Participate in the interactive webinar now: http://bit.ly/ClosedSamplingWebinar Sampling should never put your process or sample at risk of contamination resulting in loss of time, money or regulatory scrutiny. This webinar discusses simple and effective sampling options to implement a secure sampling operation across your entire process driving towards closed manufacturing. Explore our webinar library: www.emdmillipore.com/webinars

1. The life science business of Merck KGaA,
Darmstadt, Germany operates as
MilliporeSigma in the U.S. and Canada.
Closed sampling, a critical
component for every risk
mitigation strategy:
A case study
Janmeet Anant, Alex Chalmers
20th February 2020

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

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

4. Contamination
control strategy &
Closed processing

5. Aseptic processing – A path full of pitfalls…
Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.20205
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:

6. Objectives of a contamination control strategy
Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.20206
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 Products”

7. Contamination control strategy: A holistic process
Contamination control strategy & closed processing
Assess
Control and
Monitor
Identify
Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.20207
Microbiological
Chemical
Particulate
Cross-product
contamination
1
3
2

8. Key considerations for implementing an effective strategy
Contamination control strategy & closed processing
Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.20208
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

9. Microbial contamination, a major concern for Biopharma…
 In addition to the potential impact to patients, there can be an economical
consequence associated with a contaminated batch that can include but is not
limited to:
− QA investigation time & resources
− Batch scrap
− Sanitization
− Commercial impact
− Facility temporary shutdown!…
 Batch scrap due to contamination can reach USD 1 million of loss for the
company*
 And up to several hundred millions USD as missing revenue!*
Mishap can occur due to inappropriate sampling installation
Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.20209
Cost of a contamination event
Contamination control strategy & closed processing
* Westman, David. (2017, May 3). Cost And Impact Of A Bioburden Incident. Retrieved from Bioprocess
online website: https://www.bioprocessonline.com/doc/cost-and-impact-of-a-bioburden-incident-0001

10. Poll question
Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.202010

11. Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.202011
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

12. Microbial ingress
challenge and
outcomes

13. 13 Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.2020
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)

14. 14 Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.2020
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)

15. 15 Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.2020
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)

16. 16 Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.2020
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)

17. Multiple actuation microbial ingress testing
Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.202017
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

18. Multiple actuation microbial ingress testing
Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.202018
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

19. Multiple actuation microbial ingress testing
Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.202019
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

20. 20 Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.2020
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

21. Method
Microbial ingress challenge and outcomes
Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.202021
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

22. Experimental plan and acceptance criteria
Microbial ingress challenge and outcomes
Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.202022
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

23. Actuation requirement (undiluted challenge)
Development of the positive control
Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.202023
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.

24. Limit of detection (five actuations)
Development of the positive control
Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.202024
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.

25. Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.202025
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.

26. Pre-actuated NovaSeptum® sampling systems
Microbial ingress challenge and outcomes
Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.202026
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

27. NovaSeptum® GO sampling system microbial integrity
Microbial ingress challenge and outcomes
Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.202027
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.

28. 28 Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.2020
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.

29. Poll question

30. NovaSeptum® GO sterile sampling
Benefits
Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.2020
Easy
Implementation
Closed, Safe
System
Flexible
Efficient
3030
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

31. 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
Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.202031

32. Sterile sampling
value assessment

33. Sterile sampling value assessment
What do we do?
Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.202033
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

34. Overview of NovaSeptum® sampling model
Sterile sampling value assessment
Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.202034
Discover the
true
NovaSeptum®
sterile sampling
value
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
NovaSeptum® value 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.

35. Poll question

36. Scenario modeling to quantify the benefits
Sterile sampling value assessment
Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.202036
1
2
3
Evaluate your full savings potential associated with
the adoption of NovaSeptum® sterile sampling
system
Model
Assumptions
Business case
Draft
Validation

37. Case study example
Sterile sampling value assessment
37
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
Closed sampling, a critical component for every risk mitigation strategy: A case study | 20.02.2020

38. Poll question

39. © 2020 Merck KGaA, Darmstadt, Germany and/or its affiliates. All Rights Reserved. The vibrant M, Millipore, and NovaSeptum are trademarks of Merck
KGaA, Darmstadt, Germany or its affiliates. All other trademarks are the property of their respective owners. Detailed information on trademarks is available
via publicly accessible resources.