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T

he following article describes


the various steps regarding
the Installation Qualification
(IQ), Operational Qualification (OQ)
and Performance Qualification (PQ)
that were executed to complete the
validation of a pharmaceutical micro-
biology laboratory incubator. This
validation was designed and per-
formed from the viewpoint of a
microbiologist with assistance from
validation experts. The protocol used
for this validation was previously
published in the Journal of Valida-
tion Technology.
1
Descr iption of the I ncubator Unit
The incubator is a conventional unit where fresh
air enters though an air intake at the bottom. The air
is heated at a plenum below the chamber and flows
into the chamber in a uniform flow pattern. Exhaust
air is vented through a port at the top of the incubator.
The unit is a Precision Thelco Incubator catalog num-
ber 31487. This is a gravity convec-
tion unit model number 6M with an
operating temperature range of 25C
to 70C. The incubator has the fol-
lowing electrical characteristics:
Volts- 120
Hertz- 50/60
Watts- 300
The overall dimensions of the
unit are 42 X 21 X 39 with cham-
ber dimensions 36 X 18 X 27. T h e
incubator contains a dual hydraulic
thermostat, a mercury-in-glass ther-
m o m e t e r, two shelves, two heaters, double inner
doors, and double outer doors.
I nstallation Qualification (I Q)
During the IQ, personnel documented all items list-
ed in the Incubator IQ Form. Refer to Appendix A o f
validation protocol, previously published in the
Journal of Validation Te ch n o l og y.
1
A second person
reviewed and approved all verifications.
cT hi s val i dati on
was desi gned
and per for med
fr om the
vi ewpoi nt of a
mi cr obi ol ogi st
wi th assi stance
fr om val i dati on
exper ts. d
Val i dati on of a L abor ator y
I ncubator Usi ng Wi r el ess
and Cabl ed Datal ogger
Par t I I
By Laura Valdes-Mora
Elite MicroSource Corporation
and Pamela Hardt-English
PhF Specialists, Inc.
O
Laura Valdes-Mora & Pamela Hardt-English
Purpose
IQ was conducted to demonstrate that the piece
of equipment was inspected, and contains all the
required parts for its proper functioning. The pres-
ence/absence of external equipment aides (recorder
charts, etc.) was also documented.
Equipment and Materials
Incubator
Appendix A
I Q Observations
Incubator Description The unit is a Precision
Thelco Incubator, catalog number 31487. The
temperature range it is designed for is from
room temperature to 70C. The unit uses 120
volts and 5.2 amps. This incubator had not been
previously validated, and was 29 years and
seven months old at the time this validation was
conducted.
Incubator Model Number is 6M.
Incubator Serial Number 22-AG-8.
Incubator was purchased on July 14, 1971 from
Precision Scientific.
Vendor Manual is available.
Location of Vendor Manual The manual can be
found in the file cabinet of the microbiology lab-
o r a t o r y.
Drawings for the incubator are available and
located in the vendor manual.
Utilities required Electricity is required for this
unit. The power source was found to be present
and functional. A member of the maintenance
department measured the Volts and the amper-
age reading of the circuit of the electrical outlet
where the incubator is plugged in. The data were
recorded as found.
Utility Data
Circuit Rating:
130 Vo l t s 1 Phase 60 Hertz 20 A m p s
Equipment Rating:
125 Vo l t s 1 Phase 60 Hertz 5.2 A m p s
Field Measurement:
122.9 Vo l t s 1 Phase 60 Hertz 4.8 A m p s
Cord and Plug NEMA Designation: 5-15 P
Wall receptacle NEMA Designation: 5-15R
Notes: The ground wire is connected. Non-load
cycle Amps = 0.5
An Uninterrupted Power Supply (UPS) is not
used on this incubator.
This unit is not hooked to a back-up generator.
List of Standard Operating Procedures (SOPs)
associated with this incubator; the SOPs were list-
ed by number, title, and revision number. T h e y
are not included in the article as SOPs, and are
confidential documents of the company where we
conducted this validation. SOPs were reviewed
and were found to be appropriate.
The incubator does not have a temperature display.
There is an external temperature recorder in use
for this unit, which is a Dickson Temptrace Re-
corder Chart. The recorder chart is identified as
number one and contains a replaceable seven-day
chart (graph).
Acceptance criteria were met.
