Steps
Presented by:
Jose M. Galarza, PhD,
President and Founder of TechnoVax
Naomi Baer,
Business development consultant, Emerging Biotech, BioProcess division
Youssef Gaabouri, Eng. ,
Associate Director, Head of Sales Middle East & Africa, BioProcess division
Risk-Based Qualification of X-Ray Sterilization for Single-Use SystemsMerck Life Sciences
The document discusses testing done to qualify the use of x-ray sterilization for a Lynx S2S connector. Physical, chemical, and biological tests were performed on connectors that underwent either x-ray or gamma sterilization. Test results showed comparable extractable levels, thermal properties, and chromatographic profiles between the two sterilization methods. This provides evidence that x-ray sterilization is a suitable alternative to gamma sterilization for this connector.
The High Intensity Sweeteners Neotame and Sucralose: 2 Ways to ace the Patien...Merck Life Sciences
Watch the presentation of this webinar here: https://bit.ly/3vQyN7K
Bitter medicines are an important issue, especially for pediatric applications. As several APIs have bitter tasting components, high intensity sweeteners for taste optimization are of great interest. Join our webinar to discover our new sweetener toolbox enabling safe and stable formulations.
Mask bitter aftertaste for a sweeter pill to swallow! Patients’ compliance and the therapeutic benefit are supported by a pleasant taste of pharmaceutical formulations. With the high intensity sweeteners Neotame and Sucralose, you have efficient tools at hand which are superior to other sweeteners in many aspects:
• excellent sugar-like taste profile
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We will present our new toolbox of two high performance sweeteners and focus on aspects of stability, safety, the application in various dosage forms, and market perception.
In this webinar, you will learn:
• How to optimize the patients' taste experience of your pharmaceuticals
• How sweeteners can be differentiated by their sensory profiles and features
• How our new product offering Neotame can be effectively used in your targeted formulations
Presented by:
Almut von der Brelie,
Senior Manager Strategic Marketing
Excipients for Solid Applications
The Developability Classification System (DCS): Enabling an Optimized Approac...Merck Life Sciences
This whitepaper by Dr. Daniel Joseph Price outlines how poorly soluble drug formulations can be designed using the developability classification system (DCS).
The DCS identifies the root cause of low solubility and enables lean, cost-effective and effective formulations to be developed.
#solubility #pharmaceuticalmanufacturing #oralsoliddosage #drugdevelopment
The webinar discusses services from MilliporeSigma to accelerate antibody-drug conjugate (ADC) development through their ADC Express and ADCore product lines. ADC Express provides integrated antibody, linker, payload, and conjugation services to generate multiple ADC candidates for evaluation. The ADCore product line offers intermediates that simplify payload synthesis and accelerate development timelines. ChetoSensar technology incorporates a chito-oligosaccharide to enhance ADC solubility and efficacy.
Regulatory Considerations for Excipients used in Lipid NanoparticlesMerck Life Sciences
Lipid excipients and delivery systems such as lipid nanoparticles (LNPs) are essential for a wide variety of therapeutics including mRNA vaccines and therapeutics and gene therapy.
The purity and safety of novel, synthetic lipid excipients must be demonstrated due to their central role in the function of the drug product, distinct physicochemical properties, and the potential for interaction with other ingredients or the physicochemical environment. These excipients must comply with challenging and complex regulatory requirements, similar to those expected of the active pharmaceutical ingredient itself.
This whitepaper provides an overview of the regulatory classification of lipid nanoparticles, liposomes and novel excipients. Specific requirements outlined in guidance documents are shared along with strategies to stay ahead of emerging regulatory challenges.
To find more information about synthetic lipids for pharmaceutical applications and gene therapy, please visit our website:
https://www.sigmaaldrich.com/DE/en/products/pharma-and-biopharma-manufacturing/formulation/synthetic-lipids
https://www.sigmaaldrich.com/US/en/products/pharma-and-biopharma-manufacturing/formulation/synthetic-lipids
R3 Stem Cell Therapy: A New Hope for Women with Ovarian FailureR3 Stem Cell
Discover the groundbreaking advancements in stem cell therapy by R3 Stem Cell, offering new hope for women with ovarian failure. This innovative treatment aims to restore ovarian function, improve fertility, and enhance overall well-being, revolutionizing reproductive health for women worldwide.
At Apollo Hospital, Lucknow, U.P., we provide specialized care for children experiencing dehydration and other symptoms. We also offer NICU & PICU Ambulance Facility Services. Consult our expert today for the best pediatric emergency care.
