A fascinating webinar with Dora Taggert of Microbial Insights discussing the range of sophisticated yet accessible microbiological tools for contaminated site monitoring.
This document summarizes a short course on DNA barcoding methods that will cover equipment needed for PCR amplification, PCR protocols, product verification using agarose gels, minimizing contamination risks, and shipping and storing DNA extracts. The course will provide an overview of typical PCR protocols and note that protocols depend on sample quality and scale of operations. Small PCR volumes and use of trehalose are discussed as cost-saving measures. Primer selection is also highlighted as a key factor in success.
Application Note: Gross Alpha Measurements in Aqueous Samples Using Extractio...PerkinElmer, Inc.
A new approach to the measurement of gross alphas in liquid samples, which combines Eichrom's extraction technology with PerkinElmer's liquid scintillation analysis technology, is presented. Difficulties with sample preparation and detection efficiency have always been problematic with the traditional method of gas flow proportional counting (GPC). The use of Eichrom's extraction resins and alpha liquid scintillation counting (LCS) has resolved the sample preparation problems, associated with drying the sample on a planchet. With its superior detection efficiency over GPC, this combination provides a better counting platform. Thus, many of the inherent limitations associated with the traditional GPC method are overcome by using this new approach and results demonstrate faster sample preparation, lower detection limits and shorter counting times.
A Novel approach for quantitative real-time particle analysis of lentiviral v...Myriade
Lentiviral vectors are efficient vehicles for stable gene transfer in dividing and non-dividing cells. They tend to be increasingly used as a powerful tool to introduce genes into cells ex vivo, for instance in CAR-T cell therapies.
During manufacturing and production of lentiviral vectors, relevant quality control is necessary to allow batch release (1). Among standard quality control methods that can be used, quantification of lentiviral vector particles – or physical titer – is one of the most important. Up to now, this characterization can be achieved either indirectly with p24 protein quantification or with physical methods like Tunable Resistive Pulse Sensing (TRPS) for example, both methods implying prior preparation of samples (lysis, dilution or filtration). These two methods thus show important limitations as they cannot accurately reflect the true nature of the product, in addition to being relatively time-consuming (2).
Myriade, a French company created in 2017, is developing Videodrop, a new optical technique performing real-time, user-friendly, and label-free measurement of lentiviral vector physical titer. This method, based on full-field interferometry (3), was tested on various lentiviral vector samples: in a context of Drug Product (DP) release as well as in-process controls.
We compared three lentiviral physical titration methods on aratinga.bio productions: p24 ELISA, qNano and Videodrop – Myriade instrument. The correlation between Videodrop analysis and the other two methods appeared to be robust, with high R² values. These results suggest that Myriade technology is relevant for DP release as well as in-process controls, offering the ability to be a tool for continuous improvement. It is an easy-touse and fast alternative to the standard more complex and time-consuming physical titration methods.
Erica Canzler - Advances and Lessons Learned in DecontaminationMatthew Kirkby
Advances in decontamination technologies and strategies were tested to help improve response to biological incidents. The Biological Operational Test and Evaluation Project tested three decontamination methods - vaporized hydrogen peroxide, bleach, and chlorine dioxide fumigation. Chlorine dioxide was most effective at reducing spore levels, while bleach produced the most waste. Overall costs were dominated by waste management. The methyl bromide fumigation study demonstrated the feasibility of using this technology for decontamination of structures. Efforts are also underway to develop strategies to safely restore contaminated underground transportation systems like subways following an incident.
Pittcon 2012 Cs Fi Invited Sers Talks Drugs In Salivainscore
This document describes a method for rapidly detecting drugs of abuse in saliva using surface-enhanced Raman spectroscopy (SERS). SERS can detect drugs at concentrations as low as 10-8 M (parts per billion) and identify them from their unique Raman spectra within 10 minutes. The method was tested on over 35 drugs and found to accurately detect and identify drugs spiked into saliva samples at 50 parts per billion or above. If developed into a portable device, it could provide a fast, non-invasive way to screen for drug impairment in situations like roadside stops or emergency rooms.
1) The document describes an automated process for in vitro selection that was developed to generate nucleic acid aptamers faster than the traditional manual selection process.
2) An augmented Beckman Biomek 2000 pipetting robot was programmed to automate the major steps of in vitro selection, including preparing and purifying RNA, filtering RNA-target complexes, and amplifying selected sequences, in order to reduce the time needed to select aptamers from weeks or months to just days.
3) Initial attempts at automated selection yielded replication parasites but optimization suppressed their emergence and enabled the selection of true nucleic acid ligands binding to targets.
TWCA Annual Convention: Optimizing Slime Control Chemical Feed in TRWD Raw Wa...TWCA
This presentation discusses optimizing slime control and chemical feed in the raw water pipelines of Tarrant Regional Water Districts. It presents results from bench-scale and full-scale testing to evaluate chloramine decay and the impact of nitrifying bacteria in the pipelines. The testing showed that higher chlorine to nitrogen ratios, lower pH levels, and the presence of nitrifying bacteria increased chloramine decay. It also showed chloramine loss from nitrification occurring in the pipelines. The presentation recommends improvements to existing chemical systems and designing new chloramine feed systems to better control slime and protect the pipelines.
Cancer Research & the Challenges of FFPE Samples – An IntroductionQIAGEN
A cascade of complex genetic and epigenetic changes regulate tumor formation and progression. Gene expression analyses can shed light on these changes at a molecular level and identify the key genes and associated pathways involved in cancer. Often the samples used in cancer research are FFPE samples, which pose a significant challenge in terms of nucleic acid quality. The quality of nucleic acids extracted from FFPE samples depends on a number of factors, including how the samples were handled before, during and after fixation and embedding.
Dr. Vishwadeepak Tripathi describes the variability of sample purification from FFPE samples – in particular, samples to be used in cancer research. What are the challenges and solutions, and what quality control approach can ensure credible results? This webinar will focus on sample purification and the quality control of FFPE samples and compare different automated purification procedures.
This document summarizes a short course on DNA barcoding methods that will cover equipment needed for PCR amplification, PCR protocols, product verification using agarose gels, minimizing contamination risks, and shipping and storing DNA extracts. The course will provide an overview of typical PCR protocols and note that protocols depend on sample quality and scale of operations. Small PCR volumes and use of trehalose are discussed as cost-saving measures. Primer selection is also highlighted as a key factor in success.
Application Note: Gross Alpha Measurements in Aqueous Samples Using Extractio...PerkinElmer, Inc.
A new approach to the measurement of gross alphas in liquid samples, which combines Eichrom's extraction technology with PerkinElmer's liquid scintillation analysis technology, is presented. Difficulties with sample preparation and detection efficiency have always been problematic with the traditional method of gas flow proportional counting (GPC). The use of Eichrom's extraction resins and alpha liquid scintillation counting (LCS) has resolved the sample preparation problems, associated with drying the sample on a planchet. With its superior detection efficiency over GPC, this combination provides a better counting platform. Thus, many of the inherent limitations associated with the traditional GPC method are overcome by using this new approach and results demonstrate faster sample preparation, lower detection limits and shorter counting times.
A Novel approach for quantitative real-time particle analysis of lentiviral v...Myriade
Lentiviral vectors are efficient vehicles for stable gene transfer in dividing and non-dividing cells. They tend to be increasingly used as a powerful tool to introduce genes into cells ex vivo, for instance in CAR-T cell therapies.
During manufacturing and production of lentiviral vectors, relevant quality control is necessary to allow batch release (1). Among standard quality control methods that can be used, quantification of lentiviral vector particles – or physical titer – is one of the most important. Up to now, this characterization can be achieved either indirectly with p24 protein quantification or with physical methods like Tunable Resistive Pulse Sensing (TRPS) for example, both methods implying prior preparation of samples (lysis, dilution or filtration). These two methods thus show important limitations as they cannot accurately reflect the true nature of the product, in addition to being relatively time-consuming (2).