Operational Qualification: (OQ)
In this section the performance of the equipment
and its operational procedure was verified. The OQ
of an incubator consists of an empty chamber tem-
perature mapping.
Purpose
The OQ was performed to confirm that the empty
chamber works within the specified limits of tem-
perature (32 to 37C) throughout the chamber .
Equipment and Materials
Precision Thelco Incubator (32-37C)
Real Time Datalogger - Ellab Model TM9616
Real Time Datalogger
Serial Number 00270110141
Thermocouples Type T (Cu/Con)
Wireless Data Loggers
Ellab Model T S M 9 8 0 3
Serial Number 2056
Temperature Sensor Serial Number
LC 1 20022 6422
LC 2 20753 5757
LC 3 20770 5747
LC 4 20764 5770
LC 5 20804 5766
LC 6 20717 5309
Laura Valdes-Mora & Pamela Hardt-English
LC 7 20713 5219
LC 8 20703 4521
LC 9 11542 5679
LC 10 11537 4553
Windows-based PC or laptop computer to oper-
ate data loggers
External Temperature Device
Timer
Equipment Picture
See Figure 1.
Acceptance Criterion
All sites measured are within 32-37C.
OQ Observations
Calibration
The calibration of thermocouples was performed
using a Kaye LTR-140 serial number 5C339H, owned
by the pharmaceutical company where we conducted
this validation. A total of 12 Thermocouples (TCs)
were tested, and 10 of those were used for the tem-
perature profiles. Calibrations were satisfactorily con-
ducted according to the SOPs. The two extra calibrat-
ed thermocouples were kept in reserve in case of a
failure of one of the others.
Location of Thermocouples and Loggers- Dia-
grams: The purpose of the diagram is to show the
relative position of the thermocouples and wireless
sensors. The figure is not to scale.
In the diagrams seen for the top and lower shelf,
each number corresponds to the thermocouple and
the logger placed at the location. The two devices
were side-by-side for each one of the locations indi-
cated. The box in the lower shelf with an R in the
center indicates the location of the chart recorder, as
shown in Figure 2.
Tests for Temperature Setting
The incubator was set to 35-37C or 36 + 1 C .
While TCs were being placed, the incubator was
opened for 20 minutes. It took 30 minutes after clos-
ing the doors of the incubator for the unit to reach
35C. Note that the chart recorder was reading 32.4C.
An additional reading of the recorder indicated the
temperature to be 35.5C.
Test of temperature setting number one (lowest
temperature setting on this incubator). In order to
expedite the decrease in temperature, we opened the
door of the incubator for a few minutes. The temper-
ature decreased to a range between 25-29C. A f t e r
closing the door and allowing T C s to stabilize, the
lowest temperature recorded was 29C, as shown in
Fi g u re 3.
Test of Temperature Setting #9 (highest safe
temperature setting on this incubator). Safety light
came on at 66C. Chart recorder read 49.9C, while
the TC next to the recorder chart read 59.2C as
Figure 2
R elative Pos ition of the
Thermocouples and
Wireles s S ens ors
Top S helf
Lower S helf
Front
Front
1 2
3
4 5
6 7
8
9 10
R
Figure 1
E mpty Chamber S et-Up
E mpty
Laura Valdes-Mora & Pamela Hardt-English
shown in Fi g u re 4. It appears that the chart recorder
responds more slowly than the thermocouples to
temperature changes.
Due to the fact that the safety light came on during
this test, it was decided to record current tempera-
tures, and set the knob back to its regular setting of
number five. The incubator manual was reviewed,
and it was found that the safety light is related to a
safety thermostat, which operates alternatively to the
control thermostat.
After increasing the temperature as per step num-
ber three, it took the incubator 75 minutes to come
down to the set temperature range of 35-37C.
Once all thermocouples were within the tempera-
ture range established by the protocol (32-37C),
with the understanding that temperature readings
can range from 31.5 up to 37.4C (per rounding
rules), the 24-hour mapping study was started.
24-Hour Temperature Mapping
The empty chamber test was conducted for over
24 hours. Temperature readings were recorded every
two minutes. From the time the run was started
(02:10:40 PM) to its end, the time was determined to
be 26 hours, 44 minutes, and 39 seconds.
At 03:52:40 PM, all TCs were within temperature
range. There were still some hot pockets left from
the previous study. It took the chamber an addition-
al 42 minutes to stabilize all locations.
