ASTM E3371-22

Standard Test Method for Measuring the Ability of a Synthetic Polymeric Material to Resist Bacterial Adherence

SIGNIFICANCE AND USE
5.1 This method can be used to evaluate the effectiveness of incorporated or bound anti-adherent agents in synthetic polymeric materials and polymeric coatings intended to reduce the attachment of bacteria to the substrate surface.  
5.2 The synthetic polymeric substrate surface may be tested repeatedly over time for assessment of persistent ability of a material to resist bacterial adherence.  
5.3 This method is to quantify the degree of bacteria colonization of a surface to assess a materials ability to resist bacterial adherence because biofilm formation can contribute to material degradation and malfunction.
SCOPE
1.1 This method is designed to evaluate (quantitatively) the number of bacteria attached to the flat, two-dimensional surfaces of synthetic polymeric materials and polymeric coatings on various substrates that may or may not contain bound or incorporated anti-adherent agents. The method focuses on assessing the ability of the surface to reduce bacterial attachment. Other microorganisms such as yeast and fungal conidia may be tested using this method.  
1.2 This test method quantitatively determines the differences in bacterial adherence seen between synthetic polymeric surfaces that allow bacterial adherence and those that do not, comparing the number of organisms recovered from the control surface to the number recovered from the test specimen surface after the contact time. Knowledge of microbiological techniques is required for these procedures.  
1.3 This test method specifies proper methods for measuring the ability of a synthetic polymeric material to resist adherence against specified organism. Due to individual sensitivities, the result of one test organism might not be applicable for other organisms.  
1.4 This test method is designed to measure the potential ability to resist bacterial adherence of a non-porous surface compared directly to a polyester control panel known to support bacterial adherence under specific testing conditions.  
1.5 Antimicrobial treated non-porous surfaces may demonstrate ability to resist bacterial adherence in this method. This method does not purport to differentiate between anti-adherence and antimicrobial activity nor is it designed to reflect specific end-use or environmental conditions. Any product that demonstrates ability to resist bacterial adherence in this method should be measured for antimicrobial activity using a separate test technique such as Test Method E2180 or ISO 22196.  
1.6 The method focuses on assessing the ability of synthetic polymeric materials and polymeric coatings on various substrates to reduce bacterial attachment. The specimen with absorbing or adhesive surfaces may be unable to be disinfected properly before testing, or may trap inoculated organism during recovery process and thus lead to a false result. This method does not apply to specimens with absorbent or adhesive surfaces.  
1.7 The values stated in SI units are to be regarded as standard. The values given in parentheses after SI units are provided for information only and are not considered standard.  
1.8 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use.  
1.9 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

General Information

• Status: Published
• Publication Date: 30-Sep-2022
• ICS: 07.100.99 - Other standards related to microbiology
• ICS: 83.080.01 - Plastics in general
• Technical Committee: E35 - Pesticides, Antimicrobials, and Alternative Control Agents
• Drafting Committee: E35.15 - Antimicrobial Agents

Relations

• Refers: ASTM E2180-24 - Standard Test Method for Determining the Activity of Incorporated Antimicrobial Agent(s) In Polymeric or Hydrophobic Materials
• Effective Date: 01-Apr-2024
• Refers: ASTM E2756-19 - Standard Terminology Relating to Antimicrobial and Antiviral Agents
• Effective Date: 01-Nov-2019
• Refers: ASTM E2180-18 - Standard Test Method for Determining the Activity of Incorporated Antimicrobial Agent(s) In Polymeric or Hydrophobic Materials
• Effective Date: 01-May-2018
• Refers: ASTM E2756-18 - Standard Terminology Relating to Antimicrobial and Antiviral Agents
• Effective Date: 01-Apr-2018
• Refers: ASTM E3151-18 - Standard Test Method for Determining Antimicrobial Activity and Biofilm Resistance Properties of Tube, Yarn, or Fiber Specimens
• Effective Date: 01-Feb-2018
• Refers: ASTM E2180-07(2017) - Standard Test Method for Determining the Activity of Incorporated Antimicrobial Agent(s) In Polymeric or Hydrophobic Materials
• Effective Date: 01-Apr-2017
• Refers: ASTM E177-14 - Standard Practice for Use of the Terms Precision and Bias in ASTM Test Methods
• Effective Date: 01-May-2014
• Refers: ASTM E177-13 - Standard Practice for Use of the Terms Precision and Bias in ASTM Test Methods
• Effective Date: 01-May-2013
• Refers: ASTM E2180-07(2012) - Standard Test Method for Determining the Activity of Incorporated Antimicrobial Agent(s) In Polymeric or Hydrophobic Materials
• Effective Date: 01-Apr-2012
• Refers: ASTM E177-10 - Standard Practice for Use of the Terms Precision and Bias in ASTM Test Methods
• Effective Date: 01-Oct-2010
• Refers: ASTM E2756-10 - Standard Terminology Relating to Antimicrobial and Antiviral Agents
• Effective Date: 01-May-2010
• Refers: ASTM E177-08 - Standard Practice for Use of the Terms Precision and Bias in ASTM Test Methods
• Effective Date: 01-Oct-2008
• Refers: ASTM E2180-07 - Standard Test Method for Determining the Activity of Incorporated Antimicrobial Agent(s) In Polymeric or Hydrophobic Materials
• Effective Date: 01-Nov-2007
• Refers: ASTM E177-06b - Standard Practice for Use of the Terms Precision and Bias in ASTM Test Methods
• Effective Date: 15-Nov-2006
• Refers: ASTM E177-06a - Standard Practice for Use of the Terms Precision and Bias in ASTM Test Methods
• Effective Date: 01-Nov-2006