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English Drug and Alcohol Commissioners June 2024.pptxMatSouthwell1
Presentation made by Mat Southwell to the Harm Reduction Working Group of the English Drug and Alcohol Commissioners. Discuss stimulants, OAMT, NSP coverage and community-led approach to DCRs. Focussing on active drug user perspectives and interests
NURSING MANAGEMENT OF PATIENT WITH EMPHYSEMA .PPTblessyjannu21
Prepared by Prof. BLESSY THOMAS, VICE PRINCIPAL, FNCON, SPN.
Emphysema is a disease condition of respiratory system.
Emphysema is an abnormal permanent enlargement of the air spaces distal to terminal bronchioles, accompanied by destruction of their walls and without obvious fibrosis.
Emphysema of lung is defined as hyper inflation of the lung ais spaces due to obstruction of non respiratory bronchioles as due to loss of elasticity of alveoli.
It is a type of chronic obstructive
pulmonary disease.
It is a progressive disease of lungs.
The best massage spa Ajman is Chandrima Spa Ajman, which was founded in 2023 and is exclusively for men 24 hours a day. As of right now, our parent firm has been providing massage services to over 50,000+ clients in Ajman for the past 10 years. It has about 8+ branches. This demonstrates that Chandrima Spa Ajman is among the most reasonably priced spas in Ajman and the ideal place to unwind and rejuvenate. We provide a wide range of Spa massage treatments, including Indian, Pakistani, Kerala, Malayali, and body-to-body massages. Numerous massage techniques are available, including deep tissue, Swedish, Thai, Russian, and hot stone massages. Our massage therapists produce genuinely unique treatments that generate a revitalized sense of inner serenely by fusing modern techniques, the cleanest natural substances, and traditional holistic therapists.
COLOUR CODING IN THE PERIOPERATIVE NURSING PRACTICE.SamboGlo
COLOUR CODING IN THE PERIOPERATIVE ENVIRONMENT HAS COME TO STAY ,SOME SENCE OF HUMOUR WILL BE APPRECIATED AT THE RIGHT TIME BY THE PATIENT AND OTHER SURGICAL TEAM MEMBERS.
CHAPTER 1 SEMESTER V COMMUNICATION TECHNIQUES FOR CHILDREN.pdfSachin Sharma
Here are some key objectives of communication with children:
Build Trust and Security:
Establish a safe and supportive environment where children feel comfortable expressing themselves.
Encourage Expression:
Enable children to articulate their thoughts, feelings, and experiences.
Promote Emotional Understanding:
Help children identify and understand their own emotions and the emotions of others.
Enhance Listening Skills:
Develop children’s ability to listen attentively and respond appropriately.
Foster Positive Relationships:
Strengthen the bond between children and caregivers, peers, and other adults.
Support Learning and Development:
Aid cognitive and language development through engaging and meaningful conversations.
Teach Social Skills:
Encourage polite, respectful, and empathetic interactions with others.
Resolve Conflicts:
Provide tools and guidance for children to handle disagreements constructively.
Encourage Independence:
Support children in making decisions and solving problems on their own.
Provide Reassurance and Comfort:
Offer comfort and understanding during times of distress or uncertainty.
Reinforce Positive Behavior:
Acknowledge and encourage positive actions and behaviors.
Guide and Educate:
Offer clear instructions and explanations to help children understand expectations and learn new concepts.
By focusing on these objectives, communication with children can be both effective and nurturing, supporting their overall growth and well-being.
TEST BANK FOR Health Assessment in Nursing 7th Edition by Weber Chapters 1 - ...rightmanforbloodline
TEST BANK FOR Health Assessment in Nursing 7th Edition by Weber Chapters 1 - 34.
TEST BANK FOR Health Assessment in Nursing 7th Edition by Weber Chapters 1 - 34.
TEST BANK FOR Health Assessment in Nursing 7th Edition by Weber Chapters 1 - 34.
This particular slides consist of- what is Pneumothorax,what are it's causes and it's effect on body, risk factors, symptoms,complications, diagnosis and role of physiotherapy in it.
This slide is very helpful for physiotherapy students and also for other medical and healthcare students.
Here is a summary of Pneumothorax:
Pneumothorax, also known as a collapsed lung, is a condition that occurs when air leaks into the space between the lung and chest wall. This air buildup puts pressure on the lung, preventing it from expanding fully when you breathe. A pneumothorax can cause a complete or partial collapse of the lung.
Chromatography: Chromatographic strategies for IVIG purification – Part 2
1. Merck KGaA
Darmstadt, Germany
Chromatographic
strategies for IVIG
purification
Thierry Burnouf PhD.