Myriade, a French company created in 2017, is developing Videodrop, a new optical technique performing real-time, user-friendly, and label-free measurement of lentiviral vector physical titer. This method, based on full-field interferometry (3), was tested on various lentiviral vector samples: in a context of Drug Product (DP) release as well as in-process controls.
We compared three lentiviral physical titration methods on aratinga.bio productions: p24 ELISA, qNano and Videodrop – Myriade instrument. The correlation between Videodrop analysis and the other two methods appeared to be robust, with high R² values. These results suggest that Myriade technology is relevant for DP release as well as in-process controls, offering the ability to be a tool for continuous improvement. It is an easy-touse and fast alternative to the standard more complex and time-consuming physical titration methods.
Erica Canzler - Advances and Lessons Learned in DecontaminationMatthew Kirkby
Advances in decontamination technologies and strategies were tested to help improve response to biological incidents. The Biological Operational Test and Evaluation Project tested three decontamination methods - vaporized hydrogen peroxide, bleach, and chlorine dioxide fumigation. Chlorine dioxide was most effective at reducing spore levels, while bleach produced the most waste. Overall costs were dominated by waste management. The methyl bromide fumigation study demonstrated the feasibility of using this technology for decontamination of structures. Efforts are also underway to develop strategies to safely restore contaminated underground transportation systems like subways following an incident.
Pittcon 2012 Cs Fi Invited Sers Talks Drugs In Salivainscore
This document describes a method for rapidly detecting drugs of abuse in saliva using surface-enhanced Raman spectroscopy (SERS). SERS can detect drugs at concentrations as low as 10-8 M (parts per billion) and identify them from their unique Raman spectra within 10 minutes. The method was tested on over 35 drugs and found to accurately detect and identify drugs spiked into saliva samples at 50 parts per billion or above. If developed into a portable device, it could provide a fast, non-invasive way to screen for drug impairment in situations like roadside stops or emergency rooms.
1) The document describes an automated process for in vitro selection that was developed to generate nucleic acid aptamers faster than the traditional manual selection process.
2) An augmented Beckman Biomek 2000 pipetting robot was programmed to automate the major steps of in vitro selection, including preparing and purifying RNA, filtering RNA-target complexes, and amplifying selected sequences, in order to reduce the time needed to select aptamers from weeks or months to just days.
3) Initial attempts at automated selection yielded replication parasites but optimization suppressed their emergence and enabled the selection of true nucleic acid ligands binding to targets.
TWCA Annual Convention: Optimizing Slime Control Chemical Feed in TRWD Raw Wa...TWCA
This presentation discusses optimizing slime control and chemical feed in the raw water pipelines of Tarrant Regional Water Districts. It presents results from bench-scale and full-scale testing to evaluate chloramine decay and the impact of nitrifying bacteria in the pipelines. The testing showed that higher chlorine to nitrogen ratios, lower pH levels, and the presence of nitrifying bacteria increased chloramine decay. It also showed chloramine loss from nitrification occurring in the pipelines. The presentation recommends improvements to existing chemical systems and designing new chloramine feed systems to better control slime and protect the pipelines.
Cancer Research & the Challenges of FFPE Samples – An IntroductionQIAGEN
A cascade of complex genetic and epigenetic changes regulate tumor formation and progression. Gene expression analyses can shed light on these changes at a molecular level and identify the key genes and associated pathways involved in cancer. Often the samples used in cancer research are FFPE samples, which pose a significant challenge in terms of nucleic acid quality. The quality of nucleic acids extracted from FFPE samples depends on a number of factors, including how the samples were handled before, during and after fixation and embedding.
Dr. Vishwadeepak Tripathi describes the variability of sample purification from FFPE samples – in particular, samples to be used in cancer research. What are the challenges and solutions, and what quality control approach can ensure credible results? This webinar will focus on sample purification and the quality control of FFPE samples and compare different automated purification procedures.
Peritoneal Dialysis And Secondary Renal FunctionRodion Stolyar
This study aimed to develop a renal function replacement system using peritoneal dialysis (PD) with a bioartificial renal tubule construct (BREC-d). Sheep underwent bilateral nephrectomies and were placed on either cellular or acellular BREC-d with PD for up to 10 days. The cellular group showed maintenance of acceptable blood pressure and respiration rates. More frequent PD exchanges are needed to better control electrolytes and waste products. Renal cells remained viable in the BREC-d, showing potential for this approach with further optimization of PD fluid and longer study durations.
Automated Nucleic Acid Purification from Diverse Sample types using dedicated...QIAGEN
This webinar will focus on the automation of QIAGEN’s new line of DNA and RNA sample prep kits for the microbiome. We will show how automation on the QIAcube enables efficient and reliable use of these samples for sensitive downstream applications such as qPCR and NGS. In addition, you will learn how to successfully use the CLC Microbial Genomics Module for metagenome sequencing and identification of microbial composition and diversity.
This document summarizes research on the psychrophilic degradation of long chain alkanes by microorganisms. Key findings include:
1) Pseudomonas frederiksbergensis and Rhodococcus erythropolis were able to degrade n-alkanes from C10-C22 at 4-20°C with optimal degradation by P. frederiksbergensis occurring at 15°C and 7.0 pH.
2) P. frederiksbergensis degraded alkanes via terminal oxidation and detection of metabolites, starting with hydroxylation of alkanes to fatty acids.
3) Genes encoding alkane hydroxylase and alcohol dehydrogenase from P. frederik
The document describes metabolic engineering experiments to optimize Escherichia coli for efficient conversion of glucose to pyruvate. Key modifications included deleting the poxB gene encoding pyruvate oxidase to increase pyruvate production, and decreasing pyruvate dehydrogenase levels through deletion of pdhR and replacing its promoter. Initial experiments successfully deleted poxB but found contamination in strains. Batch cultures showed highest pyruvate concentrations of 6.14 g/L for strain R1 ΔpoxB. While results were mixed, the work provided experience with laboratory techniques and engineering a microbial strain through a full project.
Application Note: A Rapid Procedure for Screening Transuranium Nuclides in Ur...PerkinElmer, Inc.
One of the most extensive tasks is the field of bioassay analysis is the determination of pure alpha- (and beta-) emitting radionuclides from the nuclear fuel cycle such as (234)U and (235)U, or anthropogenic (239)Pu and (241)Am in urine samples. However, any radiochemical method, which is applied to perform such analyses, has to be highly sensitive since even small amounts of incorporated radionuclides decaying by alpha emission may contribute to harmful doses to human organs.
Since alpha radiation has an extremely short penetration length in water and solid substances, direct counting of a salt residue of dry ashed urine is not possible. Therefore, complex radiochemical techniques have been developed for efficient separation of the transuranium elements from the bulk matrix. However, in addition to several purification steps, these methods require the production of almost weightless planar sources (e.g. via electrolytic deposition) in order to perform radioassays with proportional or surface barrier detector. In contrast to the extensive preparative techniques, fast methods using alpha/beta-LSC are of increasing interest. Due to the efficient detection of alpha emitters in a liquid scintillation cocktail, extensive radiochemical purification procedures are not necessary provided the sample is homogeneous in the liquid scintillation cocktail.
(originally aired 11-14-12)
Although biofuels are an attractive alternative to fossil fuels, large scale development is currently challenging. Development of renewable fuel characterization, processes, and contaminant analysis using robust analytical methods is needed. Here, focus is on Ion Chromatography—a proven technique for providing fast, reliable answers during research to production—with HPAE-PAD technology for carbohydrate analysis in feedstock and method parameter optimization (including column chemistry) for efficient separation of mono- and disaccharides with good resolution, linearity, and accuracy over a broad dynamic range. Since some residual sucrose and cellobiose may be present, examples of monitoring them and other saccharides is covered, along with their impact on the fermentation process.
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.