Overview

ASTM E3371-22 is the Standard Test Method for Measuring the Ability of a Synthetic Polymeric Material to Resist Bacterial Adherence, developed by ASTM. This international standard provides a quantitative method to evaluate the effectiveness of synthetic polymeric materials and coatings in resisting the attachment of bacteria on flat, non-porous surfaces. The method is significant in assessing anti-adherent performance, particularly for materials intended to minimize bacterial colonization and subsequent biofilm formation, which can lead to product degradation or failure.

This test method is especially valuable for industries utilizing synthetic polymers in environments where bacterial contamination is a concern. It can be applied to materials both with and without antimicrobial or anti-adherent agents and supports the development and verification of hygienic surfaces in critical applications such as medical devices, food packaging, and consumer goods.

Key Topics

• Bacterial Adherence Measurement: Provides a standardized procedure to measure and compare the number of culturable bacteria adhering to synthetic polymeric surfaces versus control surfaces.
• Evaluation of Anti-Adherent Agents: Enables quantitative assessment of incorporated or bound agents designed to inhibit bacterial attachment.
• Surface Suitability: Applies specifically to flat, non-porous polymeric surfaces and coatings; absorbent or adhesive surfaces are excluded due to potential for contamination or inaccurate results.
• Repeatable Assessment: Supports repeated testing over time to gauge the persistent effectiveness of materials in resisting bacterial adherence, offering insight into long-term performance.
• Microbiological Techniques: Requires knowledge of microbiological methods, including sterile techniques, inoculum preparation, incubation, sonication, and data interpretation.
• Result Comparison: Involves direct comparison to polyester control panels known to support bacterial adherence under defined conditions.
• Limitations: Does not differentiate between anti-adherence and antimicrobial activity; antimicrobial efficacy must be evaluated using separate standards such as ASTM E2180 or ISO 22196.

Applications

ASTM E3371-22 offers practical value for several industry sectors, including:

• Medical Devices: Assessment of catheters, tubing, and devices where bacterial colonization can cause infections or device failure. Ensures that materials used in these products are capable of resisting initial bacterial attachment.
• Food and Beverage Packaging: Evaluation of packaging films and containers for their ability to resist bacterial contamination, thus promoting hygiene and extending shelf life.
• Consumer Products: Testing of personal care containers, household goods, and other plastic items where hygiene is important.
• Building and Construction: Assessment of polymeric surfaces in environments demanding high cleanliness, such as hospitals and laboratories.
• Material Development and R&D: Support for research and quality assurance teams in developing and validating new anti-biofouling materials.

Related Standards

For comprehensive evaluation of antibacterial and anti-adherent properties, consider these related standards:

• ASTM E2180: Test Method for Determining the Activity of Incorporated Antimicrobial Agents in Polymeric or Hydrophobic Materials – measures antimicrobial efficacy.
• ASTM E3151: Test Method for Determining Antimicrobial Activity and Biofilm Resistance Properties of Tube, Yarn, or Fiber Specimens – focuses on three-dimensional or fibrous substrates.
• ISO 22196: Measurement of Antibacterial Activity on Plastics and Other Non-Porous Surfaces – internationally recognized for antimicrobial testing on non-porous materials.
• ASTM E2756: Terminology Relating to Antimicrobial and Antiviral Agents – provides definitions used in this field.