The data obtained show that all thermocouples
were within the specified temperature range for 24
hours as shown in Figure 5.
Wi reless Datalogge r, Tra ck s e n s e : The study start-
ed at 08:43:00. All wireless loggers (LCs) were at the
t a rget temperature range at 10:07:00 as shown in
Fi g u re 6. It took the sensors 1 hour and 24 minutes to
all be stabilized. This is almost double the amount of
time it took the T C s to get into temperature range.
Figure 3
R es ults of Low
Temperature S etting
4 0 . 0
3 5 . 0
3 0 . 0
2 5 . 0
2 0 . 0
00.32 00:37 00:42
S etting
TC 1 Top Shelf Back Left
TC 2 Top Shelf Back Right
TC 3 Top Shelf Center
TC 4 Top Shelf Front Left
TC 5 Top Shelf Front Right
TC 6 Bottom Shelf Back Left
TC 7 Bottom Shelf Back Right
TC 8 Bottom Shelf Center
TC 9 Bottom Shelf Left Fr o n t
TC 10 Bottom Shelf Front Right
TC 13 Description
TC 14 Description
TC 15 Description
TC 16 Description
Figure 4
R es ults of High
Temperature S etting
7 0 . 0 0
6 0 . 0 0
5 0 . 0 0
4 0 . 0 0
3 0 . 0 0
2 0 . 0 0
1 0 . 0 0
0 . 0 0
01:08 01:13 01:18 01:23
S etting
TC 1 Top Shelf Back Left
TC 2 Top Shelf Back Right
TC 3 Top Shelf Center
TC 4 Top Shelf Front Left
TC 5 Top Shelf Front Right
TC 6 Bottom Shelf Back Left
TC 7 Bottom Shelf Back Right
TC 8 Bottom Shelf Center
TC 9 Bottom Shelf Left Fr o n t
TC 10 Bottom Shelf Front Right
TC 13 Description
TC 14 Description
TC 15 Description
TC 16 Description
Laura Valdes-Mora & Pamela Hardt-English
In order for this to be a 24-hour run, the readings
needed to last until 10:07:00 on day number two. T h e
last reading on day number two was made at
08:41:00, which completes the 24-hour run, but does
not map the temperature within specified ranges for
24 hours. This is one hour and 20 minutes short of the
requirement. This is due to the fact of not being able
to see the data until the study is complete. Note that
the wireless system is being used to determine its
suitability for validating incubators by comparing it
to the real time system. This validation study is based
on the data obtained from traditional thermocouples.
Thermocouple Test: Started at 2:40:40 and ended
at 26:52:40.
All TCs reached the target temperature range at
2:40:40. During the first hour and eight minutes, there
were 10 temperature excursions: one on TC 5 of
0.1C and the other seven on TC 8. Of the excursions
on TC 8, six were of 0.1C, and one was of 0.2C.
This was not observed at any other point in time dur-
ing the 24-hour study. This indicates that there were
still a few hot spots in the chamber after the highest
temperature setting test was completed immediately
prior to the empty chamber mapping. Future mapping
studies should allow for more time between tests
either by adding two extra hours, or by running the
mapping study for 30 hours in order to compensate
for the adjustment period. There are companies that
change temperature ranges of their incubators on a
regular basis, due to the variety of tests conducted and
the lack of incubators. In this situation, the chamber
should be allowed several hours to equilibrate prior to
loading it. However, this is not the case of this partic-
ular piece of equipment. Therefore, this phenomenon
observed during the 24-hour empty chamber test
should not occur during day-to-day use.
Tracksense: All wireless sensors were within
specified temperature range at 9:43:00. During the
first 24 minutes, three sensors reached 37.5C. This
was not observed at any other point in time during
the 24-hour study.
