Ethylene Oxide Sterilization Validation Protocol

TITLE:- VALIDATION PROTOCOL FOR ETHYLENE OXIDE STERILISATION PROCESS
VALIDATION PROTOCOL NO: EFFECTIVE DATE:-
VALIDATION PROTOCOL OF ETHYLENE OXIDE STERILIZATION PROCESS
Company Name
Eto Sterilizer manufacturer name:
Date of Purchasing:
Date of Installation:
Model number:
Capacity:
Operator Name:
Department (Place)
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Table of Contents
1. PROTOCOL APPROVAL SHEET.........................................................................................4

2. REVISION HISTORY............................................................................................................ 4
3. OBJECTIVE:-....................................................................................................................... 5
4. SCOPE:-............................................................................................................................... 5
5. RESPONSIBILITY:-..............................................................................................................5
6. STANDARDS FOR REGULATORY REQUIREMENT:-.........................................................5
7. LIST OF DOCUMENTS:-......................................................................................................6
8. PRE-REQUISITE:-............................................................................................................... 6
Equipment qualification............................................................................................................6
Calibration:........................................................................................................................... 6
Product qualification.................................................................................................................6
Product and packaging material evaluation..........................................................................6
Product grouping.................................................................................................................. 6
Selection of family representative.........................................................................................7
9. Process and Validation Approach:-.......................................................................................8
Process Validation Procedure:-................................................................................................9
Bioburden assessment............................................................................................................. 9
Selection of Process Challenge Device (PCD..........................................................................9
BI or PCD placement in the product load: -..............................................................................9
Temperature and Humidity sensor in product load:..................................................................9
Load Temperature.................................................................................................................... 9
Identification of the worst case location or cold spot...............................................................10
Process Qualification Runs....................................................................................................11
a) Half Cycle Method.......................................................................................................11
b) Bioburden /BI method..................................................................................................13
Maintainace of Validation:...............................................................................................14
Requalification.................................................................................................................... 14
10. Sampling method and test method:-...............................................................................14
LIST OF TEST TO BE EXECUTED........................................................................................14
12.Process parameter check steps............................................................................................16
Environmental preconditioning: -............................................................................................17
Initial evacuation: -................................................................................................................. 17
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Humidification: -..................................................................................................................... 18
Gas injection and gas dwell: -................................................................................................18
Post exposure gas purge and air in bleed: -...........................................................................19
Heated aeration: -.................................................................................................................. 19
Appendix 1 Simulation of Anticipated Process Conditions.........................................................21
Appendix 2:- Determination of Bioburden..................................................................................22
Determination of Bioburden.......................................................................................................22
Appendix 3(GLOSSARY)........................................................................................................... 23
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1. PROTOCOL APPROVAL SHEET
The Validation Protocol shall be Prepared, Reviewed and Approved by the concerned
personnel. It shall be signed and dated as shown below.
Prepared by:
NAME DESIGNATION SIGNATURE DATE

Assistant manager- QA(validation)
Checked by:
Approved by:
2. REVISION HISTORY
REASON FOR
REVISION REVISION DATE REVISION/CHANGE REVISED BY
REQUEST
01 Original release
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3. OBJECTIVE:-
To determine that Ethylene oxide sterilization process consistently performs as intended by

running the system and recording all relevant information. The Data and Test results must
demonstrate that the process meets pre-determined specifications under normal conditions as
well as worst case conditions.
4. SCOPE:-
The scope of validation protocol is to provide sterilization validation strategies for ethylene oxide
sterilization of medical device. This document will show two approaches for reducing or
eliminating bioburden on medical devices. This will also show test method, sampling method
and acceptance criteria used in validating ethylene oxide sterilization of medical devices.
5. RESPONSIBILITY:-
Person Responsibility
Validation team Preparation of protocol
Organization of validation activity
Collecting the samples and sending to QC
Review and interpretation of final results
Preparation of report
Quality control Review of protocol and report
Analyzing the test samples
Reporting and interpretation of results
Production Review of protocol and report
Conducting the validation activity as per the protocol
Quality assurance manager Review and approve the validation protocol
6. STANDARDS FOR REGULATORY REQUIREMENT:-
ISO 11135-1:2007, Sterilization of health care products- Ethylene oxide- Part 1:

Requirements for development, validation and routine control of a sterilization process
for medical devices
ISO 10993-7, Biological evaluation of medical devices- Part 7: Ethylene oxide

sterilization residuals
ISO 11737-1, Sterilization of medical devices Microbiological methods Part 1:

Determination of a population of microorganisms on products
ISO 11737-2, Sterilization of medical devices Microbiological methods Part 2: Tests

of sterility performed in the validation of a sterilization process
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ISO 10993-1:2002, Biological evaluation of medical devices Part 1: Evaluation and

testing
7. LIST OF DOCUMENTS:-
SOP for ethylene oxide sterilization process

SOP for environmental monitoring
SOP for sampling procedure
SOP for testing method
SOP for material handling including biological indicator
Sterilization cycle parameter specification sheet
8. PRE-REQUISITE:-
The following requisite must be fulfilled before the ethylene oxide sterilization process validation.
Equipment qualification:-
Equipment associated with ethylene oxide sterilization process must be qualified

prior to process qualification.
Calibration: - All process sensing, controlling, indicating, and recording devices

on the sterilizer or independent systems associated with sterilizer must be
calibrated and recoded. Calibration program must be documented and detailed
procedure of calibration frequency for all the instruments should be identified.
Product qualification:-
Product and packaging material evaluation: - As per ISO 11135 product and
packaging material must be evaluated for ethylene oxide and humidity
penetration.
Product grouping: - For efficient and cost effective performance validation

similar device can be grouped in to families. A family of products can be
considered to be all those products of similar design and material of construction,
similar bioburden levels but can be consist of different sizes.
Devices can be grouped based on several criteria:
Similarity of materials of construction
Same product design of different sizes, lengths or thickness
Design complexity
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Similar method of manufacture
Multiple combinations of devices
Amount and type of packaging
Selection of family representative: - Each family of products will contain a

number of devices. From these devices, the representative challenge product is
selected. The selected device will present a greater challenge to the sterilization
process and will be the most difficult to sterilize device in the family group and will
be used as the BI carrier.
Following criteria can be used to select the family representative:-
Longest tubing with the smallest lumen
The highest bioburden
Smallest opening into the interior of a device
Most subassemblies,
Most convoluted passageways
Most dense package configuration
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9. Process and Validation Approach:-
Parameters to be Verified
Bioburden Assessment No. of microorganism on product
Loading in chamber Loading Pattern
1. Temperature of Load
Environmental
Preconditioning 2. Humidity of Load
3. Duration of
1. Evacuation rate
Initial Evacuation
2. Number of nitrogen wash
1. Humidity of load
Humidification
2. Temperature of load
3. Duration of
1. Gas injection rate

Gas injection and Gas
dwell 2. Gas Concentration
3. Duration of Exposure
Post exposure gas 1. Vacuum rate

purge and air in bleed
2. Number of Nitrogen wash
1. Temperature
Heated Aeration -
2. Aeration time
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Process Validation Procedure:-

Bioburden assessment: - An understanding of the population of viable
microorganisms on a finished device (bioburden) is necessary and required to support
the validation process. The bioburden assessment includes the total number of
microorganisms with their identities. The identification needs confirmation of gram stain
characteristics and genus. It provides useful information and can be used to monitor
changes over time and as a comparison to organisms recovered during environmental
monitoring. For determination of bioburden Refer to Appendix 2
Selection of Process Challenge Device (PCD):- Process challenge device can be

selected from the family representatives in order to reduce the number of products
tested during the validation. Internal PCD is the most difficult to sterilize device seeded
with a BI. For ease of sample removal an external PCD can be used in the validation.
External PCDs are the external BI test pack that replaces the internal PCD. External
PCD should serves as a resistance greater than or equal to that of the internal PCD. The
external PCD is usually placed on the outside of the sterilization load between cases,
just inside the case or under the stretch wrap.
BI or PCD placement in the product load: - After the product load challenge has
been identified, the BI or PCD positioning and placement can be determined. BIs and
PCDs should be distributed throughout the product load and, as much as possible, in the
same orientation (e.g., vertical).
Following minimum number of BIs/PCDs to be included in each validation cycle (as per
ISO 11135-1: 2007):-
Up to 10 m3, the number of BIs is m3, with a minimum of 5.
From 10 m3 up to 100 m3, the number of additional BIs is one per additional
cubic meter.
Temperature and Humidity sensor in product load: - For humidity check in

