In this webinar, you will learn: - The challenges tied to contamination control within a biopharmaceutical environment. - What closed processing is, and how sampling solutions are an integral component towards that end. - Advantages of sterile sampling from both a technical and economical viewpoint; with the review of a technical study confirming contamination risk reduction and total cost of ownership. - Recommendations and requirements stated by these major regulatory authorities around the monitoring of the manufacturing process with the execution of sampling. Detailed description: Biopharmaceutical manufacturers are required to ensure drug product quality attributes for patient safety. Strong contamination control strategies should be considered early in process design, and have direct influence on the production environment and equipment selection. Sampling at each step is a critical component in maintaining a contamination control strategy. Regulators are critical in the sampling process, as it predicts the state of the product or process, and needs to be Representative. A case study will be presented that demonstrates a closed, robust sampling solution capable of maintaining a sterile flow path when challenged with Brevundimonas diminuta. The sampling option you select can help support your goal in achieving a closed process, improving your risk mitigation strategy and product safety.

1. The life science business of Merck KGaA,
Darmstadt, Germany operates as
MilliporeSigma in the U.S. and Canada.
Aseptic Process Sampling to address Risk of
Contamination & Containment in compliance to
Regulatory
Somasundaram G.
Senior Technical Consultant, Asia Pacific, Process Solution
Marc Antoine Kaag
Global Product Manager, Sterile Sampling, Process Solutions
August 6, 2020

2. The life science business
of Merck KGaA, Darmstadt, Germany
operates as MilliporeSigma in the U.S.
and Canada

3. Agenda
1
3
4
Contamination control strategy & closed processing
Microbial ingress challenge and outcomes
Sterile sampling value assessment
2 Sampling & Regulatory Requirements

4. Contamination control strategy
& Closed processing

5. Poll question
Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.20205

6. Aseptic processing – A path full of pitfalls…
6
Contamination control strategy & closed processing
1 Highly trained personnel
2 A risk-based approach to contamination
control and a dedicated strategy
3 Resulting in significant costs:
 Infrastructure
 Day-to-day operations
Biopharma manufacturing operations
require substantial precautions to prevent
microbial ingress:

7. Objectives of a contamination control strategy
7
Contamination control strategy & closed processing
Requirement:
“A contamination control strategy should be implemented across the facility in order to assess the
effectiveness of all the control and monitoring measures employed’’.
Implications:
• Lead to corrective and preventative actions if necessary
• Consider all aspects of contamination control and associated lifecycle
• Periodic review based on an appropriate frequency
• Particularly of utmost importance in multi-product facilities
Eudralex Volume 4, Annex 1 “Manufacture of Sterile Medicinal Products”

8. Contamination control strategy: A holistic process
Contamination control strategy & closed processing
Assess
Control and
Monitor
Identify
8
Microbiological
Chemical
Particulate
Cross-product
contamination
1
3
2

9. Key considerations for implementing an effective strategy
Contamination control strategy & closed processing
9
Process design
Designed to protect the
product against contamination
Facility design
Fit with the type of product
to be manufactured
Process simulation
Media fill
Environmental monitoring
program
Predictive system to prevent
product contamination event
Personnel training and
gowning
Primary source of
contamination!!
1
25
34
6
Cleaning and disinfecting
Use a risk-based approach
to build the plan
Inappropriate sampling operations can lead to manufacturing process
breaches and medicinal product/operator/environment contaminations

10. 10
30
Percent
1-6Months
1-14Million Euro
Percent of process deviations
caused by contamination*
Length of time to complete
an investigation
Operations cost
*Sources Langer 2013, Wiebe 2014
Biologics in-process contamination

11. 11
30
Percent
1-6Months
1-14Million Euro
*Sources Langer 2013, Wiebe 2014
Biologics in-process contamination
Impact
 Productivity losses
 Material replacement costs
 Batch loss
 Interruption of product supply
 Delay in clinical development

