Amy McDaniel, Director of Technical Operations at Pfizer, discusses moving environmental monitoring testing closer to the manufacturing floor.

1. Rapid/Automated Environmental
Monitoring on the Manufacturing Floor
Amy McDaniel, Ph.D.
Pfizer Biotech, Sanford, NC, USA

2. Outline
• Introduction to Pfizer Biotech, Sanford
• Introduction to Rapid Microbial Methods
(RMM) for Environmental Monitoring (EM)
• Implementation of the Growth Direct II in a
manufacturing facility
–Evaluation of Consumables
–Beta Instrument Evaluation
–Commercial instrument installation
• Conclusions

3. Introduction to Pfizer Biotech,
Sanford, NC
Sanford’s Role:
• Commercial manufacturing for Prev(e)nar and Prev(e)nar13 conjugate
• Commercial manufacturing for CRM197 carrier protein
• Supply clinical trial materials for microbial / conjugate products
• Commercial launch and production site for microbial / conjugate products within the PGS
network
• Contract manufacturing for microbial vaccines
Disclaimer:
This presentation is a case study of a system evaluation and implementation, it is not intended
to advertise or endorse any vendor’s technology.

4. Why RMM for EM?
4
Get materials
Sample in
Manufacturing
Wait for
settle plates
Wait for
incubation
Return to lab
Read
Results
Identify samples
Get materials
Prepare on
mfg floor Collect samples
according to role
PresentFuture?
Prepare in lab
Clean carts
Gown
Travel to manufacturing
Incubate plates
Clean up
Data entry
De Gown
QC Identifies samples
BioProcess Technicians
QC reviews results
Growth Direct communicates
with LIMS to download results,
sends alerts electronically as
programmed
Growth Direct
segregates
positive plates for
identification (in
QC lab), discards
negative plates
QC micro analysts
Reduced Time
to Results
Load samples into automated
technology

5. Why RMM for EM?
Get materials
Sample in
Manufacturing
According to
worklist
Wait for settle plates
(may exit the suite
and then return)
Wait for
incubation/transfer
plates to second inubator
Return to lab
Read
Results
Identify samples
Prepare
materials
Collect samples
according to role,
with reduced risk
of missed samples
due to automated
feedback loop
PresentFuture
Prepare in lab
Clean carts
Gown
Incubate plates
Clean up
Data entry
Into LIMS
De Gown
QC Identifies samples
QC reviews results
Growth Direct communicates
with LIMS to download results,
sends alerts electronically as
programmed
Growth Direct
segregates
positive plates for
identification (in
QC lab), discards
negative plates
QC micro analysts
Potential impact to controlled environment =
Potential for human error due to manual steps=
Potential for
Reduced Time
to Results
Load samples into automated
technology
Travel to manufacturing

6. 6
Shift huddles utilize a short, routine meeting & white board to
monitor/manage Micro lab operations
How are current EM sampling issues identified?
Relies on analyst knowledge and
memory to communicate any
missed samples.
The ideal technology should be programmed to
automatically read bar-coded labels, “know” what samples
to expect, and send email alerts if a sample is missing.
Error proof and time saving!

7. RMM for EM: Standard Work Confirms
Advantage
Break
Equipment / Process Controlled Analyst Controlled
Collecting Supplies
Travel to Area
Gowning
Cart Cleaning
Prep Equip and Materials in Manuf.
European Grade (4 carts) (20 Air sites)
Prepping Cart/Clean Up/Data Entry
Degowning/Travel to Lab
Data Download/Incubation
Collecting Supplies
Travel to Area
Gowning
Cart Cleaning
Prep Equip and Materials in Manuf.
US Classification (2 carts) (8 Air sites)
European Grade (2 carts) (9 Air sites)
Prepping Cart/Clean Up/Data Entry
Degowning/Travel to Lab
Data Download/Incubation
Collecting Supplies
Break
Travel to Area
Gowning
Cart Cleaning
Prep Equip and Materials in Manuf.
Hood Monitoring
Settle Plates (Room)
B Room
C Rooms (27 VA)
Clean Up/Data Entry/Data Download
Degown/Travel to Lab/Clean Up/Incubate
Lunch
EM
0# # # # # #12 13 14 15 16 17
Hours Evening Night
Weekly Requ
Samples /Tests
Test
Task
Type
Task
Day 1
8 9 10 11
PotentialEMRole(3)Hoods