Vice-Dean, College of Biomedical Engineering, Taipei Medical
University
Josephine Cheng
Associate Director Plasma Segment APAC, Process Solutions,
Life Science
– Part 2
2. The life science business of
Merck KGaA, Darmstadt, Germany
operates as MilliporeSigma
in the U.S. and Canada.
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.182
4. Immunoglobulin G (IgG,
intravenous and
subcutaneous)
Important plasma-derived product
used to treat patients with various
immunologic, neurologic, and
hematologic conditions, e.g. PIDD,
ITP, CIDP, leukemia, Kawasaki
Syndrome
Market growth driven by increasing
use of immunoglobulins in various
therapeutic areas, aging population,
and increasing plasma collection
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.184
5. Therapeutic IgG
production
Quality criteria including low
residual level of contamination by
other proteins such as IgA, IgM,
proteolytic enzymes, or Factor
XI/XIa
Increasing use of chromatography
as an essential step for fine-tuning
purification of IgG and removal of
protein contaminants
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.185
6. Product Characteristics
Base Material Polymethylacrylate polymer
Mean Particle Size 40-90 μm
Functional Group
Strong anion exchanger
trimethylammoniumethyl
Surface Chemistry
Tentacular ligand chemistry
composed of positively-
charged groups
Benefits
Efficient separation and a
high binding capacity even
at high flow rates, long
product lifetime
A reliable anion-exchange chromatography resin to purify
plasma-derived IgG
Fractogel® EMD TMAE (M) Resin
Tentacular ligand chemistry
6Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.186
7. Polish
+
+
+
+
+
+
-
+
++
+
-
-
-
LOAD
Load:
•IgG Feed
Flow-through
•IgG
Retention
•IgA, IgM, proteolytic
enzymes, Factor IX,
Factor IXa, and other
contaminants
Negative Mode Anion-Exchange Chromatography
Contaminants are bound onto the column, IgG in the flow-through
Fractogel®
EMD TMAE (M)
Resin
Purified IgG
IgG Feed
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.187
8. Design | Methods | Results
Case
Study
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.188
9. Case Study
One step IVIG purification by AEX chromatography
Phase 1a
Optimization of
chromatographic
conditions
Phase 2
Tests of resin stability
and process
performance up to
200 cycles, including
cleaning and
sanitization
Phase 1b
Analysis of IgG purity
and reproducibility
using optimized
conditions
Experimental design
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.189
11. Representing worst
case scenario of
precipitate (I)+II+III
Caprylic Acid treated
human plasma derived
IgG: CA-IgG
Phase 1a
Preparation of IgG fraction:
Cryoprecipitation
Caprylic Acid Treatment
Chromatography
1
2
3
4
Fractogel® EMD
TMAE (M) Resin
Fractogel® EMD TMAE (M) Resin
Fractogel® EMD TMAE (M) Resin
TFF Dialysis
Cogent® μScale TFF System
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1811
12. pH
Conductivity
Total protein (Biuret)
Albumin (bromocresol green)
IgG (ELISA and immunonephelometry)
IgA (ELISA)
IgM (ELISA)
IgG subclasses (ELISA)
Analytical methods used
Gamma protein determination by zone
electrophoresis
SDS-PAGE reducing/non-reducing conditions
Thrombogenic and proteolytic activity potential
FXIa assay (FXIa Hyphen)
Thrombin generation assay (Technoclone)
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1812
13. Phase 1a
IgG feed purity check: SDS-PAGE
←IgG
13
←Heavy chain
←Light chain
SDS-PAGE
• Under non-reduced conditions:
one main protein band of ca.
160kDa (intact IgG)
• Under reduced conditions:
protein bands of ca. 50kDa (IgG heavy
chain) and ca. 25kDa (IgG light chain)
Non-reduced Reduced
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1813
14. ELISA confirms:
• Higher content in IgA and IgM in the CA-IgG (compared to a control IgG prepared from fraction II)
• CA-IgG is roughly equivalent to a Cohn precipitate (I)+II+III (worst case fraction)
• CA-IgG fraction contains a high, physiological proportion of IgA and IgM (worst case fraction)
Phase 1a
IgG, IgA and IgM mean content in “CA” feed by ELISA
C A -Ig G
8 0 .1 4 % Ig G
1 7 .7 2 % Ig A
2 .1 4 % Ig M
C o n tro l-Ig G F e e d
9 8 .7 6 % Ig G
1 .2 2 % Ig A
0 .0 2 % Ig M
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1814
15. Chromatographic steps Typical loading chromatographic conditions
1 mL packed column (convenient for quick
investigations)
Expected loading capacity: 150-200 mg of
(purified) IgG per 1 mL of resin
Phase 1a
Chromatographic conditions testing: pH 5.7, pH 6.0 and pH 6.3
Parameter Parameter values
Residence time 6.5 min
Linear flow rate 185 cm/hr
Pre-equilibration
250 mM sodium acetate
buffer, 3 CV
Equilibration
25 mM sodium acetate
buffer, 5 CV
Wash
500 mM sodium acetate
buffer, 3 CV
Regeneration
1.5 M NaCl in 250 mM
sodium acetate, pH 4.5, 2
CV
CIP
0.5 M/1.0 M NaOH, 3 CV;
10 min. residence time
(total contact time: 30 min.)