During the review of TC temperatures, the excur-
sions stated above were not found. The highest tem-
perature found during this review using the comput-
er screen was 37.1C
Figure 5
R es ults of 24-Hour E mpty
Chamber Mapping
Thermocouples
4 0 . 0
3 0 . 0
2 0 . 0
1 0 . 0
0 . 0
02:40 07:25 12:10 16:55 21:40
S etting
TC 1 Top Shelf Back Left
TC 2 Top Shelf Back Right
TC 3 Top Shelf Center
TC 4 Top Shelf Front Left
TC 5 Top Shelf Front Right
TC 6 Bottom Shelf Back Left
TC 7 Bottom Shelf Back Right
TC 8 Bottom Shelf Center
TC 9 Bottom Shelf Left Fr o n t
TC 10 Bottom Shelf Front Right
TC 13 Description
TC 14 Description
TC 15 Description
TC 16 Description
Figure 6
R es ults of 24-Hour E mpty
Chamber Mapping
Wireles s Tracks ens e
4 0 . 0
3 0 . 0
2 0 . 0
1 0 . 0
0 . 0
08:43-1 13:28-1 18:13-1 22:58-1 03:43-2
S etting
LC 1 Top Shelf Back Left
LC 2 Top Shelf Back Right
LC 3 Top Shelf Center
LC 4 Top Shelf Front Left
LC 5 Top Shelf Front Right
LC 6 Bottom Shelf Back Left
LC 7 Bottom Shelf Back Right
LC 8 Bottom Shelf Center
LC 9 Bottom Shelf Left Fr o n t
LC 10 Bottom Shelf Front Right
Laura Valdes-Mora & Pamela Hardt-English
All temperature devices were checked, and all were
within the specified limit of + 2C from each other.
Note that the temperature loggers recorded diff e r e n t
actual clock times. The loggers were not synchronized
for this study, neither were they set to local time. Each
one was set for a different time zone. Areal time elapsed
adjustment was made to account for the diff e r e n c e .
Performance Qualification: (PQ)
In this section it was verified that the loaded incu-
bator chamber was capable of reaching and main-
taining the target temperature range.
Purpose
The PQ confirms that the loaded chamber works
within the specified limits of temperature through-
out the chamber. The effect of opening doors for two
different lengths of time, and the effect of a power
failure on the incubators ability to get back into the
temperature profile were also evaluated.
Equipment and Materials
Precision Thelco Incubator (32-37C)
Real Time Data Logger Ellab Model TM9616
Real Time Data Logger
Serial Number 00270110141
Thermocouples Type T (Cu/Con)
Wireless Data Loggers
Ellab Model TSM9803
Serial Number 2056
Temperature Sensor Serial Number
LC 1 20022 6422
LC 2 20753 5757
LC 3 20770 5747
LC 4 20764 5770
LC 5 20804 5766
LC 6 20717 5309
LC 7 20713 5219
LC 8 20703 4521
LC 9 11542 5679
LC 10 11537 4553
Windows-based PC or laptop computer to
operate data loggers.
External Temperature Device (chart recorder)
Timer
Agar plates and culture broths to create a maxi-
mum loaded chamber pattern.
Acceptance Criterion
All sites measured are within 32-37C
PQ Observations
A maximum load pattern was selected for the
PQ. In addition, the effect of opening doors for two
d i fferent lengths of time, and the effect of a power
failure on the temperature profile were evaluated in
this phase. Ty p i c a l l y, these evaluations are con-
ducted during the OQ, however, we chose the
loaded chamber for these tests as we consider this
scenario more realistic, therefore, more meaning-
f u l .
Loaded Chamber Description
Top Shelf:
4* 2000 mLscrew cap bottles of Lactose
B r o t h
4 1000 mL screw cap bottles contain-
ing 500 mL each of Lactose Broth
1 Rack of 40 screw cap test tubes with
10 mL each of Nutrient Broth
2 Baskets each containing 11 Whirlpak
bags each with 100 mL TSB**
Lower Shelf:
1 Chart recorder
1 Water bottle (used by laboratory for
temperature determinations)
15 Stacks of plates- total of 72 plates- 42
Blood Agar and 30 TSA***
12 Whirlpak bags with 250 mL Lactose
Broth each (in a basket)
4 API strips
* Two (2) 2000 mLbottles were removed from
the study. See observations for explanation.
** TSB= Tryptic Soy Broth
*** TSA= Tryptic Soy Agar
Loading Pattern Diagrams: The purpose of the
diagrams in Figure 7 is to show the relative position
of the various items in the load. The diagrams are
not to scale.
Numbers on each of the above rectangles denote
number of plates on each stack. If the number is fol-
lowed by mL, it denotes the volume of medium on
each container.
Laura Valdes-Mora & Pamela Hardt-English
Loaded Chamber Picture
An example of a loaded chamber can be seen in
Figure 8.