preconditioning and conditioning/sterilization one humidity sensor per 2.5 m3, with
minimum number of sensor is two should be placed in sterilization load and it should
include pallet centers, edges and surfaces.
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For temperature monitoring in validation one temperature sensor per cubic meter of
product volume with a minimum of 3 sensors should be placed in the load.
Load Temperature: - ISO 11135-1:2007 requires that the minimum temperature of

product permitted to enter preconditioning be determined. Load temperature prior to
entrance to preconditioning should be determined at the lowest temperature zone. Direct
placement of cold loads into preconditioning can result in excessive water condensation
and load wetting, which can cause product damage and reduced lethality. In process
validation it requires to simulate the worst condition to which the load will ever be
exposed. The anticipated load temperature extremes can be simulated during validation
by following techniques described in the appendices 1.
Identification of the worst case location or cold spot: - temperature is the easiest
variable to measure and monitor, therefore temperature is used as an indicator of the
worst case location in the sterilization load. During the preconditioning or conditioning
phase temperature profile against time of the sterilization load measured and based on
that data the worst case location or cold spot is identified.
Process Qualification Runs:-
For validation, a microbial challenge will be performed to demonstrate the adequacy of

the process to achieve the desired Sterility Assurance Level. Two methods are used
a) Half cycle method (Overkill approach)
b) Bioburden / BI method
a) Half Cycle Method: - During the bioburden assessment, if characterization of

bioburden is not performed and the bioburden level is more than 100 CFU than the
following steps are performed. In this method one fraction cycle, 3 one-half cycle
and one full cycle is performed.
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Place Biological Indicator (BI) with 10 spores of Bacillus atropheus in the PCDs and product
samples in each pallet with humidity and temperature sensor. (Place PCDs to cold spot of the
product load)
Load pallets of sterilization load in to the sterilization chamber
Run fractional cycle according to prescribed cycle parameter
(Use or gas exposure time than full cycle)
Aerate the product
Remove the product sample and PCDs from the load
Perform sterility test on product sample and BIs.
Additionally perform Bacteriostasis/Fungi stasis test on the product sample.
If product sterility samples show survival, repeat the run on new load with elevated gas
exposure time until no survival in product and growth in BI (evaluation of BI appropriateness)
Run 3 consecutive half cycle according to prescribed cycle parameter
(Use gas exposure time than full cycle)
Aerate the load
Remove the product sample and PCDs and perform sterility test on product sample and BIs
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Sterility test should show no growth for both product and BIs samples
Perform one full cycle (routine process exposure time) on the aerated sterilization load.
Place addition product sample for the additional test.
Aerate the load
Remove the product sample and PCDs
Perform following test on product sample
1. LAL test (pyrogen test)
2. Product and packaging functionality test
3. Biocompatibility test
4. Ethylene oxide residue test
b) Bioburden /BI method: - This method requires that bioburden level be

demonstrated to be relatively consistent over time and the resistance of the
bioburden be shown to be equal to, or less resistant than, the resistance of the
biological indicator. During the bioburden assessment, if the characterization of
bioburden performed and level of bioburden is less than or equal to 100 CFU which
indicating a lesser challenge than the BI than the following steps can be followed.
Establish worst case location in the product load based on temperature distribution and humidity
of load
Place BI with known amount of microorganism in the PCDs, temperature and humidity sensor
Perform 5 fractional cycles with graded exposure time (e.g. 0, 4, 8, 12, 16 minutes)
Aerate the load
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Perform sterility test on BI
Calculate rate of inactivation (D value) using survival curve or fractional negative method
Calculate half cycle and full cycle exposure time
Place product sample and PCDs in each pallet with temperature and humidity sensor
Run 3 half cycle according to prescribed cycle parameter
Aeration
Remove product sample and PCDs.
Perform sterility test on product sample and BIs.
Additionally perform fungi stasis/ Bacteriostasis test.
Perform one full cycle (routine process exposure time) on the aerated sterilization load.
Place addition product sample for the additional test.
Aerate the load and remove the product sample and PCDs
Perform following test on product sample
1. LAL test (pyrogen test)
2. Product and packaging functionality test
3. Biocompatibility test
4. Ethylene oxide residue test
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Maintainace of Validation:-
Requalification: -
1. Any significant change in product, manufacturing process, packaging,