MicroPRO, A Rapid Microbiology Method Based on Flow Cytometryguest32bcc5
This document provides information on disruptive technologies and rapid microbiology products represented by the company. It summarizes their product range including instruments for biological sample preparation, dissolution/formulation, physico-chemistry analysis, and rapid microbiology detection. It also discusses their markets in pharmaceutical, personal care, fermentation, and more. The MicroPRO instrument allows detection of bacteria, yeast and mold from various samples within 24 hours.
21st International Conference Organic Process Research & Development 2010 San...dominev
This document discusses using real-time calorimetry to improve operational efficiency. It presents case studies where ReactIR, FBRM, PVM and RTCal were used:
1) ReactIR developed kinetic models to minimize byproducts in pharmaceutical reactions and improve crystallization processes.
2) FBRM and PVM helped optimize a crystallization to reduce impurities below 0.5%.
3) RTCal validated switching to a low copper acrylamide grade for polymerization, showing a shorter induction period but similar maximum heat output. Real-time calorimetry provided process safety evaluation.
Improving Downstream Processing: Application of Excipients in DSPMilliporeSigma
Webinar summary:
This webinar will showcase the beneficial potential of using excipients during downstream processing of monoclonal antibodies.
Learning points:
In this webinar, you will see:
* An innovative excipient screening approach simulating low pH stress conditions during protein A chromatography and virus inactivation
* How the application of excipients in buffer systems can significantly improve protein stability and chromatographic performance
Abstract:
Key aspects during downstream purification of biopharmaceutical drugs are purity and process yield. Therefore, the downstream process needs to be designed in a way that the final product which will eventually end up in the patient entails low levels of product- and process related impurities (e.g. high molecular weight aggregates) as well as process related contaminants (e.g. host cell protein levels). In addition to this, the process must be capable of clearing and inactivating viruses to ensure product safety. In this webinar, we will explore the benefits of adding excipients during downstream processing on protein stability, chromatographic performance and viral inactivation.
1. The document describes the quality assurance system for sputum smear microscopy in India's Revised National Tuberculosis Control Programme (RNTCP).
2. It establishes a three level laboratory network of National Reference Laboratories, Intermediate Reference Laboratories at the state level, and Designated Microscopy Centers.
3. Quality assurance activities include internal quality control, external quality assessment including on-site evaluations, panel testing of lab staff, and random blinded rechecking of slides to evaluate performance at each level of the network.
PCR Detection of SNP Markers for Warfarin Sensitivity Using Oral Fluids
1. DNA was extracted from oral fluid samples using different methods and quantified, with yields ranging from 3-12 μg/ml. Method 2 was chosen for future work due to simplicity and consistency.
2. PCR amplification of extracted DNA was successful, demonstrating that DNA could be extracted and amplified from oral fluid samples. Inhibitors in oral fluids varied between samples.
3. Primers were designed to detect SNPs associated with warfarin sensitivity. Amplification with LNA primers distinguished between wild-type and mutant samples, showing potential for an oral fluid-based pharmacogenomic test.
The document discusses a presentation on LOINC (Logical Observation Identifiers Names and Codes) given at the 2011 Public Health Informatics conference in Atlanta, GA. The presentation provides an introduction to LOINC and covers topics such as the origins of LOINC, common elements in LOINC terms, LOINC collections like forms and surveys, and domain-specific approaches to mapping standards and terminologies in areas like microbiology. It also discusses LOINC tools and resources for mapping terms and codes.
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
Cowin Biotech is a Chinese biotech company founded in 2007 that provides molecular diagnostic products and services. The company has R&D centers in Jiangsu, Beijing, and Boston and produces over 1,000 products through independent research. Cowin's core capabilities include viral transport media, nucleic acid extraction kits, PCR reagents, and enzymes. It aims to provide a complete solution for clinical diagnostic applications.
ReaMetrix is a life science technology company founded in 2003 with facilities in San Carlos, CA and Bangalore, India. It is committed to developing high-quality, affordable diagnostic solutions for resource-limited settings. ReaMetrix has expertise in assay development, protein expression and labeling, instrumentation, and developing integrated diagnostic solutions. It currently offers products like conjugates for flow cytometry-based tests and is working on projects like affordable point-of-care tests for bacterial detection and disease monitoring.
Introduction to real-Time Quantitative PCR (qPCR) - Download the slidesQIAGEN
This slidedeck introduces the concepts of real-time PCR and how to conduct a real-time PCR assay. The topics that are covered include an overview of real-time PCR chemistries, protocols, quantification methods, real-time PCR applications and factors for success.
This document discusses the polymerase chain reaction (PCR) technique. It describes how PCR was developed by Kary Mullis in the 1980s and allows for the in vitro amplification of specific DNA sequences. The key steps in PCR include denaturation of DNA, annealing of primers, and extension of the DNA strand by a DNA polymerase. The document outlines the main components required for PCR including primers, DNA polymerase, dNTPs, buffer, and magnesium chloride. It also discusses applications of PCR like DNA sequencing, genetic testing, and the Human Genome Project. Real-time PCR is introduced as a variation that allows for detection and quantification of DNA during the PCR process. Diagrams illustrate the PCR process and setup in a thermal cycler machine
The document provides information about upcoming community events and programs in the West End neighborhood of Providence, Rhode Island. It announces the opening of the Sankofa World Market on Wednesdays from July 1st to September 30th at Knight Memorial Library, featuring food, crafts, and resources celebrating the neighborhood's diversity. It also lists dates for resident engagement meetings, homebuyer workshops, photography walks, and POWER workshops. Angie Ankoma, president of the neighborhood organization's board, is awarded the Dorothy Richardson Award for Resident Leadership for her long-time community involvement.
Este documento describe las principales biomoléculas que forman los seres vivos, incluyendo proteínas, carbohidratos, lípidos y ácidos nucleicos. Explica la estructura, función y clasificación de cada biomolécula. También cubre temas como la estructura de las proteínas a nivel primario, secundario, terciario y cuaternario. Finalmente, concluye que las biomoléculas son fundamentales para el funcionamiento normal de los organismos.
Peritoneal Dialysis And Secondary Renal FunctionRodion Stolyar
This study aimed to develop a renal function replacement system using peritoneal dialysis (PD) with a bioartificial renal tubule construct (BREC-d). Sheep underwent bilateral nephrectomies and were placed on either cellular or acellular BREC-d with PD for up to 10 days. The cellular group showed maintenance of acceptable blood pressure and respiration rates. More frequent PD exchanges are needed to better control electrolytes and waste products. Renal cells remained viable in the BREC-d, showing potential for this approach with further optimization of PD fluid and longer study durations.
Automated Nucleic Acid Purification from Diverse Sample types using dedicated...QIAGEN
This webinar will focus on the automation of QIAGEN’s new line of DNA and RNA sample prep kits for the microbiome. We will show how automation on the QIAcube enables efficient and reliable use of these samples for sensitive downstream applications such as qPCR and NGS. In addition, you will learn how to successfully use the CLC Microbial Genomics Module for metagenome sequencing and identification of microbial composition and diversity.
This document summarizes research on the psychrophilic degradation of long chain alkanes by microorganisms. Key findings include:
1) Pseudomonas frederiksbergensis and Rhodococcus erythropolis were able to degrade n-alkanes from C10-C22 at 4-20°C with optimal degradation by P. frederiksbergensis occurring at 15°C and 7.0 pH.
2) P. frederiksbergensis degraded alkanes via terminal oxidation and detection of metabolites, starting with hydroxylation of alkanes to fatty acids.
3) Genes encoding alkane hydroxylase and alcohol dehydrogenase from P. frederik
The document describes metabolic engineering experiments to optimize Escherichia coli for efficient conversion of glucose to pyruvate. Key modifications included deleting the poxB gene encoding pyruvate oxidase to increase pyruvate production, and decreasing pyruvate dehydrogenase levels through deletion of pdhR and replacing its promoter. Initial experiments successfully deleted poxB but found contamination in strains. Batch cultures showed highest pyruvate concentrations of 6.14 g/L for strain R1 ΔpoxB. While results were mixed, the work provided experience with laboratory techniques and engineering a microbial strain through a full project.