12. 12
Closed processing – The future of biomanufacturing
Contamination control strategy & closed processing
Closed process definition
 A process condition when the product, materials, critical
components or container/closure surfaces are contained and
separated from the immediate process environment within
closed/sealed process equipment. A process step (or system)
in which the product and product contact surfaces are not
exposed to the immediate room environment.’’
ISPE (International Society for Pharmaceutical Engineering)
Advantages
 Risk of contamination greatly reduced
 Operations can take place in lower classified environments
 Operating time & cost optimizations
Sterile sampling helps to reach this final state

13. Sampling & Regulatory
Requirements

14. 14
 Representative Sample :
 Composition
 Non-operator dependent
 Repeatable
 Protect the Process (Contamination-free)
 Protect the Person (Contained)
 Protect the Product
 Easy of Use
 Save Time and Cost Efficient
Introduction to sampling
What are The Requirements and Challenges?
Process
People Sample
Validated
Sampling
Procedure
Validated
Sampling
Procedure
Equipment
In today’s biopharmaceutical market, sampling is critical during every step in
the manufacturing process. An imprecise or false positive result can lead to a
quarantine as well as the need to repeat the analysis.

15. 15
Drivers for Aseptic Process Sampling
Sampling in Bioprocesses
Bioreactor
Seed train
Buffer/Media
Preparation
Bioreactor
Production
Purification Sterile
FIltration
pH     
Conductivity/ Osmol     
Cells  
Metabolites  
Protein analysis   
Bioburden & Archiving     
Endotoxin     
Other (virus, by products)     

16. Aseptic Filler
Sterile Hold
Tank
Vent
Filter
Vent
Filter
Bioburden
Reduction
Filter
Sterilizing
Filter
Sterilizing
Filter
Prefiltered
Formulation
Aseptic Filler
Sterile Hold
Tank
Vent
Filter
Vent
Filter
Bioburden
Reduction
Filter
Sterilizing
Filter
Sterilizing
Filter
Vent
Filter
Sterilizing
Filter
Prefiltered
Formulation
Sampling point
Final Filling
Single SGF
Redundant
Filtration
Drivers for Aseptic Process Sampling

17. 17
Open Sampling
Valve
Steam in place
valve
Septum sampling Aseptic Connectors Tube Welding
Large number of
samples
Low cost per sample
Large number of
samples
Closed sampling
Low cost per sample
Large number of
samples
Low cost per sample
Flexible & Reliable
Aseptic sampling
Safe and disposable
Large number of
samples
Aseptic sampling
Flexible
Dead-leg
Loss of product (flush)
Open sampling
Impossible to sterilize
Complex operation (SIP)
Safety hazard (heat)
Risk of sample
dilution
Container limitations
Not steam sterilizable
Safety hazard (needles)
Sample volume
limitation
Extra cost
Potential dead-leg
(tubing)
Limited disconnection
Waste of product
Requires utility
Piece of hardware
High risk of
contamination
Operator & Process
safety
Sample
representativeness
Operator training &
safety
Sample
representativeness
High risk of
contamination
Operator safety
Sample
representativeness
Operator training
Requires
maintenance Operator
training
What are the Ways to Sample?
Traditional sampling: Pros, Cons and Limitations

18. 18
2. Sampling process
2.3 Storage and retention
“The container used to store a
sample should not interact
with the sampled material
nor allow contamination.
[…]. As a general rule the
container should be sealed
and preferably tamper-evident.”
Regulatory requirements
Sampling Process and Precautions
Annex 4, WHO guidelines for sampling of pharmaceutical products and related materials
2. Sampling process
2.1 Preparation for sampling
”All sampling tools and implements
should be made of inert materials
and kept scrupulously clean. […].
The cleaning procedure used for all
sampling tools and implements
should be documented and
recorded. […]. The use of
disposable sampling materials
has distinct advantages.”
2. Sampling process
2.2 Sampling operation and
precautions
“There should be a written
procedure describing the
sampling operation. This should
include details of the health and
safety aspects of sampling. It
should ensure that
representative samples are
taken in sufficient quantity for
testing in accordance with
specifications.”