Keywords

bacterial adherence, anti-adherent agents, polymeric materials, antibacterial, biofilm resistance, synthetic polymers, ASTM E3371, surface hygiene, non-porous coatings, standard test method

ASTM E3371-22 is a valuable standard for organizations seeking to enhance the hygienic performance of synthetic polymer surfaces, offering a reliable framework for testing and comparison.

Standards Content (Sample)

This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the
Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.
Designation: E3371 − 22
Standard Test Method for
Measuring the Ability of a Synthetic Polymeric Material to
Resist Bacterial Adherence
This standard is issued under the fixed designation E3371; the number immediately following the designation indicates the year of
original adoption or, in the case of revision, the year of last revision.Anumber in parentheses indicates the year of last reapproval.A
superscript epsilon (´) indicates an editorial change since the last revision or reapproval.
INTRODUCTION
Synthetic polymeric materials, used in packaging for food containers, personal care products and
other items, may have inherent antimicrobial properties. Others may contain antimicrobial additives.
Many methods, such as Test Method E2180 are used to determine quantitative bacterial reductions
caused by these additives. Test Method E3151 specifically examines the ability of tubular, yarn and
fiber specimens to resist bacterial colonization/adherence. However, these methods do not quantita-
tively measure the number of culturable bacteria adhering to the flat surfaces of these synthetic
polymeric materials, as measured in this test method.
1. Scope 1.5 Antimicrobial treated non-porous surfaces may demon-
strate ability to resist bacterial adherence in this method. This
1.1 This method is designed to evaluate (quantitatively) the
method does not purport to differentiate between anti-
number of bacteria attached to the flat, two-dimensional
adherenceandantimicrobialactivitynorisitdesignedtoreflect
surfaces of synthetic polymeric materials and polymeric coat-
specificend-useorenvironmentalconditions.Anyproductthat
ings on various substrates that may or may not contain bound
demonstratesabilitytoresistbacterialadherenceinthismethod
or incorporated anti-adherent agents. The method focuses on
should be measured for antimicrobial activity using a separate
assessing the ability of the surface to reduce bacterial attach-
ment. Other microorganisms such as yeast and fungal conidia test technique such as Test Method E2180 or ISO 22196.
may be tested using this method.
1.6 Themethodfocusesonassessingtheabilityofsynthetic
1.2 This test method quantitatively determines the differ-
polymeric materials and polymeric coatings on various sub-
ences in bacterial adherence seen between synthetic polymeric
strates to reduce bacterial attachment. The specimen with
surfaces that allow bacterial adherence and those that do not,
absorbingoradhesivesurfacesmaybeunabletobedisinfected
comparingthenumberoforganismsrecoveredfromthecontrol
properly before testing, or may trap inoculated organism
surfacetothenumberrecoveredfromthetestspecimensurface
during recovery process and thus lead to a false result. This
after the contact time. Knowledge of microbiological tech-
method does not apply to specimens with absorbent or adhe-
niques is required for these procedures.
sive surfaces.
1.3 Thistestmethodspecifiespropermethodsformeasuring
1.7 The values stated in SI units are to be regarded as
theabilityofasyntheticpolymericmaterialtoresistadherence
standard. The values given in parentheses after SI units are
against specified organism. Due to individual sensitivities, the
providedforinformationonlyandarenotconsideredstandard.
result of one test organism might not be applicable for other
organisms.
1.8 This standard does not purport to address all of the
safety concerns, if any, associated with its use. It is the
1.4 This test method is designed to measure the potential
ability to resist bacterial adherence of a non-porous surface responsibility of the user of this standard to establish appro-
compared directly to a polyester control panel known to priate safety, health, and environmental practices and deter-
support bacterial adherence under specific testing conditions.
mine the applicability of regulatory limitations prior to use.
1.9 This international standard was developed in accor-
dance with internationally recognized principles on standard-
This test method is under the jurisdiction of ASTM Committee E35 on ization established in the Decision on Principles for the