0
1000
2000
mL
mAu
80604020
IgG flow-through
IgA and IgM waste
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1815
16. Phase 1a Result
pH 5.7 to pH 6.3 range has limited impact on chromatographic
performance
Mean of 3 cycles at each pH
10 15 20 25 30 35 40
0
200
400
600
800
1000
IgG injection (mL)
IgG(mg/dl)
pH5.7
pH6.0
pH6.3Feed
Content of IgG in flow-through
Wash
• Limited pH impact
• Gel robustness
Limited loss of IgG
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1816
17. Phase 1a Result
Impact of pH on IgA chromatographic behavior
10 15 20 25 30 35 40
0
50
100
150
IgG injection (mL)
IgA(mg/dl)
pH5.7
pH6.0
pH6.3
Mean of 3 cycles at each pH
Feed
Content of IgA in
flow-through
fractions
Wash
IgA binding on gel
Limited pH impact
Gel robustness
Limited impact of pH on IgA
content in flow-through at
least until a volume of
injection of ca. 20 mL of Ig
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1817
18. Phase 1a Result
Impact of pH on IgM chromatographic behavior
10 15 20 25 30 35 40
0
5
10
15
20
25
IgG injection (mL)
IgM(mg/dl)
pH5.7
pH6.0
pH6.3
Mean of 3 cycles at each pH
Feed
Content of IgM in
flow-through fractions
Wash
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1818
19. Even under worst-case conditions (crude IgG feed and high
linear flow rate of 185 cm/hr):
✓ Obvious preferential and reproducible binding (and
therefore removal) of IgA at pH 5.7-6.3 allowing to
purify IgG in a negative chromatographic mode
✓ Removal of IgM is also apparent, but apparently less
efficient than for IgA, possibly due to IgM content in this
CA-IgG fraction
No apparent impact of pH on IgG recovery (flow-through)
and IgA and IgM removal (binding), clearly proving
Fractogel® EMD TMAE (M) resin robustness in the pH 5.7 to
pH 6.3 range
The data allowed to select loading conditions for further
studies (phase 1b)
Phase 1a
Conclusions
19 Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.18
20. Optimal Loading Volume Determination
Considering the content in IgA and IgM of the CA-IgG fraction
IgG recovery reaches
90% at IgG injection
volume of 12.5 mL
IgA removal reaches
80% with a volume of
IgG of 12.5 mL
30 20 12.5 10
0
20
40
60
80
100
IgG Injection volume (mL)
IgGRecovery(percentage)
IgG recovery
30 mL 89%
20 mL 93%
12.5 mL 94%
10 mL 90%
30 20 12.5 10
0
20
40
60
80
100
IgG Injection volume (mL)
IgARemoval(percentage)
IgA removal
30 mL 46%
20 mL 59%
12.5 mL 84%
10 mL 89%
IgM removal reaches
90% at IgG injection
volume of 12.5 mL
30 20 12.5 10
0
20
40
60
80
100
IgG Injection volume (mL)
IgMRemoval(percentage)
IgM removal
30 mL 39%
20 mL 60%
12.5 mL 90%
10 mL 92%
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1820
21. Optimal loading volume
12.5 mL CA-IgG chosen
corresponding to about 137 mg of total proteins
Volume used for phase 1b
Phase 1a Conclusions
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1821
22. Phase 1b
Analysis of IgG purity
using optimized
conditions: 10 cycles
pH 6.0
12.5 mL
185 cm/hr