sterilization equipment or process, product loading or density may necessitate
requalification. A documented formal review of any change should be undertaken
to make this determination.
2. To ensure continued control, periodic repetition (annually is recommended) of all

or part of the performance validation should be considered.
10. Sampling method and test method:-
Sampling method:-
Sample taken for validating sterilization process should be representative of entire batch
and it should be taken from most critical part of load configuration. Sample must be
taken from each separate packaging system place on.
LIST OF TEST TO BE EXECUTED

No Test Objective References
1 Bioburden To check the liable micro organism AS ISO 11737:1 and ISO
determination in the product. 11737:3
2 Sterility testing Product Sterility Testing is AS ISO 11737:2
determine any viable
microorganisms remain on
product during sterilization
3 Biological Indicator To challenge the sterilization ISO11139:2006
Testing process and are used during
validation and routine processing.
4 Fungi stasis/ To determine the recovery AS ISO 11737:1

Bacteriostasis percentage, microbial growth and
will be used to correct Bioburden
results.
5 LAL test To determine Endotoxin level TGO NO. 50, USP (161)
(pyrogen-end product of gram & (85)
negative bacteria)
6 Product package To evaluate the outer packaging of ISO 11607-1
functionality test a device.
7 Product functionality To evaluate the changes in the ISO 11607-1

test product characteristic.
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7 Biocompatibility Biocompatibility testing is ensure ISO 10993-1- 9 to 12

o Cytotoxicity test that sterile devices are compatible
o Biodegradation test with biological systems
o Carcinogenicity test
o Sensitization test
o Irritation test
8 Ethylene oxide To ensure the amount of ethylene ISO 10993-7
residue test oxide residual levels are within
limits.
11. Test result and acceptances criteria:-
No. Test Acceptance criteria Test results
1 Sterility Testing No Growth for product sample
2 Biological Indicator Fractional cycles:-

Sterility Testing
No growth
Half Cycles:-
No growth
Full Cycles:-
No growth
All positive controls must show

growth
3 Fungi stasis/ Must be negative

Bacteriostasis
4 LAL test Endotoxin and pyrogen must be
absent
5 Product Package Packaging integrity must be intact

Functionality Test
6 Product Product should not show any
Functionality Test degradation, discoloration and
physical changes
7 Biocompatibility Product must be biocompatible

Test
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8 Ethylene Oxide Ethylene oxide residue levels must

Residue Test be in limit at the day of release
9 Process parameters All parameter must meet with

specified cycle parameters
12. Process parameter check steps:-
Environmental preconditioning: -
Parameters to be checked Actual Specification
Temperature of
preconditioning room
Humidity of the
preconditioning room
Duration of preconditioning
Acceptance Criteria met [yes]/ [no]
Comments:
Verified by: _____________________ Date: ________________________
Initial evacuation: -
Point of consideration Actual Specification
The evacuation rate(vacuum) to