Maximum Load Pattern
After loading the chamber and waiting one hour
and 45 minutes, it was observed that only 2 TCs were
within the target temperature range. We analyzed the
content of the load, and laboratory personnel re-
viewed the loading pattern. It was concluded that the
incubator was loaded above the worst-case scenario.
Four 2-liter flasks are never placed into this incubator
by laboratory personnel. This was done only once
during the validation of a microbial limit test, but as it
is not a regular procedure, it was chosen to decrease
the loaded chamber by two 2-liter flasks, based on
previously performed work.
Following this action, it took the chamber two
hours and 45 minutes to reach the set temperature.
Five-Minute Opened Door Test
This incubator consists of two double doors.
During this test all doors were opened for five-min-
utes. The purpose of the test is to simulate a worse case
scenario when laboratory analysts are loading inocu-
lated samples and their controls into the incubator.
During this test time, the temperature of the incu-
bator decreased. Temperature ranged from 22.9 to
30.2C across the loaded chamber as shown in Fig-
ure 9. The chart recorder reading was 30.5C. It took
50.5 minutes for the loaded chamber to return to the
set temperature.
Three-Hour Power Outage Test
The purpose of the test was to simulate the eff e c t
of a power outage on the temperature profile of a
fully loaded incubator chamber. The TCs were
checked to ensure all sites were at the set temperature
and proceeded to disconnect the incubator. A f t e r
three hours, the temperature across the chamber was
reviewed and the incubator was reconnected to its
power supply.
There was a decrease in temperature that took the
loaded chamber outside the set temperature and out-
side the target temperature. Temperatures ranged
from 27.7 to 29.4C as shown in Fi g u re 10. The chart
recorder was reading 28.7C.
Figure 7
R elative Pos ition of the Various
Items in the Load
B ottom S helf
T1 T2
T3
T4 T5
T6 T7
T8
T9 T 1 0
2000
mL
Top S helf
2000
mL
Test Tube
Rack
10
A
10
B
3
A
4
B
4
A
5
A
4
B
4
B
4
B
6
B
8
A
R C
2 B
2 B 3 B
3 B
Basket
#1
1
0
0
0
mL
1
0
0
0
mL
1
0
0
0
mL
1
0
0
0
mL
Basket
#2
Basket
API
R= Chart recorder
A= TSA plates
C= Water Control
B= Blood Agar Plates
T= Thermocouple number (a
wireless sensor is next to
each thermocouple)
Figure 8
Picture of Fully Loaded Chamber
Full
Laura Valdes-Mora & Pamela Hardt-English
Following the power start-up, it took one hour
and 35.5 minutes for the temperature to reach the set
point of 35-37C. This unexpectedly long ramp time
signals that the load also dropped in temperature,
thus requiring a longer time to return to the set tem-
perature.
One-Minute Opened Door Test:
The protocol was originally written for a five-
minute and a 15-minute opened door test. After per-
forming the five-minute opened door test, it was
determined that most of the time the incubator doors
do not remain open longer than just a few minutes,
typically not even reaching the five-minute mark.
Based on this, it was decided to change the 15-
minute test to a one-minute test in order to represent
a true scenario. In addition, for this test, only one side
of the double doors was opened, not both, for the
same reason of mimicking actual use.
During the one-minute opened door test, the tem-
perature decreased and was outside the set tempera-
ture in some areas of the chamber. The temperatures
ranged from 29.3 to 35.2C as shown in Fi g u re 11.
The chart recorder was reading 33.6C. It took 15 min-
utes for the chamber to return to the set temperature
once the door was closed.
24-Hour Loaded Chamber Temperature Mapping
The loaded chamber test was conducted for 24
hours. Temperature readings were recorded every
two minutes. The study started at 08:32:00 and ended
at 32:16:00. This is 16 seconds short of precisely 24-
hours, however, due to rounding rules, this small
Figure 10
R es ults of Three-Hour
Power Outage Tes t
7 0 . 0 0
6 0 . 0 0
5 0 . 0 0
4 0 . 0 0
3 0 . 0 0
2 0 . 0 0
1 0 . 0 0
0 . 0 0
03:36 04:31 05:26 06:21 07:16
S etting
TC 1 Top Back Left
TC 2 Top Back Right
TC 3 Top Center
TC 4 Top Front Left
TC 5 Top Front Right
TC 6 Bottom Back Left
TC 7 Bottom Back Right
TC 8 Bottom Center
TC 9 Bottom Front Left
TC 10 Bottom Front Right
Figure 9
R es ults of Five-Minute
Opened Door Tes t
7 0 . 0 0
6 0 . 0 0
5 0 . 0 0
4 0 . 0 0
3 0 . 0 0
2 0 . 0 0
1 0 . 0 0
0 . 0 0
02:34 02:44 02:54 03:04 03:14 03:24 03:34
S etting
TA 1 Top Back Left
TA 2 Top Back Right
TA 3 Top Center
TA 4 Top Front Left
TA 5 Top Front Right
TA 6 Bottom Back Left
TA 7 Bottom Back Right
TA 8 Bottom Center
Laura Valdes-Mora & Pamela Hardt-English
shortage is considered acceptable as this is a 24-hour
run, not a 24.0-hour run.