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1822
24. Phase 1b Results
The concentration of total protein (Biuret)
Sample Total Protein (mg/mL)
Feed 11
C1 Flow-through 4
C2 Flow-through 4
C3 Flow-through 4
C4 Flow-through 4
C5 Flow-through 4
C6 Flow-through 4
C7 Flow-through 4
C8 Flow-through 4
C9 Flow-through 4
C10 Flow-through 4
Sample Total Protein (mg/mL)
C1 Wash 6
C2 Wash 7
C3 Wash 7
C4 Wash 7
C5 Wash 6
C6 Wash 7
C7 Wash 7
C8 Wash 7
C9 Wash 7
C10 Wash 6
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1824
25. Phase 1b Results
Recovery of IgG, IgA, IgM
IgGRecovery(percentage)
C
y
c
le
1
C
y
c
le
2
C
y
c
le
3
C
y
c
le
4
C
y
c
le
5
C
y
c
le
6
C
y
c
le
7
C
y
c
le
8
C
y
c
le
9
C
y
c
le
1
0
0
2 0
4 0
6 0
8 0
1 0 0
Mean IgG recovery in the flow-
through is 94.18% (89.66-97.66%)
IgARemoval(percentage)
C
y
c
le
1
C
y
c
le
2
C
y
c
le
3
C
y
c
le
4
C
y
c
le
5
C
y
c
le
6
C
y
c
le
7
C
y
c
le
8
C
y
c
le
9
C
y
c
le
1
0
0
2 0
4 0
6 0
8 0
1 0 0
Mean IgA content in the wash is
84.00% (82.18-86.18%)
IgMRemoval(percentage)
C
y
c
le
1C
y
c
le
2C
y
c
le
3C
y
c
le
4C
y
c
le
5C
y
c
le
6C
y
c
le
7C
y
c
le
8C
y
c
le
9
C
y
c
le
1
0
0
2 0
4 0
6 0
8 0
1 0 0
Mean IgM content in the wash is
89.82% (81.81-99.80%)
*IgG (ELISA) *IgA (ELISA) *IgM (ELISA)
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1825
26. Phase 1b Results
IgG, IgA and IgM in feed, flow-through and wash:
10 cycles with good reproducibility
Blue: IgG
Red: IgA
Green: IgM
• Good reproducibility of Fractogel® EMD TMAE (M) resin to remove IgA and IgM from CA-IgG fraction
• Improved IgG purity
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1826
27. Phase 1b Results
IgG subclass distribution in feed: Flow-through remains throughout
10 cycles
• Reproducible content in all 4 subclasses
• Slight decrease in subclass 4 content in the flow-through (subclass 4 has a pI closer to that of
IgA, so more likely to be adsorbed on the resin)
Mean of 10 cycles
58.40% IgG1
38.58% IgG2
1.37% IgG3
1.63% IgG4
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1827
28. Phase 1b Results
SDS PAGE shows reproducible removal of contaminants in the wash
Flow-through
←IgG
←LC
←HC
Flow-through
Non-reduced Reduced
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1828
29. Phase 1b Results
FXIa content
Sample Factor XIa Concentration (mg/dL)
Feed
<0.0000575
Cycle 1 Flow-through
Cycle 2 Flow-through
Cycle 3 Flow-through
Cycle 4 Flow-through
Cycle 5 Flow-through
Cycle 6 Flow-through
Cycle 7 Flow-through
Cycle 8 Flow-through
Cycle 9 Flow-through
Cycle 10 Flow-through
* FXIa activity tested by FXIa Hyphen
• Chromatographic
separation on
Fractogel® EMD
TMAE (M) resin did
not induce FXIa
activity
• The process does
not induce
detectable risks of
thrombogenicity
linked to activation
of Factor XI
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1829
30. Phase 1b Results
Thrombin generation assay (TGA)
*Thrombin generation assay (Technoclone)
RC High
Thrombin
[nmol/L]
Time to Peak
[min.]