maintain the seal integrity
Amount of negative pressure
Number of Nitrogen wash
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Comments:
Verified by: _____________________ Date: ________________________
Humidification: -
Level of moisture
Heat
Comments:
Verified by: _____________________ Date: ________________________
Gas injection and gas dwell: -
Gas concentration
Duration of exposure
Comments:
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Verified by: _____________________ Date: ________________________
Post exposure gas purge and air in bleed: -
Number of Nitrogen wash
Vacuum rate
Comments:
Verified by: _____________________ Date: ________________________
Heated aeration: -
Temperature of room
Aeration duration
Comments:
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Verified by: _____________________ Date: ________________________
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Appendix 1 Simulation of Anticipated Process Conditions
Anticipated load temperature extremes during transport, handling, and storage can be simulated
by following techniques:-
a) Use of cold storageFor example, the product load is stored under refrigeration or
deliberately induced cold temperature. The temperature of the storage area should be
less than or equal to the lowest temperature the product is expected to be exposed to
throughout the year. Product temperature data are recorded during the storage time. The
time in storage should be equal to or greater than the maximum time any product load is
expected to exist under such conditions, or the maximum product temperature while in
storage becomes the minimum acceptable product temperature for a load to be admitted
to the preconditioning phase. This applies to preconditioning or to conditioning when
preconditioning is not used. The use of refrigeration can result in unrealistically low
humidity levels, with the resultant desiccation of the load. A desiccated load can be much
more difficult to sterilize than a non-desiccated load.
b) Temperature modelingData analysis of load temperature studies may be augmented
by modeling to establish the additional time required for starting temperatures lower than
those studied. If lower starting temperatures are allowed, the effect of additional
condensation should be evaluated. For example, at the time of validation, the lowest
product load temperature point is 60 F prior to entering preconditioning. Once in
preconditioning, data yield a temperature profile for the worst-case position showing that
the load temperature increases to 100 F by the end of preconditioning. When the
temperatures over time data are graphed, the linear part of the time/temperature
relationship (which is generally the initial few hours of preconditioning when the
difference between load temperature and preconditioning area temperature is the
greatest) can be used to extrapolate the necessary preconditioning times for sterilization
loads at temperatures lower than 60 F. Using this technique to calculate a minimum
preconditioning time will yield a time that is greater than that actually required. Other
techniques of temperature modeling can be used.
c) Seasonal validationpreconditioning validation can target those seasons that present
the most extreme temperature and humidity conditions. For example, a validation
conducted during the coldest part of the year (which would also present the lowest
ambient humidity level) could yield minimum preconditioning parameters valid for routine
production cycles during the entire year. However, in cases of validations performed
during the summer, it is advisable to validate the preconditioning process with simulated
cold storage, mathematical modeling or to repeat the preconditioning validation at least
once during the winter to confirm the validity of the parameters.
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Appendix 2:- Determination of Bioburden

Determination of Bioburden:-
Procure ten (10) each product samples randomly from three (3) different batches of routine
production processes.
Send to approved test lab for bioburden evaluation (enumeration of aerobes, fungus, and spores.)
At the test lab, perform the bioburden determinations using a standard method as outlined in IS0 1
1737-1 by extracting each device individually and filtering the extract through a sterile bacterial
retentive filter.
A validation of bioburden recovery should be performed. (An additional five non sterile samples
are required.)
Determine the average bioburden per device for each lot, as well as the overall batch average
bioburden
Calculate the overall average bioburden. If a spike (a single value at least 2 X the overall average)
occurs, the spike value may be used rather than the average for the cycle selection.
Sample item portion (SIP) method for bioburden determination:
Based on following criteria product sample portion determined for bioburden assessment.
1) If the product with an average Bioburden equal to or greater than 1.0 whenever practicable, an
entire product (SIP equal to 1.0) should be used for testing in accordance with ISO 11137:2. When
the use of an entire product is not practicable, a selected portion of product (sample item portion)
may be substituted. The SIP should be as large a portion of item as practicable in order to
manipulate in the laboratory, and should be of a size that can be handled during testing.
2) If a product with an average Bioburden equal to or less than 0.9, an entire product (SIP equal to
1.0) shall be used for testing in accordance with ISO 11137:2.
3) If the Bioburden is evenly distributed on and/or in the item, the SIP may be selected from any
portion of the item. If the Bioburden is not evenly distributed, the SIP shall consist of portions of
product selected at random, which proportionally represent each of the materials from which the
product is made. If the Bioburden distribution is known, the SIP may be selected from the portion
of the product that is considered to be the most severe challenge to the sterilization process.
The value of SIP can be calculated on the basis of length, mass, volume or surface area
Basis for SIP Product
Length Tubing (consistent diameter)
Mass Powders
Gowns
Implants (absorbable)
Volume Fluid
Surface area Implants (non-absorbable)