Application Note: A Rapid Procedure for Screening Transuranium Nuclides in Ur...PerkinElmer, Inc.
One of the most extensive tasks is the field of bioassay analysis is the determination of pure alpha- (and beta-) emitting radionuclides from the nuclear fuel cycle such as (234)U and (235)U, or anthropogenic (239)Pu and (241)Am in urine samples. However, any radiochemical method, which is applied to perform such analyses, has to be highly sensitive since even small amounts of incorporated radionuclides decaying by alpha emission may contribute to harmful doses to human organs.
Since alpha radiation has an extremely short penetration length in water and solid substances, direct counting of a salt residue of dry ashed urine is not possible. Therefore, complex radiochemical techniques have been developed for efficient separation of the transuranium elements from the bulk matrix. However, in addition to several purification steps, these methods require the production of almost weightless planar sources (e.g. via electrolytic deposition) in order to perform radioassays with proportional or surface barrier detector. In contrast to the extensive preparative techniques, fast methods using alpha/beta-LSC are of increasing interest. Due to the efficient detection of alpha emitters in a liquid scintillation cocktail, extensive radiochemical purification procedures are not necessary provided the sample is homogeneous in the liquid scintillation cocktail.
(originally aired 11-14-12)
Although biofuels are an attractive alternative to fossil fuels, large scale development is currently challenging. Development of renewable fuel characterization, processes, and contaminant analysis using robust analytical methods is needed. Here, focus is on Ion Chromatography—a proven technique for providing fast, reliable answers during research to production—with HPAE-PAD technology for carbohydrate analysis in feedstock and method parameter optimization (including column chemistry) for efficient separation of mono- and disaccharides with good resolution, linearity, and accuracy over a broad dynamic range. Since some residual sucrose and cellobiose may be present, examples of monitoring them and other saccharides is covered, along with their impact on the fermentation process.
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.
MicroPRO, A Rapid Microbiology Method Based on Flow Cytometryguest32bcc5
This document provides information on disruptive technologies and rapid microbiology products represented by the company. It summarizes their product range including instruments for biological sample preparation, dissolution/formulation, physico-chemistry analysis, and rapid microbiology detection. It also discusses their markets in pharmaceutical, personal care, fermentation, and more. The MicroPRO instrument allows detection of bacteria, yeast and mold from various samples within 24 hours.
21st International Conference Organic Process Research & Development 2010 San...dominev
This document discusses using real-time calorimetry to improve operational efficiency. It presents case studies where ReactIR, FBRM, PVM and RTCal were used:
1) ReactIR developed kinetic models to minimize byproducts in pharmaceutical reactions and improve crystallization processes.
2) FBRM and PVM helped optimize a crystallization to reduce impurities below 0.5%.
3) RTCal validated switching to a low copper acrylamide grade for polymerization, showing a shorter induction period but similar maximum heat output. Real-time calorimetry provided process safety evaluation.
Improving Downstream Processing: Application of Excipients in DSPMilliporeSigma
Webinar summary:
This webinar will showcase the beneficial potential of using excipients during downstream processing of monoclonal antibodies.
Learning points:
In this webinar, you will see:
* An innovative excipient screening approach simulating low pH stress conditions during protein A chromatography and virus inactivation
* How the application of excipients in buffer systems can significantly improve protein stability and chromatographic performance
Abstract:
Key aspects during downstream purification of biopharmaceutical drugs are purity and process yield. Therefore, the downstream process needs to be designed in a way that the final product which will eventually end up in the patient entails low levels of product- and process related impurities (e.g. high molecular weight aggregates) as well as process related contaminants (e.g. host cell protein levels). In addition to this, the process must be capable of clearing and inactivating viruses to ensure product safety. In this webinar, we will explore the benefits of adding excipients during downstream processing on protein stability, chromatographic performance and viral inactivation.
1. The document describes the quality assurance system for sputum smear microscopy in India's Revised National Tuberculosis Control Programme (RNTCP).
2. It establishes a three level laboratory network of National Reference Laboratories, Intermediate Reference Laboratories at the state level, and Designated Microscopy Centers.
3. Quality assurance activities include internal quality control, external quality assessment including on-site evaluations, panel testing of lab staff, and random blinded rechecking of slides to evaluate performance at each level of the network.
PCR Detection of SNP Markers for Warfarin Sensitivity Using Oral Fluids
1. DNA was extracted from oral fluid samples using different methods and quantified, with yields ranging from 3-12 μg/ml. Method 2 was chosen for future work due to simplicity and consistency.
2. PCR amplification of extracted DNA was successful, demonstrating that DNA could be extracted and amplified from oral fluid samples. Inhibitors in oral fluids varied between samples.
3. Primers were designed to detect SNPs associated with warfarin sensitivity. Amplification with LNA primers distinguished between wild-type and mutant samples, showing potential for an oral fluid-based pharmacogenomic test.
The document discusses a presentation on LOINC (Logical Observation Identifiers Names and Codes) given at the 2011 Public Health Informatics conference in Atlanta, GA. The presentation provides an introduction to LOINC and covers topics such as the origins of LOINC, common elements in LOINC terms, LOINC collections like forms and surveys, and domain-specific approaches to mapping standards and terminologies in areas like microbiology. It also discusses LOINC tools and resources for mapping terms and codes.
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
Cowin Biotech is a Chinese biotech company founded in 2007 that provides molecular diagnostic products and services. The company has R&D centers in Jiangsu, Beijing, and Boston and produces over 1,000 products through independent research. Cowin's core capabilities include viral transport media, nucleic acid extraction kits, PCR reagents, and enzymes. It aims to provide a complete solution for clinical diagnostic applications.
ReaMetrix is a life science technology company founded in 2003 with facilities in San Carlos, CA and Bangalore, India. It is committed to developing high-quality, affordable diagnostic solutions for resource-limited settings. ReaMetrix has expertise in assay development, protein expression and labeling, instrumentation, and developing integrated diagnostic solutions. It currently offers products like conjugates for flow cytometry-based tests and is working on projects like affordable point-of-care tests for bacterial detection and disease monitoring.
Introduction to real-Time Quantitative PCR (qPCR) - Download the slidesQIAGEN
This slidedeck introduces the concepts of real-time PCR and how to conduct a real-time PCR assay. The topics that are covered include an overview of real-time PCR chemistries, protocols, quantification methods, real-time PCR applications and factors for success.
This document discusses the polymerase chain reaction (PCR) technique. It describes how PCR was developed by Kary Mullis in the 1980s and allows for the in vitro amplification of specific DNA sequences. The key steps in PCR include denaturation of DNA, annealing of primers, and extension of the DNA strand by a DNA polymerase. The document outlines the main components required for PCR including primers, DNA polymerase, dNTPs, buffer, and magnesium chloride. It also discusses applications of PCR like DNA sequencing, genetic testing, and the Human Genome Project. Real-time PCR is introduced as a variation that allows for detection and quantification of DNA during the PCR process. Diagrams illustrate the PCR process and setup in a thermal cycler machine
The document provides information about upcoming community events and programs in the West End neighborhood of Providence, Rhode Island. It announces the opening of the Sankofa World Market on Wednesdays from July 1st to September 30th at Knight Memorial Library, featuring food, crafts, and resources celebrating the neighborhood's diversity. It also lists dates for resident engagement meetings, homebuyer workshops, photography walks, and POWER workshops. Angie Ankoma, president of the neighborhood organization's board, is awarded the Dorothy Richardson Award for Resident Leadership for her long-time community involvement.
Este documento describe las principales biomoléculas que forman los seres vivos, incluyendo proteínas, carbohidratos, lípidos y ácidos nucleicos. Explica la estructura, función y clasificación de cada biomolécula. También cubre temas como la estructura de las proteínas a nivel primario, secundario, terciario y cuaternario. Finalmente, concluye que las biomoléculas son fundamentales para el funcionamiento normal de los organismos.