Velocity
Index
AUC
Feed 16.16 35.00 0.52 94.11
Feed (diluted 2.46X) ND ND ND ND
Cycle 1 Flow-through ND ND ND ND
Cycle 2 Flow-through ND ND ND ND
Cycle 3 Flow-through ND ND ND ND
Cycle 4 Flow-through ND ND ND ND
Cycle 5 Flow-through ND ND ND ND
Cycle 6 Flow-through ND ND ND ND
Cycle 7 Flow-through ND ND ND ND
Cycle 8 Flow-through ND ND ND ND
Cycle 9 Flow-through ND ND ND ND
Cycle 10 Flow-through ND ND ND ND
0,00
2,00
4,00
6,00
8,00
10,00
12,00
14,00
16,00
18,00
0 10 20 30 40 50 60 70 80 90 100
nMThrombin
time [min]
Thrombin
• Slight tendency for thrombin generation in
the feed
• No thrombin generation was reproducibly
found in the 10 flow-through cycles
• Fractogel® EMD TMAE (M) chromatography
resin may contribute to the removal of
factors (proteolytic enzymes) responsible
for thrombin generation
• Spiking experiments with pro-thrombotic
factors may be considered to confirm
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1830
31. Even under worst-case conditions (crude IgG feed + high linear flow
rate):
✓ Powerful removal of IgA and IgM at 12.5 mL CA-IgG/mL of gel
(=75-90 mg IgG)
Robustness of Fractogel® EMD TMAE resin at pH 6.0:
✓ Consistent removal of IgA and IgM (constant proportion present
in the flow-through)
The wash fraction is proportionally enriched in IgA and IgM,
confirming adsorption on TMAE
Good consistent recovery of IgG subclasses (only limited decrease in
the proportion of IgG subclass 4)
Improvement in IgG purity and removal of IgA and IgM observed by:
✓ Immunonephelometry
✓ ELISA
✓ Zone electrophoresis
✓ SDS-PAGE under reduced and non-reduced conditions, which also
reveals removal of various protein contaminants
No generation of thrombogenicity (based on FXIa and TGA data)
during Fractogel® EMD TMAE chromatography resin, and potential
removal of thrombogenic factors
Phase 1b
Conclusion
using CA-IgG
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1831
32. • Control fraction II IgG containing more purified IgG,
199 mg of IgG/mL of gel loading
• 5 cycles done
• Checked:
• Chromatographic profile
• IgG recovery: 97.50% (range 91.76-103.72%)
• IgA removal: 92.16% (range 88.64-94.18%)
• IgM removal: 66.75% (range 59.77-74.14%)
• IgG/A/M distributions in Feed/Flow-through/Wash
over 5 cycles show good consistency
• IgG subclasses distribution consistent, with IgG4
dropped a little
Phase 1b Conclusion
Control experiment using Fraction II IgG
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1832
Flow-through Cycle 5
Flow-through Cycle 5
33. Cycle 1
2017Nov22no002:10_UV 2017Nov22no002:10_Cond 2017Nov22no002:10_pH
2017Nov22no002:10_Fractions
0
200
400
600
800
1000
mAu
0 20 40 60 80 ml
1 2 3 4 5 6 7 Waste 8 1011 Waste
Flow-through
Wash
Regeneration/CIP
Chromatographic profile obtained with purified IgG
Phase 1b Conclusion
Evidence of adsorption of contaminants on the resin
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1833
34. Phase 2
Tests of stability and
robustness of resin
performance up to 200
cycles pH 6.0
12.5 mL
185 cm/hr
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1834
35. Phase 2
Test conditions and a typical chromatographic profile
Buffer Contents pH
Conductivity
(mS)
Pre-
equilibration
500 mM sodium acetate 6 20.8
Equilibration 25 mM sodium acetate 6 1.97
Wash 500 mM sodium acetate 4.5 22.8
Regeneration
1.5 M NaCl in 250 mM
sodium acetate
4.5 >30.00
CIP
0.5 M NaOH (odd run
numbers)
1 M NaOH (even run
numbers)
Sample pH Conductivity (mS)
Cycle 1-IgG 6 1.737
Cycle 201-IgG 6 1.744
2018Feb11no004:10_UV 2018Feb11no004:10_Cond
2018Feb11no004:10_pH 2018Feb11no004:10_Fractions
2018Feb11no004:10_Logbook
0
500
1000