Tubing (variable diameter)
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Appendix 3(GLOSSARY)
ETHYLENE OXIDE STERILIZATION GLOSSARY
Below is an alphabetical list of terms that may be used in discussions of ethylene oxide (EO)
sterilization as recognized by medical device manufacturers inspected by the U.S. FDA. All
definitions or explanations are to be taken as applied specifically in the context of EO
sterilization processing.
Aeration
Part of the sterilization process during which ethylene oxide and/ or its reaction products desorb
(outgas) from the medical device until predetermined levels are reached. This may be
performed within the sterilizer and/or in a separate chamber or room.
Aeration Area
Either a chamber or a room in which aeration occurs. The temperature in the aeration area is
elevated and controlled to assure that the aeration process is accelerated and repeatable. Air is
recirculated in the room to maintain good heat distribution with a portion of the air stripped off
and cleansed with the pollution control equipment. The stripping of the air reduces the amount
of free ethylene oxide resident in the rooms, thus prevents recontamination of product load.
AAMI
Abbreviation for the Association for the Advancement of Medical Instrumentation. AAMI
generates guidelines which are used in the industry for validating, monitoring, and performing
routine EO sterilization processes.
ANSI/AAMI/ISO 11135:2007
International Organization for Standardization (ISO) Guidance document which has been
adopted and published by AMI (and ANSI) to address validation and routine control of ethylene
oxide sterilization.
Bacteriostasis/Fungi stasis Test