Image morphing provides the tool to generate the flexible and powerful visual effect. Morphing depicts the transformation of one image into another image. The process of image morphing starts with the feature specification phase and then proceeds to warp generation phase, followed by the transition control phase. This paper surveys the various techniques available for all three stages of image morphing.
Santanu Chakraborty has over 11 years of experience in electronics, telecommunications, and teaching. He has a diploma in electronics and telecommunications and passed AMIE engineering. He has worked as a service engineer, senior supervisor maintaining machines, and as a faculty member and administrator for various educational institutions. Currently he is a faculty member teaching electronics, electrical, and physics subjects at a technical institution in Kolkata.
Single Jersey Slub is a type of knit fabric made from cotton or cotton blends. It is warm, flexible, stretchy, and insulating, making it popular for layers worn close to the body like t-shirts. Single Jersey Slub has different faces on each side, curls at the edges, clearly visible wales on one side, is twice as extensible widthwise than lengthwise, and can unravel from either side. It provides comfort, shape retention, drape, and crease recovery. Single Jersey Slub is often used for t-shirts and as a substrate for leather or shoe uppers due to its stretch and recovery properties.
Ethnocentric Behaviour and Business Performance of Multinational Enterprises ...IJAEMSJORNAL
This document summarizes a study that examined how ethnocentric behavior of multinational enterprises (MNEs) impacts business performance in Nigeria. The study assessed how foreign culture, use of expatriates, and standardized marketing strategies affected operating costs, employee acceptance, and customer loyalty. It found that an MNE's foreign culture negatively impacted acceptance of management techniques by local employees. It also found that using expatriates in strategic positions increased operating costs, while standardized marketing strategies encouraged customer loyalty in Nigeria. The study recommends incorporating local employees in decisions and guiding MNEs to avoid culture shock to improve effectiveness in host countries.
Site Operation Manual for a Typical Air Monitoring SiteTAMUK
1. Mount the sensor on a mast at least 10 feet above any obstructions.
2. Wire the sensor leads to the datalogger channels for voltage input.
3. Program the datalogger to record the wind speed and direction at the appropriate intervals.
Nagios Conference 2013 - Thomas Dunbar - Building Technology for Storage Syst...Nagios
Thomas Dunbar's presentation on Building Technology for Storage Systems Monitoring.
The presentation was given during the Nagios World Conference North America held Sept 20-Oct 2nd, 2013 in Saint Paul, MN. For more information on the conference (including photos and videos), visit: http://go.nagios.com/nwcna
LabVIEW Based Monitoring the Building in wireless communicationSathish Kumar
The presented wireless system for building monitoring takes advantage of the unique features of
custom-developed MEMS sensors and read-out ASIC combined with an optimized network and module
architecture, to realize a solution which offers long battery lifetime and potentially low cost in
manufacturing, installation and maintenance, while providing high-quality sensor data at the right
time which can be proposed for monitoring building to assess earthquake damage. Low power network
architecture was implemented over an 802.15.4 MAC in the 900-MHz band.A custom patchantenna
was designed in thisfrequency band to obtain robustlinks in real-worldconditions. The modules have
been validatedin a full-scale laboratory setup with simulated earthquakes. This will be illustrated via
LABVIEW software which shows the Temperature level in the time of Earthquake and which helps in
preventing the human life.
Khulisa Management Services- ECD Site Monitoring Instrumentkaleylemottee
This document provides an overview and summary of monitoring conducted of early childhood development conditional grant sites in South Africa between 2003-2004. It finds that:
1) Provincial and district officials were generally satisfied with the training provided and demonstrated relevant learning, though application and impact on organizational performance was only partially achieved.
2) The monitoring and support system was implemented with officials noting it provided structure, but many sites were only visited once and coverage varied significantly between provinces.
3) Provinces generally delivered the conditional grant funds on time, though a minority of sites could not show they received resource kits as intended.
4) Sites that received multiple visits showed progress, with most achieving average or strong performance, though a
Presentation Mrs.Smolka Ursula, Ramboll: costs and benefits when monitoring s...Torben Haagh
Ursula Smolka, Project Engineer at Ramboll in Germany, discusses state of the art load monitoring approaches as well as challenges and benefits of collecting site specific load data for reliability based maintenance as presented at last year’s “Substructures for Offshore UK Wind” conference.
Read the full presentation for free here: http://bit.ly/presentation_Ursulasmolkaramboll
Building and Monitoring Services at LithiumPaul Cichonski
Paul Cichonski's presentation from SF CloudOps Meetup on building and monitoring fault tolerant systems. (http://www.meetup.com/CloudOps/events/159397622/)
The student team tested various aspects of water quality at River Monitoring Site #7, including habitat, macroinvertebrates, total solids, nitrates, phosphates, pH, and dissolved oxygen. The habitat scored poorly due to lack of pools and narrow riparian vegetation. Macroinvertebrate testing found the river's health to be poor. Total solids and phosphates were high, while nitrates and pH were acceptable. Dissolved oxygen was moderate. To improve the river's quality, the team proposes dredging sediment, establishing an "Adopt a River" program, and restricting certain fertilizers. Costs of dredging could be offset through beach fees, equipment rentals, and fertilizer sales
This document discusses using quantitative PCR (qPCR) to analyze microbial communities at contaminated sites. qPCR allows quantification of bacteria, genes, and activities important for bioremediation. It provides information on monitored natural attenuation and whether biostimulation or bioaugmentation is needed. The document summarizes qPCR analysis of 869 wells at 56 sites, finding correlations between Dehalococcoides abundance, reductive dehalogenase genes, and ethene production. The highest ethene formation occurred with near-equal ratios of VC reductase genes to Dhc cells. Developing high-throughput qPCR arrays allows monitoring many targets from multiple samples simultaneously and cost-effectively.
This document provides an overview of DNA and RNA isolation, purification, and amplification techniques. It discusses preparing genomic DNA from bacterial cells using lysis and phenol-chloroform extraction. Plasmid DNA can be purified from bacterial cells using alkaline lysis and silica binding. The document also reviews RNA isolation, which requires rapid processing to prevent RNase degradation, and describes mRNA selection using oligo-dT cellulose. Reverse transcription PCR and quantitative PCR are introduced as methods to study gene expression.
1. CENTRAL DOGMA OF MOLECULAR BIOLOGY
2. NUCLEIC ACID PREPARATION & APPLICATIONS
3. FUNDAMENTAL STEPS IN DNA PURIFICATION
4. ANALYSIS OF NUCLEIC ACIDS
5. STORAGE CONDITIONS
Over the past decade, there have been a growing number of mAb candidates entering the clinical pipeline. This results in a large increase on the demand for analytical characterization. This seminar discusses advances in analytical method development with analytical run times below 10 minutes for all routine methods with intelligent, integrated chromatography workflows. Orbitrap technology has been established as the most powerful MS technology for protein characterization. How this can be incorporated into a complete workflow for bio-pharma analysis is also discussed.
Principle, Procedure and applications of Digital PCR.pptxVikramadityaupmanyu
Digital PCR (dPCR) is the new generation PCR that enables absolute quantification of target gene by separating reac¬tion mixture in several compartments. In this system, copies of target nucleic acid are distributed randomly from 0, 1 or many in the several small volume compartments. Amplification is occurred in the compartment and resulting absorbance is measured. Integrated fluidic circuits, chip based microchambers, and water in oil droplets are the methods are used for separation of reaction mixture in to several compartments. BioMark HD system (Fluidigm, USA) and QuantStudio3D system (Thermofisher Scientific, USA) uses integrated fluidic circuits harbor¬ing 10000 to 40000 microchambers and integrated chip containing 20000 microchambers, respectively. In droplet digital PCR System (Biorad, USA), reaction mixture is separated into 20,000 water in droplets. After cycles of reaction using any of the above technologies, fluorescence is detected with an imaging sys¬tem in the each compartment and copy numbers of the target is calculated with imaging software.