1500
2000
mAu
4.0
6.0
8.0
10.0
0 20 40 60 80 ml
Break point 2 Break point 5 Break point 9 Break point 10 Break point 12
1 3 4 5 6 Waste 10 Waste
Flow-through
Wash Regeneration/CIP
Cycle 1
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1835
36. Phase 2
Consistent chromatographic profile throughout 201 cycles
Cycle 11
2018Feb12no002:10_UV 2018Feb12no002:10_Cond
2018Feb12no002:10_pH 2018Feb12no002:10_Fractions
2018Feb12no002:10_Logbook
0
500
1000
1500
2000
mAu
4.0
6.0
8.0
10.0
0 20 40 60 80 ml
Break point 2 Break point 5 Break point 9 Break point 10 Break point 12
1 3 4 5 6 Waste 10 Waste
Cycle 31
2018Feb21no001:10_UV 2018Feb21no001:10_Cond
2018Feb21no001:10_pH 2018Feb21no001:10_Fractions
2018Feb21no001:10_Logbook
0
500
1000
1500
2000
mAu
4.0
6.0
8.0
10.0
0 20 40 60 80 ml
Break point 2 Break point 5 Break point 9 Break point 10 Break point 12
1 3 4 5 6 Waste 10 Waste
Cycle 512018Feb23no002:10_UV 2018Feb23no002:10_Cond
2018Feb23no002:10_pH 2018Feb23no002:10_Fractions
2018Feb23no002:10_Logbook
0
500
1000
1500
2000
mAu
4.0
6.0
8.0
10.0
0 20 40 60 80 ml
Break point 2 Break point 5 Break point 9 Break point 10 Break point 12
1 3 4 5 6 Waste 10 Waste
Cycle 71
2018Feb25no002:10_UV 2018Feb25no002:10_Cond
2018Feb25no002:10_pH 2018Feb25no002:10_Fractions
2018Feb25no002:10_Logbook
0
500
1000
1500
2000
mAu
4.0
6.0
8.0
10.0
0 20 40 60 80 ml
Break point 2 Break point 5 Break point 9 Break point 10 Break point 12
1 3 4 5 6 Waste 10 Waste
Cycle 91
2018Feb27no001:10_UV 2018Feb27no001:10_Cond
2018Feb27no001:10_pH 2018Feb27no001:10_Fractions
2018Feb27no001:10_Logbook
0
500
1000
1500
2000
mAu
4.0
6.0
8.0
10.0
0 20 40 60 80 ml
Break point 2 Break point 5 Break point 9 Break point 10 Break point 12
1 3 5 6 Waste 10 Waste
Cycle 111
2018Mar02no002:10_UV 2018Mar02no002:10_Cond
2018Mar02no002:10_pH 2018Mar02no002:10_Fractions
2018Mar02no002:10_Logbook
0
500
1000
1500
2000
mAu
4.0
6.0
8.0
10.0
0 20 40 60 80 ml
Break point 2 Break point 5 Break point 9 Break point 10 Break point 12
1 3 4 5 6 Waste 10 Waste
2018Mar04no002:10_UV 2018Mar04no002:10_Cond
2018Mar04no002:10_pH 2018Mar04no002:10_Fractions
2018Mar04no002:10_Logbook
0
500
1000
1500
2000
mAu
4.0
6.0
8.0
10.0
0 20 40 60 80 ml
Break point 2 Break point 5 Break point 9 Break point 10 Break point 12
1 3 4 5 6 Waste 10 Waste
Cycle 131 Cycle 151
2018Mar09no002:10_UV 2018Mar09no002:10_Cond
2018Mar09no002:10_pH 2018Mar09no002:10_Fractions
2018Mar09no002:10_Logbook
0
500
1000
1500
2000
mAu
4.0
6.0
8.0
10.0
0 20 40 60 80 ml
Break point 2 Break point 5 Break point 9 Break point 10 Break point 12
1 3 4 5 6 Waste 10 Waste
Cycle 181
2018Mar14no001:10_UV 2018Mar14no001:10_Cond
2018Mar14no001:10_pH 2018Mar14no001:10_Fractions
2018Mar14no001:10_Logbook
0
500
1000
1500
2000
mAu
4.0
6.0
8.0
10.0
0 20 40 60 80 ml
Break point 2 Break point 5 Break point 9 Break point 10 Break point 12
1 3 4 5 6 Waste 10 Waste
Cycle 201
2018Mar16no001:10_UV 2018Mar16no001:10_Cond
2018Mar16no001:10_pH 2018Mar16no001:10_Fractions
2018Mar16no001:10_Logbook
0
500
1000
1500
2000
mAu
4.0
6.0
8.0
10.0
0.0 20.0 40.0 60.0 80.0 ml
Break point 2 Break point 5 Break point 9 Break point 10 Flow 0.0 ml/min
12 3 4 5 6 7 8 10 Waste
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1836
37. Phase 2
Total protein consistent throughout 201 cycles
Sample Total Protein (mg/mL) Albumin (mg/mL)
C1 Feed 12 < 2
C201 Feed 10 < 2
C1 Flow-through 4 < 2
C201 Flow-through 4 < 2
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1837
38. mean IgG recovery in the flow-
through is 94.22% (range 91.09-
97.78%)
Phase 2
IgG Recovery, IgA Removal, IgM Removal consistency test