8. Why GDII? What technologies are
available for Rapid/Automated EM?
Air Sampling
• Active Air Sampling:
– IMD (Instant Microbial
Detection, Biovigilant)
– BioLaz (Particle Measuring
Systems)
– BioTrak (TSI)
– Growth Direct II (Rapid Micro
Biosystems)
• Passive Air:
– Growth Direct II
Surface Sampling
• Swabbing & preparation
with:
– ScanRDI (BioMerieux)
– ATP systems (e.g., Celsis,
PallCheck, MilliFlex Rapid)
– Growth Direct II
• RODACS:
– Growth Direct II

9. Growth Direct II: What is it?
9
User Interface
Touch Screen
Interface to
Network or LIMS
Incubators
1 or 2 temperatures
300+
capacity/incubator
Interior
View
Input Queue
Cassette
Elevator
Slide graphics courtesy of Rapid Micro Biosystems

10. GDII: How does it work?
Patented technology uses a
blue light causing the micro-
colonies to autofluoresce:
this is captured on a CCD
chip
Growth Direct™ Imaging Visual Plate Counting
Day 5Day 4Day 3Day 2Day 1
The Growth Direct™
counts the same colonies
in half the time of the
traditional method.
0.5µ
Powerful software starts to detect colonies within hours, enabling real-time enumeration of organisms
12 hrs 16 hrs 20 hrs 24 hrs 28 hrs 32 hrs
An A. brasiliensis Microcolony in CHO cells

11. Technology overview: Growth Direct II
Traditional EM Growth Direct II
Media: TSA w/PS80 & lecithin (surface)
TSA (air)
TSA w/PS80 & lecithin with membrane overlay (based
on technology requirements)
Manual sampling by qualified personnel Manual sampling by qualified personnel
Automatic confirmation of required samples
Load plates into incubator at 1st temperature
Transfer plates to 2nd temperature
Automatic incubation at 1st temperature
Automatic transfer to second temperature at specified
time
Manual reading of plates Automatic reading of plates based on natural
fluorescence
Manual entry of data into LIMS Automatic download of results to LIMS
Discard plates or save for microbial ID Automatic handling rules to allow retrieval of plates for
ID based on program, or automatic disposal followed
by manual waste removal

12. Evaluating the Growth Direct II (GDII)
• Phase I: Evaluation of Consumables
• Phase II: Evaluation of Beta Unit

13. GDII: Evaluation of Consumables
• Why?
– The intention was to train manufacturing
technicians to take EM samples
– Designed the study to compare an
experienced analyst (QC Micro) with a
new technician
– Assess handling of plates, switching
lids, general equivalence of results with
offline incubation

14. GDII: Evaluation of Consumables
Active air sampling
• Same sampling
apparatus able to be
used for GDII and
traditional testing
• Media accommodated
within the sampler with
an adaptor (made by
manufacturer of the
sampling device)

15. Passive Air sampling:
• No difference in testing set
up for GDII and traditional
• Traditional plates are
slightly larger diameter than
GDII
GDII: Evaluation of Consumables

16. Surface Sampling with RODACs:
• Currently the only technology capable of automating this
element of EM
• Same convex surface of the plate with black membrane
overlayed on agar
• Diameter of the test surface is the same, GDII plates
have a wider base
GDII: Evaluation of Consumables