19. 19
FDA cGMP for phase 1 drugs: “recommends the use of closed system to minimize the risk of
contamination”
Guidance for the industry investigational drugs section
F. Laboratory Controls / 1. Testing
“We recommend that the sample consist of a quantity adequate to perform additional testing or
investigation if required at a later date […]. We recommend that you appropriately store and retain
the samples for at least two years […].”
Q7A GMP guidance for manufacturing API – section C. In-process Sampling and Controls (8.3)
“In-process sampling should be conducted using procedures designed to prevent contamination […].
[…] ensure the integrity of samples after collection.”
Regulatory requirements
FDA cGMP: Sampling of Pharmaceutical Products and Related
Materials

20. 8.94 Bioburden samples should
be taken from the bulk product
and immediately prior to the final
sterile filtration. Systems for taking
samples should be designed so as
not to introduce contamination. 9.7 Sampling methods should
not pose a risk of contamination
to the manufacturing
operations.
7.3 Non-essential processes
such as product inspection
and in process testing should be
conducted à with the crimping
option there is a validated and easy
disconnection in place to further
treat the sample
outside the clean areas wherever
possible.
Annex 1 EU GMP (Draft)
20
Regulatory requirements

21. Poll question
Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.202021

22. Microbial ingress challenge and
outcomes

23. 23
Introduction to NovaSeptum® sampling system
 Closed design to sample from aseptic and sterile processes
 Different connection types to the process (Tri-clamp, Ingold®,
In-line)
 Sample is isolated from point of sampling to analysis in
disposable containers (bags, bottles, and syringes)

24. 24
Introduction to NovaSeptum® sampling system
 Closed design to sample from aseptic and sterile processes
 Different connection types to the process (Tri-clamp, Ingold®,
In-line)
 Sample is isolated from point of sampling to analysis in
disposable containers (bags, bottles, and syringes)

25. 25
Introduction to NovaSeptum® sampling system
 Closed design to sample from aseptic and sterile processes
 Different connection types to the process (Tri-clamp, Ingold®,
In-line)
 Sample is isolated from point of sampling to analysis in
disposable containers (bags, bottles, and syringes)

26. 26
Introduction to NovaSeptum® sampling system
 Closed design to sample from aseptic and sterile processes
 Different connection types to the process (Tri-clamp, Ingold®,
In-line)
 Sample is isolated from point of sampling to analysis in
disposable containers (bags, bottles, and syringes)

27. Multiple actuation microbial ingress testing
27
Microbial ingress challenge and outcomes
Purpose
 Determine if multiple actuations of the trigger system can be performed while maintaining microbial
integrity.
Study Scope
• NovaSeptum® sampling system
 1 and 2 mm gauge sampling needles
• NovaSeptum® GO sampling system
 2 mm gauge sampling needle
• Gamma and beta-irradiated products
Challenge Organism
 Brevundimonas diminuta (on average 0.3 by 0.7 µm in size)
Assay
 Presence/absence of microbial growth (turbidity) in medium flow-through with bacterial identification
confirmation

28. Multiple actuation microbial ingress testing
28
Microbial ingress challenge and outcomes
Purpose
 Determine if multiple actuations of the trigger system can be performed while maintaining microbial
integrity.
Study Scope
• NovaSeptum® sampling system
 1 and 2 mm gauge sampling needles
• NovaSeptum® GO sampling system
 2 mm gauge sampling needle
• Gamma and beta-irradiated products
Challenge Organism
 Brevundimonas diminuta (on average 0.3 by 0.7 µm in size)
Assay
 Presence/absence of microbial growth (turbidity) in medium flow-through with bacterial identification
confirmation

29. Multiple actuation microbial ingress testing
29
Microbial ingress challenge and outcomes
Purpose
 Determine if multiple actuations of the trigger system can be performed while maintaining microbial
integrity.
Study Scope
• NovaSeptum® sampling system
 1 and 2 mm gauge sampling needles
• NovaSeptum® GO sampling system
 2 mm gauge sampling needle
• Gamma and beta-irradiated products
Challenge Organism
 Brevundimonas diminuta (on average 0.3 by 0.7 µm in size)
Assay
 Presence/absence of microbial growth (turbidity) in medium flow-through with bacterial identification
confirmation

30. 30
Sterile flow path medium: Trypticase soy broth
Challenge organism: Brevundimonas diminuta inoculation >108 colony forming units per test article
Operating pressure: 5 psi
Red arrow indicates medium flow direction
NovaSeptum® Sampling System
Medium Collection
Bottles
Valve Bank
Pressure Vessel