All thermocouples were within temperature range
at the start of the mapping, as shown in Fi g u re 12.
No excursions were found at any point in time,
thus corroborating that the excursions seen during
the empty chamber test were due to hot pockets left
from the highest temperature setting test.
The loaded chamber maintained a tight temperature
range throughout the 24-hour mapping study on a per
TC basis. The lowest temperature recorded was
34.5C found on TC 8, while the highest was 36.9C
found on TC 9.
Tra ck s e n s e : All loggers were within temperature
range at the start of the mapping as shown in Fi g u re
1 3. The study started at 09:50:00, and ended at
33:52:00, for a total of 24 hours and two seconds.
There were no excursions found at any point in time.
The lowest temperature recorded was 34.3C found on
LC 8, while the highest was 37.0C found on LC 6.
All temperature devices were checked and all
were within the specified limit of + 2C from each
other.
Note that the temperature loggers recorded diff e r-
ent actual clock times. The loggers were not syn-
chronized for this study, neither were they set to local
time. Each one was set for a different time zone.
Compar isons Between Data Collected by
T her mocouples and by Wir eless L ogger s
The thermocouples used are type T, copper-con-
stantan thermocouples. The quality of wire and con-
struction can significantly affect the accuracy of the
thermocouples. The thermocouples used in these
tests are Ellabs sealed thermocouples with a stain-
less steel tip. The Ellab Tracksense logger sensor is
a thermister made of a Silicon chip. Each logger and
sensor has a separate ID number, and is factory cal-
ibrated at five points in the range of -40 to 140C.
The accuracy of the Ellab thermocouples and the
Ellab Tracksense sensors is +/-0.2C.
Figure 11
R es ults of One-Minute
Open Door Tes t
4 5 . 0
4 0 . 0
3 5 . 0
3 0 . 0
2 5 . 0
2 0 . 0
07:58 08:08 08:18 08:28
S etting
TC 1 Top Back Left
TC 2 Top Back Right
TC 3 Top Center
TC 4 Top Front Left
TC 5 Top Front Right
TC 6 Bottom Back Left
TC 7 Bottom Back Right
TC 8 Bottom Center
TC 9 Bottom Front Left
TC 10 Bottom Front Right
Figure 12
R es ults of 24-Hour Loaded
Chamber Temperature Mapping
Thermocouples
4 0 . 0
3 0 . 0
2 0 . 0
1 0 . 0
0 . 0
08:32 13:17 18:02 22:47 27:32
S etting
TC 1 Top Back Left
TC 2 Top Back Right
TC 3 Top Center
TC 4 Top Front Left
TC 5 Top Front Right
TC 6 Bottom Back Left
TC 7 Bottom Back Right
TC 8 Bottom Center
TC 9 Bottom Left Fr o n t
TC 10 Bottom Front Right
Laura Valdes-Mora & Pamela Hardt-English
Can these data collection systems be used inter-
changeably? Fi g u re 14 shows a summary of 10 hours of
data from both the empty chamber 24-hour test and the
fully loaded incubator test for both the wireless and
thermocouple studies. The thermocouple data were col-
lected on a real time system, EVAL, and the wireless
data were collected using Ellabs Tracksense system.
This summary demonstrates that both systems
generated comparable results. The differences in val-
ues are well within the accuracy of both the thermo-
couples and the thermisters, which is +/-0.2C.