Test performed to evaluate the presence of microbial growth inhibiting properties of a health
care product. This test assures that any reaction of the growth medium with the different
materials used in the manufacture of the device will not mask or prevent growth from any viable
organism remaining on the device after being subjected to a sterilization process.
Bioburden
Population of viable microorganisms on a raw material, component, finished product, and/or
package. Unlike the gamma irradiation process, bioburden data is not utilized in ethylene oxide
sterilization to determine the sterilization process. It is collected to quantify the amount of
product contamination, which is then compared to the population of the biological indicator used
in the EO sterilization process.
Biological Indicator
Inoculated carrier contained within its primary pack ready for use and providing a defined
resistance to the specified sterilization process.
Calibration
Comparison of a measurement system or device of unknown accuracy to a measurement
system or device of known accuracy (traceable to national standards) to detect, correlate,
report, or eliminate by adjustment, any variation from the required performance limits of the
unverified measurement system or device. All critical measuring devices utilized on ethylene
oxide sterilizers which may impact the quality of the process are calibrated to traceable national
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or international standards.
Chamber
Enclosed area which only accommodates sufficient product to fill the sterilizer. EO chambers are
constructed of steel or stainless steel and are designed to withstand the extreme pressures and
elevated temperatures utilized in the EO sterilization process.
Commissioning
Obtaining and documenting evidence that equipment has been provided and installed in
accordance with its specification and that it functions within predetermined limits when operated
in accordance with operational instructions. All EO sterilization equipment which complies with
ANSI/AAMI/ISO 11135 is commissioned.
Conditioning
Treatment of product within the sterilization cycle, but prior to sterilant admission, to attain a
predetermined temperature and relative humidity. This part of the sterilization process may be
carried out either at atmospheric pressure or under vacuum (see also preconditioning).
Critical Parameters
Parameters identified as being essential to the sterilization process and requiring monitoring.
D-Value
Time (expressed in minutes) required to achieve inactivation of 90 percent of a population of a
test organism under stated exposure conditions. Also referenced as the D10 Value or decimal
reduction value.
Exposure Time
Time for which the sterilizer chamber is maintained within the specified range for temperature,
sterilant concentration, pressure, and relative humidity. Also may be referred to as the time for
which a medical device (load) is exposed at the specified sterilizing conditions.
Failure
Event in which a component does not perform one or more of its required functions within the
specified limits under specified conditions.
Flushing
Procedure by which sterilant is removed from the load and chamber by either multiple alternate
admissions of filtered air or inert gas and evacuations of the chamber or continuous passage of
filtered air or inert gas through the load and chamber.
Inactivation
Loss of the ability of microorganisms to grow and/or multiply under specified culture conditions.
Indicator
Combination of the indicator agent and its substrate in the form in which it is intended to be
used.
Inoculated Carrier
Carrier on which a defined number of test organisms have been deposited.
Installation Qualification
Obtaining and documenting evidence that equipment has been provided and installed in
accordance with its specifications and that it functions within predetermined limits when
operated in accordance with the operational instructions.
Manufacturer
Natural or legal person packaging or sterilizing a medical device.
Medical Device
Any instrument, apparatus, appliance, material, or other article, whether used alone or in
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combination, including the software necessary for its proper application intended by the
manufacturer to be used for human beings for the purpose of:
diagnosis, prevention, monitoring, treatment, or alleviation of disease;
diagnosis, monitoring, treatment, alleviation of, or compensation for an injury or handicap;
investigation, replacement, or modification of the anatomy or of a physiological process;
control of conception; and which does not achieve its principal intended action in or on the
human body by pharmacological, immunological, or metabolic means, but which may be
assisted in its function by such means.
Microbial Barrier
Ability of the packaging system to prevent the ingress of microorganisms under specified
conditions.
Microbiological Challenge
Biological indicators, biological-indicator test packs, or inoculated product that contain known
populations of microorganisms and can be used in testing sterilization cycles.
Overkill Sterilization Process
Process which is sufficient to provide at least a 12-logarithmic reduction or 12 D inactivation of
an appropriately resistant biological indicator with an established D value.
Package Integrity
Unimpaired physical condition of a final package.
Parametric Release
Declaring product as sterile based on physical and/or chemical process data rather than on the
basis of sample testing or biological indicator results.
Performance Qualification
Obtaining and documenting evidence that the equipment, as commissioned, will produce
acceptable product when operated according to the processing specifications.
Positive Sterility Test
Sterility test samples which exhibit detectable microbial growth after incubation.
Preconditioning
Treatment of product prior to the sterilization cycle in a room or chamber to attain specified limits
for temperature and relative humidity.
Preconditioning Area
Either a chamber or a room in which preconditioning occurs.
Process Lethality
Capability of the sterilization process to destroy microorganisms.
Process Qualification
Obtaining and documenting evidence that the sterilization process will produce acceptable
health care products.
Product
Generic term used to describe raw materials, intermediate products, subassemblies, and
finished medical devices.
Product Qualification
Obtaining and documenting evidence that the health care product will be acceptable for its
intended use after exposure to the sterilization process.
Qualification
Documented evidence that all prescribed design and performance requirements are met.
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Requalification
Repetition of part of all of the validation test requirements for the purpose of reconfirming
process reliability.
Sterilant
Microbicidal agent in the physical form in which it is active.
Sterile
Free from viable microorganisms. In practice, no such absolute statement regarding the
absence of microorganisms can be proven (see sterilization).
Sterility
State of being free from viable microorganisms. In practice, no such absolute statement
regarding the absence of microorganisms can be proven (see sterilization).
Sterility Assurance Level (SAL)
Probability of a viable microorganism being present on a product unit after sterilization. SAL is
normally expressed at 10-n.
Sterility Test
Test performed to determine if viable microorganisms are present.
Sterilization
Validated process used to render a product free from viable microorganisms. In a sterilization
process, the nature of microbial death is described by an exponential function. Therefore, the
presence of viable microorganisms on any individual item can be expressed in terms of
probability. While this probability may be reduced to a very low number, it can never be reduced
to zero. The probability can be expressed as a sterility assurance level (SAL).
Sterilization Cycle
Defined sequence of operational steps designed to achieve sterilization that is carried out in a
sealed chamber. Specifically for EO sterilization, the treatment in a sealed chamber comprising
air removal, conditioning, injection of sterilant, exposure to ethylene oxide, and removal of
ethylene oxide.
Sterilization Load
Goods that are to be or have been sterilized simultaneously in the same sterilization chamber.
Sterilization Process
All treatments which are required to accomplish sterilization, including preconditioning, the
sterilization cycle, and aeration.
Validation
A documented procedure for obtaining, recording, and interpreting the results needed to show
that a process will consistently yield a product complying with predetermined specifications.
Validation is considered as a total process that includes written protocol, evidence that the
equipment as installed meets design criteria and specifications (equipment qualification), use of
calibrated instruments to collect data, and evidence that the equipment can deliver the process
within specified tolerances under established operating conditions and is reproducible as
demonstrated by replicate runs and process challenges (performance qualification).
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Ethylene Oxide Sterilization Validation Protocol