In droplets digital PCR, after amplification, droplets containing target gene are detected by fluorescence and scored as positive, and droplets without fluorescence are scored as negative. Poisson statistical analysis of the numbers of positive and negative droplets yields absolute quantitation of the target sequence. Digital PCR is the preferred technique for absolute quantification of target gene without need of standard curve and higher sensitivity and produces highly reproducible results, and also less susceptible to inhibitors than conventional RT-qPCR
Clinical isolates of urinary tract infection and candidiasis inhibited by T. ...Diganta Dey
This document summarizes a study that evaluated the antimicrobial activity of extracts from Terminalia arjuna bark. Methanol and water extracts were tested against bacterial and fungal clinical isolates. The methanol extract showed higher total phenolic and flavonoid content. Both extracts inhibited the growth of pathogenic bacteria like S. aureus and fungi like Candida species. Minimum inhibitory concentration values ranged from 0.04-1 mg/mL. Comet assay results also indicated that the extracts induced DNA damage in Candida tropicalis. Overall, the study demonstrated the antibacterial and antifungal properties of T. arjuna bark extracts.
Multi-residue pesticide analysis of food samples using acetonitrile extractio...Kate?ina Svobodov
This document describes a method for multi-residue pesticide analysis of food samples using two-dimensional liquid chromatography coupled with tandem mass spectrometry (2D-LC-MS/MS). The method involves acetonitrile extraction of samples followed by separation using reverse phase and HILIC columns connected by a switching valve. The method showed improved peak shape and sensitivity for polar pesticides compared to single column methods. Recoveries of 79.67% of analytes were between 60-100% and reporting limits were 1 ppb or lower for most pesticides tested, demonstrating this 2D-LC-MS/MS method is effective for broad-scope pesticide residue testing in foods.
This document provides a directory of services for BestCare Laboratory Services, LLC. It outlines policies and procedures for specimen collection, processing, storage, and transport. Key points include:
- Proper patient preparation, specimen collection techniques, handling, and storage are essential for accurate test results.
- Specimen collection follows a specific order including verifying patient identity, using the correct collection container and additive, and proper labeling.
- The laboratory offers a variety of routine and stat testing as well as specialized services like microbiology and therapeutic drug monitoring.
- Detailed guidelines are provided for collection of different specimen types like blood, urine, stool, and cultures to ensure specimen integrity.
This document provides an overview of key water quality parameters for recirculating aquaculture systems. It discusses parameters like dissolved oxygen, temperature, pH, ammonia, nitrite, and nitrate. It explains their interactions and ideal concentrations for fish health. It also covers measurement techniques for these parameters as well as other important water quality metrics like alkalinity, hardness, carbon dioxide, and solids. Measurement methods include test kits, colorimetric analysis, and Winkler titration for dissolved oxygen. Maintaining optimal conditions for these water quality parameters is essential for fish health and productivity in recirculating aquaculture.
This document provides an overview of key water quality parameters for recirculating aquaculture systems including dissolved oxygen, temperature, pH, ammonia, nitrite, nitrate, carbon dioxide, alkalinity, and solids. It discusses their interactions and optimal ranges for fish health. Measurement techniques including titration, colorimetry, and ion-specific electrodes are presented. The importance of proper chemical analysis for monitoring water quality is emphasized.
This document provides an overview of key water quality parameters for recirculating aquaculture systems including dissolved oxygen, temperature, pH, ammonia, nitrite, nitrate, carbon dioxide, alkalinity, and solids. It discusses their interactions and optimal ranges for fish health. Measurement techniques including titration, colorimetry, and ion-specific electrodes are presented. The importance of proper chemical analysis for monitoring water quality is emphasized.
This document provides an overview of key water quality parameters for recirculating aquaculture systems including dissolved oxygen, temperature, pH, ammonia, nitrite, nitrate, carbon dioxide, alkalinity, and solids. It discusses their interactions and optimal ranges for fish health. Measurement techniques including titration, colorimetry, and ion-specific electrodes are presented. The importance of proper chemical analysis for monitoring water quality is emphasized.
The document discusses polymerase chain reaction (PCR) and its use for detecting COVID-19 through RT-PCR testing. RT-PCR is considered the gold standard for detecting viruses like SARS-CoV-2 due to its rapid detection, high sensitivity, and specificity. The document outlines the RT-PCR testing process, which involves collecting a sample, extracting RNA from the sample, converting the RNA to cDNA, and amplifying the cDNA using fluorescent dyes to detect the presence of the virus. Real-time PCR allows visualization and quantification of PCR product accumulation in real-time by using fluorescent dyes that bind to DNA.
This document provides an overview of reverse transcription polymerase chain reaction (RT-PCR). It explains that RT-PCR is used to detect and quantify mRNA by first converting it to cDNA using reverse transcriptase. There are two main approaches to RT-PCR - a one-step approach where reverse transcription and PCR occur in the same tube, and a two-step approach where they occur in separate tubes. The document outlines the basic protocol for each approach and discusses some technical issues and applications of RT-PCR, such as distinguishing between infectious and non-infectious viruses.
This document provides an overview of reverse transcription polymerase chain reaction (RT-PCR), including its objectives, introduction, history, principle, protocol for one-step and two-step RT-PCR, technical issues, and literature applications. RT-PCR is a technique that allows detection and quantification of mRNA by first converting RNA to cDNA using reverse transcriptase, then exponentially amplifying the cDNA using PCR. It is often used to detect gene expression and distinguish between infectious and non-infectious viruses or variants in samples. Care must be taken to prevent contamination during sample preparation and RT-PCR.
This document provides an overview of key water quality parameters for recirculating aquaculture systems. It discusses parameters like dissolved oxygen, temperature, pH, ammonia, nitrite, and nitrate. For each parameter, it explains their importance and how they interact with each other. It also summarizes different types of recirculating systems and provides water quality standards. Measurement techniques for various parameters and chemical analysis methods are also outlined.
HIV Vaccines Process Development & Manufacturing - Pitfalls & PossibilitiesKBI Biopharma
Originally presented at the HIV Vaccine Manufacturing Workshop –July 19th& 20th, 2017 by Abhinav A. Shukla, Ph.D.Senior Vice PresidentDevelopment & ManufacturingKBI Biopharma, Durham NC
Flyer of our new product Cleanpid® Easy Purification Kit for detecting Legionella. Cleanpid is for purification the sample before doing the PCR. After using CLEANPID you can collect the Legionella cells and with freeze/thaw cycles you can obtain the DNA and with PCR, the customer could say if there is or not Legionella presence.
Legionellae DNA extraction is particularly problematic when different types of microorganisms may be present.
Cleanpid® Easy Purification kit is the simple method for the separation and purification of Legionella in water samples. The method is based on Legionella binding magnetic beads for capturing cells (immunomagnetic separation).
No columns or centrifugations are necessary.
A summary of the advantages of Cleanpid is:
Quick and Easy-To-Use
The kit only requires a magnetic rack (we provide it)
Improves the sensitivity and reproducibility of qPCR
Reduction of false positive rate for PCR:
o Detects only legionella viable cells (not exogenous DNA, not other microbiota...)
Reduction of false negative rate for PCR:
o Obtains a larger amount of DNA from the water samples (avoid dilution of the DNA samples for eliminating inhibitors)
The purified organism is additionally suitable for DNA extraction by simply freeze/thaw cycles (simply and cost-efficient lysis)
http://www.biotica.es/en/Products/Cleanpid%20Easy%20Purification%20kit%20for%20Legionella
Kenyatta university. dna extration docxLando Elvis
1. The document describes a HotSHOT DNA extraction method to isolate DNA from fish fins. The method uses sodium hydroxide (NaOH) to lyse cells and release DNA. The NaOH treatment denatures the DNA. Tris buffer is then used to neutralize the solution and renature the DNA.
2. The method involves homogenizing fish fins, treating with hot NaOH to lyse cells and denature DNA, cooling and neutralizing with Tris buffer. Centrifugation pellets debris and collects DNA in the supernatant.