31. Method
Microbial ingress challenge and outcomes
31
Microbial Ingress TestsNegative Control
Collect medium flow-
through prior to microbial
challenge
Actuation Series
1. Steam-in-place
2. Challenge
3. Collect medium flow-
through after 1, 20,
40, and 50 actuations
Pre-actuated
1. Actuate test article 50
times
2. Steam-in-place
3. Challenge
4. Collect medium flow-
through after 5
additional actuations
Positive Control
1. Rupture the sheath at
the challenge site
(red rectangle)
2. Challenge
3. Collect medium flow-
through after 5
actuations
Challenge

32. Experimental plan and acceptance criteria
Microbial ingress challenge and outcomes
32
Sampling Needle Test Article Pretreatment Experiments
2 mm Gamma-irradiation Pre-actuated test article
Beta-irradiation Actuation series
1 mm Gamma-irradiation Actuation series
Beta-irradiation Pre-actuated test article
2 mm
NovaSeptum® GO
Beta-irradiation Actuation series
*Negative and positive control performed for each experiment
Acceptance Criteria:
 Negative Control: Absence of the challenge organism
 Positive Control: Presence of the challenge organism in media flow-through
 Microbial Ingress: Presence of the challenge organism in media flow-through

33. Actuation requirement (undiluted challenge)
Development of the positive control
33
Presence (+)/Absence (-) of Challenge Organism
Treatment—Number of Actuations
Test Article
Negative
Control
1 2 3 4 5 10
2 mm needle,
beta-irradiation
- - + + + + +
Negative Control: (-) indicates absence of the challenge organism
Actuated Test Article: (+) indicates microbial ingress after the
corresponding number of actuations
A positive control was used
to have the challenge
organism in direct contact
with the needle and
septum.
Two actuations were required to
force microbial ingress.

34. Limit of detection (five actuations)
Development of the positive control
34
Negative Control: (-) indicates absence of the challenge organism
Actuated Test Article: (+) indicates microbial ingress for the
corresponding challenge level
Presence (+)/Absence (-) of Challenge Organism
Treatment—Challenge Concentration (CFU)
Test Article
Negative
Control
62 620 6.2x104 6.2x106 6.2x108
2 mm
Needle,
Beta-
Irradiation
- - + + + +
Deliberately damaged
devices required five
actuations with >62
colony forming units
(CFUs) per test article
to demonstrate
microbial ingress.

35. 35
Actuation series for NovaSeptum® sampling systems
Microbial ingress challenge and outcomes
Negative Control: (-) indicates absence of the challenge organism
Positive Control: (+) indicates a deliberately damaged sheath allows ingress and the system has sufficient sensitivity to detect ingress
Actuated Test Article = (-) indicates no microbial ingress
Presence (+)/Absence (-) of Challenge Organism
Treatment—Number of Actuations
Test Article Sample
Negative
Control
1 20 40 50
Positive
Control
2 mm Needle,
Beta-irradiation
1 - - - - - +
2 - - - - - +
1 mm Needle,
Gamma-irradiation
1 - - - - - +
2 - - - - - +
NovaSeptum® sampling systems showed no microbial
ingress up to ≥50 actuations.

36. Pre-actuated NovaSeptum® sampling systems
Microbial ingress challenge and outcomes
36
Presence (+)/Absence (-) of Challenge Organism
Test Article Sample
Negative
Control
50 Actuations (pre-sterilization) +
Five (post-inoculation)
Positive
Control
2 mm Needle,
Gamma-irradiation
1 - -
+2 - -
3 - -
1 mm Needle,
Beta-irradiation
1 - -
+2 - -
3 - -
Two actuations were required to
force microbial ingress.
Pre-actuated NovaSeptum® sampling systems showed no
microbial ingress up to ≥55 actuations.
Negative Control: (-) indicates absence of the challenge organism
Positive Control: (+) indicates a deliberately damaged sheath allows ingress and the system has sufficient sensitivity to detect ingress
Actuated Test Article = (-) indicates no microbial ingress

37. NovaSeptum® GO sampling system microbial integrity
Microbial ingress challenge and outcomes
37
Presence (+)/Absence (-) of Challenge Organism
Treatment—Number of Actuations
Test Article Sample
Negative
Control
1 10 20 40 50
Positive
Control
2 mm,
Beta-irradiation
1 - - - - - - +
2 - - - - - - +
Negative Control: (-) indicates absence of the challenge organism
Positive Control: (+) indicates a deliberately damaged sheath allows ingress and the system has sufficient sensitivity to detect ingress
Actuated Test Article = (-) indicates no microbial ingress
NovaSeptum® GO sampling systems showed no microbial
ingress up to ≥50 actuations.