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Ethylene Oxide Sterilization Validation Protocol

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TITLE:- VALIDATION PROTOCOL FOR ETHYLENE OXIDE STERILISATION PROCESS

VALIDATION PROTOCOL NO: EFFECTIVE DATE:-

VALIDATION PROTOCOL OF ETHYLENE OXIDE STERILIZATION PROCESS

Eto Sterilizer manufacturer name:

VALIDATION PROTOCOL NO: EFFECTIVE DATE:-

1. PROTOCOL APPROVAL SHEET.........................................................................................4

VALIDATION PROTOCOL NO: EFFECTIVE DATE:-

VALIDATION PROTOCOL NO: EFFECTIVE DATE:-

1. PROTOCOL APPROVAL SHEET

NAME DESIGNATION SIGNATURE DATE

NAME DESIGNATION SIGNATURE DATE

NAME DESIGNATION SIGNATURE DATE

VALIDATION PROTOCOL NO: EFFECTIVE DATE:-

To determine that Ethylene oxide sterilization process consistently performs as intended by

6. STANDARDS FOR REGULATORY REQUIREMENT:-

ISO 11135-1:2007, Sterilization of health care products- Ethylene oxide- Part 1:

ISO 10993-7, Biological evaluation of medical devices- Part 7: Ethylene oxide

ISO 11737-1, Sterilization of medical devices Microbiological methods Part 1:

ISO 11737-2, Sterilization of medical devices Microbiological methods Part 2: Tests

VALIDATION PROTOCOL NO: EFFECTIVE DATE:-

ISO 10993-1:2002, Biological evaluation of medical devices Part 1: Evaluation and

SOP for ethylene oxide sterilization process

Equipment associated with ethylene oxide sterilization process must be qualified

Calibration: - All process sensing, controlling, indicating, and recording devices

Product grouping: - For efficient and cost effective performance validation

Similarity of materials of construction

Same product design of different sizes, lengths or thickness

VALIDATION PROTOCOL NO: EFFECTIVE DATE:-

Similar method of manufacture

Multiple combinations of devices

Amount and type of packaging

Selection of family representative: - Each family of products will contain a

Following criteria can be used to select the family representative:-

Longest tubing with the smallest lumen

The highest bioburden

Smallest opening into the interior of a device

Most convoluted passageways

Most dense package configuration

VALIDATION PROTOCOL NO: EFFECTIVE DATE:-

9. Process and Validation Approach:-

Bioburden Assessment No. of microorganism on product

Loading in chamber Loading Pattern

1. Gas injection rate

Post exposure gas 1. Vacuum rate

VALIDATION PROTOCOL NO: EFFECTIVE DATE:-

Process Validation Procedure:-

Selection of Process Challenge Device (PCD):- Process challenge device can be

Up to 10 m3, the number of BIs is m3, with a minimum of 5.

Temperature and Humidity sensor in product load: - For humidity check in

VALIDATION PROTOCOL NO: EFFECTIVE DATE:-

Load Temperature: - ISO 11135-1:2007 requires that the minimum temperature of

Process Qualification Runs:-

For validation, a microbial challenge will be performed to demonstrate the adequacy of

a) Half cycle method (Overkill approach)

a) Half Cycle Method: - During the bioburden assessment, if characterization of

VALIDATION PROTOCOL NO: EFFECTIVE DATE:-

Load pallets of sterilization load in to the sterilization chamber

Run fractional cycle according to prescribed cycle parameter

(Use or gas exposure time than full cycle)

Aerate the product

Remove the product sample and PCDs from the load

Perform sterility test on product sample and BIs.

Additionally perform Bacteriostasis/Fungi stasis test on the product sample.

Run 3 consecutive half cycle according to prescribed cycle parameter

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