When complementing thermocouples with wire-
less loggers or using wireless loggers in place of ther-
mocouples, the issue of real time versus historical
data collection must be addressed. Thermocouples are
connected to real time data loggers, i.e., the data are
displayed as collected. The wireless system is histor-
ical, i.e., the data are downloaded and displayed at the
conclusion of the test. Since the data cannot be seen,
preliminary tests might need to be conducted to deter-
mine the start time on a test, such as the 24-hour fully
loaded incubator, because the temperature of the incu-
bator might drop during loading.
With a wireless system, there are no wires to get
tangled and assembly can proceed quickly. The Ellab
wireless data loggers are more stable and sealed, and
need less frequent calibration than thermocouples.
Thermocouples are frequently not sealed, and there-
fore subject to corrosion and degradation that will
a ffect the calibration. It must be also kept in mind
that not all thermocouples and wireless data loggers
are as accurate as the sensors used in this study.
Summary of Deviations:
The following are the deviations that occurred dur-
ing the execution of this validation. All deviations from
Figure 13
R es ults of 24-Hour Loaded
Chamber Temperature Mapping
Wireles s Tracks ens e
4 0 . 0
3 0 . 0
2 0 . 0
1 0 . 0
0 . 0
09:50 14:35 19:20 24:05 28:50
S etting
LC 1 L # 20022 S # 6422
LC 2 L # 20753 S # 5757
LC 3 L # 20770 S # 5747
LC 4 L # 20764 S # 5770
LC 5 L # 20804 S # 5766
LC 6 L # 20717 S # 5309
LC 7 L # 20713 S # 5219
LC 8 L # 20703 S # 4521
LC 9 L # 11542 S # 5679
LC 10 L # 11537 S # 4553
Figure 14
Data Collection Comparis on Between Thermocouples and Wireles s Dataloge rs
All Values are in Degrees Centigrade
Type of Data File T i m e M i n i mu m M a x i mu m Av e r age Av e r age Delta T Av e r age
Te s t N a m e / S y s t e m I n t e rva l for all for all of Minimu m of Maximum ( R a n ge of all
(10 M i n i mu m M a x i mu m Va l u e s Va l u e s f ro m Va l u e s
H o u rs ) Va l u e s Va l u e s Av e r age )
Empty Eval1.dt 6:00:00- 36.3 37.4 36.5 37.1 0.3 36.8
Chamber 4/EVAL 16:00:00
Track1.dt 6:00:00- 36.3 37.4 36.7 37.1 0.2 36.8
4/Tracksense 16:00:00
Loaded Eval 22.dt 12:00:00- 35.5 36.9 35.9 36.3 0.3 36.1
Chamber 4 22:00:00
Track2.dt 12:00:00- 35.4 36.8 35.9 36.3 0.5 36.2
4/Tracksense
Laura Valdes-Mora & Pamela Hardt-English
the original protocol were justified and approved.
There was no reference available for the Kaye
dry bath. Equipment was sent out to the manufactur-
er for routine calibration. The internal bath reference
was used for all calibrations of the dry bath per-
formed for this validation.
The protocol indicated that the chamber should be
allowed to be at temperature range for at least 30
minutes prior to conducting the five-minute opened
door test. Based on OQ data, it was shown that the
incubator is capable of maintaining the temperature
within the range for extended periods; therefore wait-
ing 30 minutes would not have made a diff e r e n c e .
A 15-minute Opened Door Test was described on
step 15 of the PQ protocol. This was changed to a one-
minute Opened Door Test, as typically incubator doors
are opened for not more than a couple of minutes. T h e
new test length was considered to be reflective of a
real situation.
The final post calibration of the thermocouples had
to be repeated because there was not enough data col-
lected to confirm the high temperature calibration at
40C. The EVA L Datalogger collects and displays
data every two seconds, but the program was set to
save data every two minutes, and the system was shut
down before the final data were stored. Therefore, the
calibration was repeated at Star Services, Hayward,
California, using a Kaye Liquid bath at two tempera-
tures, 30 and 40C to confirm that the thermocouples
were within specifications.
Conclusions
I Q
It is concluded that the unit was installed proper-
ly. All necessary components were found to be pre-
sent, and all the required SOPs were found to be in
place. The utilities (power) supplied to the unit, as
per testing, were found to meet the requirements.
OQ
All test results met the protocol acceptance crite-
ria. The unit is capable of maintaining the target
temperature.
PQ
All test results met the protocol acceptance crite-
ria. Based on data obtained from the opened door
tests and power failure, it can be concluded that nor-
mal laboratory practices are acceptable, and that
extending the incubation time for the same length of
time of the power failure is an acceptable solution.