3. The HotSHOT method provides a rapid, inexpensive way to obtain PCR-quality DNA from fish fins compared to traditional methods. The isolated DNA is suitable for
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Unveiling the Dynamic Personalities, Key Dates, and Horoscope Insights: Gemin...my Pandit
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The APCO Geopolitical Radar - Q3 2024 The Global Operating Environment for Bu...APCO
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Anny Serafina Love - Letter of Recommendation by Kellen Harkins, MS.AnnySerafinaLove
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How are Lilac French Bulldogs Beauty Charming the World and Capturing Hearts....Lacey Max
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How to Implement a Real Estate CRM SoftwareSalesTown
To implement a CRM for real estate, set clear goals, choose a CRM with key real estate features, and customize it to your needs. Migrate your data, train your team, and use automation to save time. Monitor performance, ensure data security, and use the CRM to enhance marketing. Regularly check its effectiveness to improve your business.
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Introduction
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2. Why
do
we
need
MBTs?
Contaminant
concentra,ons
and
geochemistry
don’t
always
provide
the
complete
picture.
Plate
counts
do
not
accurately
reflect
in
situ
microbial
community
<
1
%
of
bacteria
can
be
cultured
in
the
laboratory
3. Ques(ons
that
MBTs
can
answer
What
is
the
concentra,on
of
contaminant
degraders?
qPCR
QuantArray
Is
biodegrada,on
occurring?
Stable
Isotope
Probing
(SIP)
Compound
Specific
Isotope
Analysis
(CSIA)
What
microorganisms
are
present?
Next
Genera,on
Sequencing
(metagenomics)
What
treatment
strategy
should
be
selected?
In
Situ
Microcosms
(ISMs)
4. CENSUS®
qPCR
and
QuantArray®
What
is
the
concentra(on
of
contaminant
degraders?
5. • qPCR
Amplifica,on
– Primers
&
probe
bind
to
target
gene
– Fluorescence
signal
increase
propor,onal
to
concentra,on
• Two
main
types
of
target
genes
– Taxonomic
(16S
rRNA
gene)
– Func,onal
(Reductases,
oxygenases)
qPCR
Basics
Rapidly
detect
and
quan,fy
a
target
gene
or
microbial
popula,on
12. Es(ma(ng
Cometabolism
Contribu(on
0.01
0.1
1
10
1.0E+00
1.0E+01
1.0E+02
1.0E+03
1.0E+04
1.0E+05
TCE
Degrada(on
Rate
Constant
(per
year)
PHE
(gene
copies/mL)
PHE
is
ND
(X)
or
Low
PHE
is
Average
PHE
is
High
Significant
Degrada(on
No
Degrada(on
Some
Degrada(on
Faster
Degrada(on
ESTCP
ER-‐201584
13. • Former
chemical
manufacturing
facility
• Superfund
site
• Groundwater
impacted
by
– Chloroethanes
– Chloroethenes
– Chloropropanes
QuantArray®-‐Chlor
Case
Study
14. Site
Management
Ques(ons
qPCR
Is
complete
reduc,ve
dechlorina,on
likely?
Should
an
electron
donor
be
added?
Was
electron
donor
injec,on
effec,ve?
Is
bioaugmenta,on
needed?
What
is
the
concentra,on
of
contaminant
degraders?
qPCR
QuantArray
26. Low
vinyl
chloride
concentra(ons
(<5
µg/L)
Rescaled
Y
axis
TOC
decreases
to
Non-‐Detect
DHC
con(nues
to
decrease
with
consump(on
of
e-‐donor
Vinyl
chloride
detected
27. • Effec,ve
adsorp,on
and
biodegrada,on
– Dehalococcoides
is
an
obligate
halorespiring
microbe
– Dehalococcoides
decreased
when
e-‐
donor
was
consumed
– Daughter
products
only
detected
when
Dehalococcoides
had
likely
dropped
to
low
concentra,ons
• Microbial
monitoring
cri,cal
aoer
Ac,vated
Carbon
– Daughter
products
not
detected
during
biodegrada,on
– Daughters
only
detected
aoer
biodegrada,on
slowed
(e-‐
donor
consumed
and
redox
condi,ons
less
favorable)
Conclusions
29. Next
Genera(on
Sequencing
What
microorganisms
are
present?
Next
Genera,on
Sequencing
(metagenomics)
Microbial
Insights
EMD
Webinar
Series
hmp://www.microbe.com/webinars/
Metagenomics
&
Next
Genera(on
Sequencing:
How
to
Make
the
Most
of
Your
Data
without
Jumping
to
Conclusions
30. “Sequencing”
“Sequencing”
Amplicon
Sequencing
(16S
rRNA
gene)
Specific
target
region
is
amplified
to
improve
coverage
level
Who
is
there?
(Taxonomy
Classifica,on)
31. What
do
you
get?
Sample
ID
Reads
Passing
Quality
Filtering
%
Reads
Classified
to
Genus
Shannon
Genus
Diversity
MW6
607,795
92.2%
3.0
MW7
577,170
93.6%
2.9
MW8
719,650
93.7%
2.3
MW9
736,200
94.1%
2.3
MW10
734,080
93.6%
2.7
99.6%
97.8%
97.3%
95.4%
94.5%
92.2%
49.2%
0%
20%
40%
60%
80%
100%
Kingdom
Phylum
Class
Order
Family
Genus
Species
%
Total
Reads
Classified
32. Top
Genus
Classifica(on
Results
Classifica(on
Number
of
Reads
%
Total
Reads
Descrip(on
Dechloromonas
146,290
24.1%
Faculta,ve
anaerobic
bacteria
(uses
oxygen
as
electron
acceptor
when
available).
Some
strains
u,lize
nitrate
as
an
electron
acceptor
and
some
can
reduce
perchlorate
and
chlorate.
Geobacter
108,799
17.9%
Anaerobic,
gram-‐nega,ve,
iron
reducing
bacteria.
Some
species
can
also
reduce
sulfur.
Unclassified
at
Genus
Level
74,511
12.3%
Pseudomonas
26,248
4.3%
Pseudomonas
is
a
metabolically
diverse
genus
of
aerobic
organisms.
Some
species
can
also
denitrify.
Some
strains
use
common
hydrocarbons
as
carbon
sources.
Rhodoferax
25,011
4.1%
anaerobic
genus
that
oxidizes
acetate
with
the
reduc,on
of
Fe
(III).
Gallionella
23,727
3.9%
Aerobic,
iron
oxidizing
bacteria
Sulfuritalea
18,234
3.0%
Genus
of
faculta,ve
anaerobes
bacteria
(uses
oxygen
as
electron
acceptor
when
available)
that
also
reduce
nitrate.
Grows
chemolithoautotrophically
by
oxida,on
of
reduced
sulfur
compounds
and
hydrogen
under
anoxic
condi,ons.
Heterotrophic
growth
on
organic
acids.
Methylotenera
16,927
2.8%
Facultative
methylotrophs
that
utilize
methylamine.
Some
may
utilize
methanol,
ethanol
and
pyruvate.
37. Ques(ons
MBTs
can
answer
Is
biodegrada,on
occurring?
Stable
Isotope
Probing
(SIP)
Compound
Specific
Isotope
Analysis
(CSIA)
Microbial
Insights
EMD
Webinar
Series
hmp://www.microbe.com/webinars/
CSIA
vs.
SIP
What
is
the
difference
and
how
do
I
use
them?
39. • As
organic
compounds
degrade,
the
ra,o
of
stable
isotopes
(13C/
12C,
2H/H,
37Cl/35Cl)
in
the
frac,on
remaining
aoer
degrading
can
change
in
a
predictable
way.