38. 38
Conclusion
 All NovaSeptum® and NovaSeptum® GO devices showed no
ingress of bacteria up to ≥50 actuations*
 All controls performed as expected, demonstrating the test
system has sufficient sensitivity to detect microbial ingress
when known defects are present.
*The decision to perform multiple actuations, and how many actuations, must be
assessed independently depending upon the nature of the fluid being sampled (or
is in contact with the face of the NovaSeptum® being actuated) and critical nature
of the application.

39. NovaSeptum® GO sterile sampling
Benefits
Easy
Implementation
Closed, Safe
System
Flexible
Efficient
3939
Closed, easy to use and
validate, the system
improves operational
efficiency
Pre-loaded or
configure onsite
accommodating a
wide range of holders
and sampling options
for adaptable and
flexible sampling
throughout a process
Sampling actuation
evidence and lock
control ensures
process integrity and
a representative
sample
Sterile sampling is
increasingly
recommended by
regulatory bodies

40. NovaSeptum® GO Sterile Sampling
A sterile, secure, and flexible
sampling solution for efficient
sampling across your entire
manufacturing process
1
Flexible:
• Broad range of standard and custom containers
and connectors to meet every sampling need
• Batch to batch ajustable in standalone mode
2
Closed, safe system:
• First actuation locking tag and safety ring to
strengthen risk mitigation plan
3
Easy Implementation:
• Inert material of construction for reliable sample
representativity
4
Efficiency:
• Preconfigured or configure on site
• Ready-to-use
40

41. Poll question

42. Sterile sampling value
assessment

43. Sterile sampling value assessment
What do we do?
43
Quantified
Value
Propositions
Support
Process
Efficiency
Initiatives
Evaluation of
New Products
&
Technologies
VALUE
MANAGEMENT
CENTER
(VMC) Facilitate
Sustainable
Collaboration
Support
Customers
Strategic
Objectives
Partner with
our Customers

44. Overview of the model of NovaSeptum® sampling
Sterile sampling value assessment
44
Discover the true
value of
NovaSeptum®
sterile sampling
system
Scope
Contact
 Get in touch with us directly
 Contact one of our single-use
specialists
 Ask your local account manager to
contact us
Scenario Manager tool compares
traditional sampling techniques
with NovaSeptum® sterile sampling
system.
Objective Guidance
We offer guidance throughout your
journey to quantify the value of
NovaSeptum® sterile sampling system
for your process to ensure a robust
analysis.
Supporting customers to evaluate the
economic, process efficiency and
risk mitigation benefits associated
with the adoption of NovaSeptum®
sterile sampling system.

45. Scenario modeling to quantify the benefits
Sterile sampling value assessment
45
1
2
3
Evaluate your full savings potential associated with
the adoption of NovaSeptum® sterile sampling
system
Model
Assumptions
Business case
Draft
Validation

46. Case study example
Sterile sampling value assessment
46
62% Reduced labor hours
35% Reduced product loss during
sampling
80% Reduced deviation costs
400 Autoclave cycles eliminated
10%
Overall cost reduction
Key facts
Background
Replace standard open sampling with
NovaSeptum® sterile sampling assembly
Outcome

47. Global Product Manager, Sterile
Sampling, Process Solutions
Marc-antoine.kaag@emdgroup.com
Senior Technical Consultant, Asia Pacific,
Process Solutions
Somasundaram.g@emdgroup.com
MarcAntoineKaagSomasundaramG.
The vibrant M, NovaSeptum are trademarks of Merck KGaA, Darmstadt, Germany or its affiliates. All
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