17. 17
Grade Sample Type
Results (Avg CFU/plate)
Growth Direct Traditional
A Active Air 0 0
Settle Plates 0 0
Surface #1 0 0
Surface #2 0 0
B Active Air 3 3
Settle Plates 1 1
Surface #1 0 0
Surface #2 1 1
C Active Air Site #1 9 11
Active Air Site #2 7 8
Settle Plates 3 1
Surface #1 0 0
Surface #2 5 1
Surface #3 0 0
Surface #4 2 1
Surface #5 0 0
D Active Air Site #1 23 24
Active Air Site #2 13 14
Settle Plates 1 2
Surface #1 5 5
Surface #2 0 0
Surface #3 1 1
Surface #4 0 0
Surface #5 0 0
Grade A: 1 active air site, 1 settling plate, 2 analysts, two reps each
2 surface sites, 2 analysts, 2 reps each
Grade B: 2 active air locations, 1 settling plate, 2 analysts, 2 reps each
3 surface sites, 3 analysts, 2 reps each
Grade C/D 2 active air sites, 1 settling plate, 2 analysts, 2 reps each
GDII: Evaluation of Consumables: Data

18. • Preparation for onsite evaluation in a GMP manufacturing
facility:
– Project plan
– Change control
– Impact assessment
– Evaluation Protocol
• Install the unit
• Execute evaluation protocol
GDII: Beta Evaluation

19. GDII: Beta Evaluation - Planning
• Proposal was to locate it within the classified area in manufacturing
– To gain the full benefit of efficiency, plates not moving out of the area, carts
not moving in
• Equipment pass through (Grade D)
– Evaluated options with project manager
– System requires water (to humidify incubators), air (to power the robotics),
230V power, gives off 4200BTU/hr of heat (all assessed for impact and
accommodated)
• Evaluated Risk of microbial growth in controlled area
– Closed incubators
– Plates are individually closed (lids lock to the base)
– Disposal chamber is contained within the instrument
– Increased monitoring around the instrument performed during evaluation
19

20. GDII: Beta Evaluation – Site Installation
Equipment pass through to
manufacturing. Uncontrolled
corridor looking into the pass
through.
Modular installation of unit
was preceded by vendor
assessment of area.

21. GDII: Beta Evaluation – Site Installation
Stacked incubators showing location of plates (the
parking deck), touch screen and keyboard interface.

22. GDII: Beta Evaluation - Project Plan
• Month 1:
– System install
– Train technicians on EM
– Program the test methods (two tiered incubation, load sampling sites,
load trend rules, load response to growth – output queue)
– Evaluate LIMS interface capability, network interface capability
• Success criteria
– Successful install (operational without rework) – June 26-28 (sample
ran successfully)
– Successful test method input (methods are accepted without rework
or computer issues, or with successful & timely resolution)
– Integration with LIMS is assessed as feasible
– Integration with network is assessed as feasible (“Pfizerized” PC on
7/2, connected to network)
22
√
√
√
√

23. • Months 2/3:
– 156 samples completed
– Inconsistencies evaluated (no discrepancies in action limits reached
by one system and not the other)
– Results indicated acceptability of the technology
GDII: Beta Evaluation - Project Plan
1133 C Air Site 2 3 6
1133 C Tank 120185 Top 0 0
1133 C Tank 120180 Top 2 0
1134 C Air Site 1 2 0
1134 C Air Site 2 2 2
1134 C Air Site 3 7 2
1134 C Tank 120110 Bottom 0 0
1134 C Wall 0 0
1134 C Counter 1 0
1134 C UP-1134 0 0
1135 C Air Site 1 4 1
1135 C Air Site 2 2 3
1135 C Air Site 3 8 1
1135 C Door to Rm 1131 0 0
1135 C Door to Rm 1145 11 0
1135 C Door to Rm 1153 0 0
1135 C Wall 0 0
1137 C Air Site 1 0 0
1137 C Air Site 2 0 1
Room Grade Sample Location
Growth
Dire/methodct
Result #CFU
Traditional EM
Result #CFU