IgGRecovery(percentage)
C
1C
1
1
C
2
1
C
3
1C
4
1
C
5
1
C
6
1
C
7
1C
8
1
C
9
1C
1
0
1C
1
1
1C
1
2
1C
1
3
1C
1
4
1C
1
5
1C
1
6
1C
1
7
1C
1
8
1C
1
9
1C
2
0
1
0
5 0
1 0 0
1 5 0
IgARemoval(percentage)
C
1C
1
1
C
2
1
C
3
1C
4
1
C
5
1
C
6
1
C
7
1C
8
1
C
9
1C
1
0
1C
1
1
1C
1
2
1C
1
3
1C
1
4
1C
1
5
1C
1
6
1C
1
7
1C
1
8
1C
1
9
1C
2
0
1
0
5 0
1 0 0
1 5 0
mean IgA content in the wash is
96.16% (range 87.48-107.15%)
IgMRemoval(percentage)
C
1C
1
1
C
2
1
C
3
1C
4
1
C
5
1
C
6
1
C
7
1C
8
1
C
9
1C
1
0
1C
1
1
1C
1
2
1C
1
3
1C
1
4
1C
1
5
1C
1
6
1C
1
7
1C
1
8
1C
1
9
1C
2
0
1
0
5 0
1 0 0
1 5 0
mean IgM content in the wash is
96.89% (range 87.22-108.06%)
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1838
39. Phase 2 Results
Reproducible removal of IgA and IgM over the 201 cycles performed
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1839
40. Phase 2 Results
Pie chart of IgG, IgA and IgM in Wash for 201 cycles
C 1 W a s h C 1 1 W a s h C 2 1 W a s h C 3 1 W a s h C 4 1 W a s h C 5 1 W a s h
C 6 1 W a s h C 7 1 W a s h C 8 1 W a s h C 9 1 W a s h C 1 0 1 W a s h C 1 1 1 W a s h
C 1 2 1 W a s h C 1 3 1 W a s h C 1 4 1 W a s h C 1 5 1 W a s h C 1 6 1 W a s h C 1 7 1 W a s h
C 1 8 1 W a s h C 1 9 1 W a s h C 2 0 1 W a s h
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1840
41. Phase 2 Results
Reproducible IgG subclass distribution in the purified flow-through
over the 201 cycles performed
C101 Flow-through
Feed
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1841
42. Percentage(%)
C
1
F
e
e
d
C
2
0
1
F
e
e
d
C
1
F
T
(c
o
n
c
.
3
X
)C
2
0
1
F
T
(c
o
n
c
.
3
X
)
0
1 0
2 0
3 0
4 0
5 0
6 0
7 0
8 0
9 0
1 0 0
G a m m a + B e ta
A lp h a 1
A lp h a 2
A lb u m in
Zone electrophoresis
C1
Feed
C201
Feed
C1 FT
(conc. 3X)
C201 FT
(conc. 3X)
Albumin 1.7% 1.8% 1.1% 1.4%
Alpha 2 9.2% 9.4% 4.3% 3.2%
Alpha 1 4.4% 2.8% 0.3% 0.3%
Gamma +
Beta
84.7% 86.0% 94.3% 95.1%
C1 Feed C201 Feed
C1 Flow-through
(conc. 3X)
C201 Flow-through
(conc. 3X)
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1842
44. • Chromatographic separation on Fractogel® EMD
TMAE (M) resin did not induce the generation of
FXIa activity
• Process does not increase possible risks of
thrombogenicity linked to activation of Factor XI
• These findings are constant over 201 cycles
Phase 2
Factor XIa assay and TGA activity
Sample Factor XIa concentration (mg/dL)
Cycle 1 Feed
<0.0000575
Cycle 1 Flow-through
Cycle 201 Feed
Cycle 201 Flow-through
0,00
0,50
1,00
1,50
2,00
0 10 20 30 40 50 60 70 80 90 100
nMThrombin
time [min]
Cycle 1 Feed (2.25X)
Cycle 201 Feed (2.25X)
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1844
45. Phase 2
Conclusion
1
Fractogel® EMD TMAE (M) resin demonstrates to have
excellent resistance for over 201 cycles in its capacity
to robustly remove IgA and IgM from a crude CA-IgG
preparation, under good recovery of IgG and without
affecting the IgG subclass distribution
Excellent resistance to cleaning/sanitization cycles
2
The chromatographic purification process does not
generate detectable thrombin activity and may remove
thrombogenic factors present in the CA-IgG feed
3
Optimal loading conditions would have to be
determined by each plasma fractionator based on the
level of IgA or IgM present in the IgG fraction to be
purified
Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1845
46. Chromatographic Strategies for IVIG Purification - Part 2 | 29.11.1846
How to
systematically
set up an IVIG
purification
strategy
How to integrate
chromatography
In a downstream
process of IVIG
How to evaluate the
quality of purified
IVIG
How to confirm
the robustness of
an upgraded IVIG
purification
process