• CSIA
can
provide
a
conserva,ve
boundary
on
the
extent
of
degrada,on
EPA
Guidance
40. Unit
of
measure
Amount
of
13C
rela,ve
to
12C
is
expressed
by
the
δ13C
nota,on
The
standard
is
a
specific
carbon-‐containing
mineral
from
a
specific
loca,on:
Pee
Dee
Belimnite
(PDB)
Units
of
δ13C
are
o/oo
or
“per
mill”
[ ] 10001
)/(
)/(
‰
Standard
1213
Sample
1213
13
⋅
⎟
⎟
⎠
⎞
⎜
⎜
⎝
⎛
−=
CC
CC
Cδ
41. • Chemical
bonds
with
the
lighter
isotope
(12C)
are
slightly
weaker
than
those
formed
with
the
heavier
isotope
(13C)
and
react
more
quickly.
• The
parent
compound
becomes
enriched
in
the
heavier
isotope
(increasing
δ13C).
• The
daughter
product
is
ini,ally
very
depleted
in
the
heavy
isotope
(lower
or
“more
nega,ve”
δ13C).
CSIA
–
Why
it
works
42. 13Chocolate
Frac(ona(on
Decreasing
total
M
&
M’s
Decreasing
ra,o
M
:
M
(Increasing
ra,o
M
:
M)
12C
13C
Time
46. • Specially
produced
“heavy”
compounds
which
are
composed
of
99+%
13C
– Natural
compounds
are
99%
12C
– Same
characteris,cs
as
original
compound
– Behave
similar
to
the
natural
compound
• Used
as
a
“probe”
or
“tracer” to
determine
if
biodegrada(on
is
occurring
– If
biodegrada,on
occurs,
the
13C
will
be
incorporated
biomass
or
mineralized
to
13CO2.
Stable
Isotope
Probes
47. Overview
of
Bio-‐Trap
SIP
Approach
13C labeled
Benzene
Beads loaded
with 13C
compound
Bio-Trap® with 13C-
benzene loaded
beads
In-Situ deployment
in monitoring well
Beads
analyzed
following
deployment
48. • Passive microbial sampling tool
• Colonized by active microbes
• 25% Nomex and 75% PAC
• Used in conjunction with
– Stable isotope probing
– qPCR and QuantArray
– Other MBTs
What
Are
Bio-‐Trap®
Samplers?
49. Bio-‐Trap
SIP
Analysis
Residual
13C-‐Compound
13C/12C
Dissolved
Inorganic
Carbon
13C/12C
of
Biomarkers
U(liza(on
Mineraliza(on
(C
for
energy)
Metabolism
(C
for
growth)
PLFA
DNA
RNA
50. ü Contaminant
concentra,ons
ü Geochemistry
• Molecular
Biological
Tools
MNA
Assessment
-‐
SIP
Case
Study
Concentra,ons
of
contaminant
degrading
microorganisms?
Is
biodegrada,on
occurring?
Stable
Isotope
Probing
(SIP)
QuantArray
&
qPCR
51. Study
Wells
–
Weathered
Limestone
Well
Naphthalene
2-‐Methylnaphthalene
UMW-‐7C
13
1,000
Well
Naphthalene
2-‐Methylnaphthalene
UMW-‐44
15
100
Well
MMW-‐17D
52. Is
naphthalene
biodegrada(on
occurring?
-‐50
0
50
100
150
200
250
Background
UMW-‐7C
DIC
δ13C
(‰)
13C
naphthalene
mineralized
to
CO2
UMW-‐7C
55. MNA
Assessment
Chemical
Microbiological
Decreasing
contaminant
concentra,on?
Stable
Isotope
Probing
Did
biodegrada,on
occur?
QuantArray
Concentra,ons
of
contaminant
degraders?
Naphthalene
56. • Conclusive
evidence
of
in
situ
biodegrada,on
• Don’t
need
to
know
organisms
or
pathways
involved
• Broad
applicability
(carbon
and
energy
sources)
– BTEX,
MTBE,
TBA
– Naphthalene
– Chlorobenzene
– Emerging
contaminants
(dioxane,
sulfolane)
• Inexpensive
for
many
common
contaminants
SIP
Strengths
57. Links
–
SIP
&
In
Situ
Microcosms
(ISMs)
-‐500
0
500
1000
1500
2000
2500
3000
Average
Background
MNA
ORC
Advanced
PLFA
Del
(‰)
13C
Incorpora(on
into
Biomass
59. In
Situ
Microcosms
(ISMs)
What
treatment
strategy
should
be
selected?
In
Situ
Microcosms
(ISMs)
Control
(MNA)
Treatment
Op,on
1
Treatment
Op,on
2
60. Unit
Samplers
Assembly
Control
(MNA)
Treatment
Op,on
1
Treatment
Op,on
2
GEO
COC
Bio-‐Trap
Supplier
Supplier
61. • Shallow
aquifer
impacted
by
TCE.
• Daughter
product
cis-‐1,2
dichloroethene
(DCE)
has
been
detected.
• DCE
appears
to
be
accumula,ng
(“DCE
stall”)
• Considering
– Bios,mula,on
(BioS,m)
with
electron
donor
– Bioaugmenta,on
(BioAug)
w/culture
and
electron
donor
Site
Background
62. ISM
Study
–
Microbial
Lines
of
Evidence
qPCR
MNA
(Control)
Are
halorespiring
bacteria
(e.g.
Dehalococcoides)
present?
BioS(m
Will
electron
donor
addi,on
s,mulate
growth
of
halorespiring
bacteria?
Bioaugmenta,on
needed?
BioAug
Will
a
bioaugmenta,on
culture
survive?
63. ISM
Study
–
Chemical
Lines
of
Evidence
qPCR
MNA
(Control)
BioS(m
BioAug
Contaminant
concentra,ons
under
exis,ng
condi,ons?
Will
electron
donor
addi,on
enhance
daughter
product
forma,on?
Ethene?
Full
dechlorina,on?
Will
bioaugmenta,on
enhance
biodegrada,on
compared
to
electron
donor
alone?
65. ISM
Study
–
Microbial
Lines
of
Evidence
qPCR
MNA
(Control)
Are
halorespiring
bacteria
(e.g.
Dehalococcoides)
present?
Yes,
but
at
a
low
concentra,on
BioS(m
Will
electron
donor
addi,on
s,mulate
growth
of
halorespiring
bacteria?
Yes,
a
noteworthy
increase
was
observed
Bioaugmenta,on
needed?
Probably
not
BioAug
Will
a
bioaugmenta,on
culture
survive?
Yes,
DHC
remained
high
during
the
deployment
period
67. ISM
Study
–
Chemical
Lines
of
Evidence
qPCR
MNA
(Control)
BioS(m
BioAug
Contaminant
concentra,ons
under
exis,ng
condi,ons?
Mainly
TCE
(60%)
with
DCE
(40%)
–
no
vinyl
chloride,
ethene
Will
electron
donor
addi,on
enhance
daughter
product
forma,on?
Yes,
enhanced
DCE
produc,on
(90%)
Ethene?
Full
dechlorina,on?
Yes
Will
bioaugmenta,on
enhance
biodegrada,on
compared
to
electron
donor
alone?
Yes
but
not
substan,ally
68. Site
Management
Decision
Overall
Ques,on
MNA
or
Bios,mula,on
or
Bioaugmenta,on?
Bios,mula,on
Client’s
Ac,on
69. • Chlorinated
hydrocarbon
degrada,on
– Enhanced
anaerobic
biodegrada,on
• Petroleum
hydrocarbons
– BTEX,
MTBE
and
TBA
Links
-‐
ISM
70. A
litle
info
about
Microbial
Insights
Founded
in
1992
as
a
technology
transfer
company
based
on
the
research
of
Dr.
D.C.
White
at
the
University
of
Tennessee
71. • Experience
• Accuracy,
precision
and
quality
control
• Innova,on
– Comprehensive
suite
of
MBT
analyses
– Microbial
Insights
Database
– QuantArray
&
con,nuous
assay
development
– Next
Genera,on
Sequencing
• Customer
Service
A
litle
more
about
Microbial
Insights
72. • www.microbe.com
• Contacts
– Kate
Clark
(kclark@microbe.com)
– Casey
Brown
(cbrown@microbe.com)
• Telephone
(865)
573-‐8188
For
more
informa(on