24. GDII Beta Evaluation:
Potential Standard Work Efficiencies
Break
Equipment / Process Controlled Analyst Controlled
Collecting Supplies
Travel to Area
Gowning
Cart Cleaning
Prep Equip and Materials in Manuf.
European Grade (4 carts) (20 Air sites)
Prepping Cart/Clean Up/Data Entry
Degowning/Travel to Lab
Data Download/Incubation
Collecting Supplies
Travel to Area
Gowning
Cart Cleaning
Prep Equip and Materials in Manuf.
US Classification (2 carts) (8 Air sites)
European Grade (2 carts) (9 Air sites)
Prepping Cart/Clean Up/Data Entry
Degowning/Travel to Lab
Data Download/Incubation
Collecting Supplies
Break
Travel to Area
Gowning
Cart Cleaning
Prep Equip and Materials in Manuf.
Hood Monitoring
Settle Plates (Room)
B Room
C Rooms (27 VA)
Clean Up/Data Entry/Data Download
Degown/Travel to Lab/Clean Up/Incubate
Lunch
EM
0# # # # # #12 13 14 15 16 17
Hours Evening Night
Weekly Requ
Samples /Tests
Test
Task
Type
Task
Day 1
8 9 10 11
PotentialEMRole(3)Hoods
X
X
X
X
X
X
X
X
X
X
*
*
*
*
*
*
* Timing/collection supplies (carts)
could be reduced
X Step could be eliminated

25. GDII: Commercial Unit installation
Successful Beta Evaluation led to purchase of a commercial unit – Installed April 2014
Validation work slowed through the summer months due to resources

26. Growth Direct II:
Overall Validation Strategy
Instrument Qualification
• Per USP <1058>
• IQ by vendor/Pfizer
– Software/hardware components
– Utilities verified
– System set up/documentation
• OQ by vendor/verified by Pfizer
– Sequence of operations
• PQ by Pfizer
– E. coli, S. aureus, A. brasiliensis spike
study verifying method successfully stored
– Handling rules verified
– Response to growth verified (alert/action
notifications sent)
Method Validation
• Post-exposure recovery
• Accuracy / precision
• Concurrent testing (traditional/GDII)
• Strategy for settling plates
• Incubation parameters
– 2-tiered temp, 3 day/2 day
(current traditional method)
– 2-tiered 2 day/1 day*
– Single temp (32°C)**
– Single temp (22.5°C)**
*Potential to reduce incubation time
and receive more real-time results
**Potential to increase incubator
capacity with a single temp.
incubation (both incubators at one
temp.)

27. Why Transfer Sampling to Manufacturing?
• Aligns with current sampling responsibilities
– Manufacturing technicians currently take in process & release
samples from the process
• Minimizes flow of materials and personnel
– Reduces risk of contamination in controlled areas
• Increased awareness & ownership of EM
– Visual status of results and automated tracking of samples provides
real time feedback
• Better root cause analysis for events due to depth of
knowledge of manufacturing processes ongoing during
sampling

28. Category Action
Training QC oversight & periodic audits
LIMS Limited access role for technicians
Procedures
• Specify process flow & roles/responsibilities
• Impact evaluation for any program changes
• Material handling
• Continuity plan
Processes
Standard work
Evaluation of EM data trends
QC Oversight
 QC release of EM media
 QC authorization of data
 QC review and reporting of trend analysis
Periodic Review Periodic review of program & controls
Implementation Controls for Transfer of
Sampling Responsibilities
Documented in an approved risk assessment

29. Summary
Changes & Benefits: No Changes:
• Automation/incubation
•Reduce human error
•Earlier detection & response
• Sampling by manufacturing
•Minimize flow of personnel & materials
•Increased awareness of EM
• Consumables
•Membrane overlay – supports technology
•Base easier to manipulate
•Fundamental methodology is same
•Media type
•All positive plates evaluated by QC
•Release of consumables by QC
•Program oversight by QC
•Environment
•Air classification
•Clean utilities
•Personnel/material flow
•Acceptance criteria for action levels

30. Conclusions
• Rapid/Automated methods for EM can add value:
– Standard work efficiencies
– Environmental control (with faster results and notifications of trends
or events)
– Right First time (by automating receipt of expected samples and
counting errors)
• Moving EM technology and sampling to the
manufacturing area:
– Increases efficiencies mentioned above
– Decreases movement of materials in and out of controlled area
– Improves awareness of technicians to EM
– Ownership of the EM program resides with QC, Manufacturing is a
sampling resource
• The Growth Direct II is a significant benefit for an
environmental monitoring program