Note that time should be calculated from the time
the incubator reaches the temperature range (see rec-
ommendations).
Conclusion
The incubator was tested and verified to operate
properly according to the manufacturer and protocol
requirements. This unit is therefore considered vali-
dated.
Recommendations
In order to have enough data collected, one should
consider running the temperature mappings with a
safety factor of six hours, that is, for a 24-hour run,
conduct a 30-hour mapping study, especially if wire-
less sensors will be used.
The validation presented here was completed in
three days of continuous work. Scheduling and exe-
cuting the work in this manner is taxing and compli-
cated. At times, one will be working very early or
very late hours. Careful pre-scheduling is highly rec-
ommended. Consider adding extra time to the sched-
ule for unforeseen situations (example: chamber may
not get to temperature in the time you have predict-
e d ) .
Older incubators that do not have a digital temper-
ature display can benefit from having one installed.
Digital temperature displays are available; they have
a Real Time Datalogger (RTD) which is placed in the
incubator either through a port (if available) or
through the top vent.
Microbiologists should consider that after open-
ing the incubator doors, the equipment loses heat,
causing a decrease in temperature, which typically
falls outside the teststemperature range. Based on
this, incubation times should be recorded from the
time the incubator is at temperature range, not from
the time samples are placed in the incubator.
Caution must be exercised when reading temper-
ature from an external chart recorder. Data from this
validation clearly showed that chart recorders are
slow in capturing accurate temperature. J
Laura Valdes-Mora & Pamela Hardt-English
About the Author s
L a u ra Va l d e s - M o ra serves as Technical S ervices Dir-
ector, acting as Principal Consultant and National
S p e a ker on US P Microbiology Updates and related
topics at E lite MicroS ource Corporation, a R esource
Center for Pharmaceutical and Biotechnology M i c r o-
b i o l o g y. S he received a B. S. d e gree in Biology from the
U n i versity of Puerto R ico in 1982, and an M.S. d e gr e e
in Microbiology from North Carolina State University in
1 9 8 6 . M s. Va l d e s - M o ra was a Director at AAI P h a rm a ,
was fo rm e rly in charge of production of bacteri a l
endospores at AMS CO, and has been the President
of the Pharmaceutical Microbiology Forum (PMF)
since 1994. Ms. Valdes-Mora can be reached by
phone at 850-763-5453, by fax at 850-913-9735, or by
e-mail at E MS ource@ aol.com.
Pamela Hardt-E nglish, is President of PhF S pecial-
i s t s. S he has M.S. d e grees in Food Science and
A gri c u l t u ral E ngineering from the University of Califo r-
nia (Davis) and a B. A . in Computer S cience from the
University of California (Berkeley). For the last 23
years at PhF Specialists and FMC Corp o ration, Ms.
Hardt-E nglish has served as a recognized FDA and
U S DA process authori t y, instituted HACCP progra m s,
conducted thermal processing studies, and valida-
tions of autoclave s, incubators, and other chambers.
S he has designed and taught numerous food pro-
cessing courses. M s. Hardt-E nglish can be reached
by phone at 408-275-0161, by fax at 408-280-0979,
or by email at phfspec@ pacbell.net.
Suggested R eading
The following publications were consulted during the preparation of the
protocol, execution of the work, and preparation of the final report.
Cloud, P.A., Validating a Laboratory Incubator, B i o p h a r m30,
pp. 39-42. November (1997).
Phil Cloud, Pharmaceutical Equipment Validation: T h e
Ultimate Qualification Guidebook (Interpharm Press, Buffalo
Grove, IL1998).
Cloud, P., How to Develop and Manage Qualification Proto-
cols for FDACompliance. (Interpharm Press, Buffalo Grove,
IL 1999).
R efer ences
1. Institute of Validation Te c h n o l o g y. Journal of Va l i d a t i o n
Technology. Vol. 8 No. 1, November 2001. pp. 63-72.
IQ: Installation Qualification
LC: Wireless Loggers
OQ: Operational Qualification
PQ: Performance Qualification
RT D : Real Time Datalogger
S O P : Standard Operating Procedure
T C : T h e r m o c o u p l e
UPS: Uninterrupted Power Supply
TSA: Tryptic Soy Agar
TSB: Tryptic Soy Broth
Ar ticle Acr onym L isting

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