Pesticides, Antimicrobials, and Alternative Control Agents and is the direct
Development of International Standards, Guides and Recom-
responsibility of Subcommittee E35.15 on Antimicrobial Agents.
mendations issued by the World Trade Organization Technical
Current edition approved Oct. 1, 2022. Published November 2022. DOI:
10.1520/E3371–22. Barriers to Trade (TBT) Committee.
Copyright © ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959. United States
E3371 − 22
2. Referenced Documents 4. Summary of Test Method
2.1 ASTM Standards: 4.1 A fixed inoculum volume (0.4 mL 6 0.02 mL) at a
known concentration is pipetted onto the surface of the test
E177Practice for Use of the Terms Precision and Bias in
ASTM Test Methods specimens and controls and then incubated under appropriate
conditions for the test organism.
E2180Test Method for Determining the Activity of Incor-
porated Antimicrobial Agent(s) In Polymeric or Hydro-
4.2 After the specified contact time (24 h), the testing
phobic Materials
surface is gently washed with sterile saline with the aid of
E2756Terminology Relating toAntimicrobial andAntiviral
rocker to remove the unattached bacteria.
Agents
4.3 The remaining attached bacteria are collected by soni-
E3151Test Method for Determining Antimicrobial Activity
cation and then counted.
and Biofilm Resistance Properties of Tube,Yarn, or Fiber
Specimens 4.4 Ability of a material to resist bacterial adherence is
calculated from the numbers of bacteria recovered from the
2.2 ISO Standard:
control and specimen surfaces.
ISO22196MeasurementofAntibacterialActivityonPlastic
and Non-Porous Surfaces
5. Significance and Use
2.3 FDA Manuals:
5.1 Thismethodcanbeusedtoevaluatetheeffectivenessof
Bacteriological Analytical Manual (BAM)
incorporated or bound anti-adherent agents in synthetic poly-
meric materials and polymeric coatings intended to reduce the
3. Terminology
attachment of bacteria to the substrate surface.
3.1 Defintions:
5.2 The synthetic polymeric substrate surface may be tested
3.1.1 For definitions of terms used in this test method, refer
repeatedly over time for assessment of persistent ability of a
to Terminology E2756.
material to resist bacterial adherence.
3.2 Definitions of Terms Specific to This Standard:
5.3 This method is to quantify the degree of bacteria
3.2.1 anti-adherence, n—term describing a state where the colonization of a surface to assess a materials ability to resist
attachment of bacteria to the surface of products is suppressed
bacterial adherence because biofilm formation can contribute
ordescribingtheeffectofanagentwhichreducestheabilityof
to material degradation and malfunction.
bacteria to attach to product surfaces.
6. Apparatus
3.2.2 anti-adherent, adj—term describing a state where the
6.1 Autoclave, capable of producing 103 kPa (15 psi) of
attachment of bacteria to the surface of products is suppressed
steam pressure at 121°C and maintaining it for a minimum of
ordescribingtheeffectofanagentwhichreducestheabilityof
15 minutes.
bacteria to attach to product surfaces.
6.2 Balance, capable of weighing to 6 0.01 g.
3.2.3 ability to resist bacterial adherence, n—percent
reduction, calculated from the number of bacteria recovered
6.3 Binder clip, 15mm to 25 mm width, or equivalent.
from the surface of the test specimen and the reference control
6.4 Cell counting chamber (optional).
after inoculation with bacteria, incubation, saline wash, and
6.5 Clear polyester panels (Leneta – Order #P300-7G or
quantification of attached organisms.
equivalent)
3.2.4 antimicrobial, n—difference in the logarithm of the
6.6 Colony counter (optional).
culturable cell count found on supernatant of an antibacterial-
treated product and an untreated product after inoculation with
6.7 Cotton swabs, sterile.
and incubation of bacteria.
6.8 Culture tubes, screw top, sterile, or equivalent.
3.2.5 reference control material, n—synthetic polymeric
6.9 Cuvettes.
material, that does not inhibit the attachment of the test
6.10 Forceps, sterile.
bacteria.
3.2.5.1 Discussion—This material is commonly referred to
6.11 Incubator, set at required temperature (35°C 6 1 °C).
as the untreated control and is used for test validation calcu-
6.12 Inoculating loops, sterile, 4mm ring diameter.
lations.
6.13 Petri dishes, (15 mm× 100 mm), sterile.
6.14 pH meter, any capable of measuring to 0.2 units.
For referenced ASTM standards, visit the ASTM website, www.astm.org, or
6.15 Pipettes, (100 µL and 1000 µL) positive displacement.
contact ASTM Customer Service at [e-mail]. For Annual Book of ASTM
Standards volume information, refer to the standard’s Document Summary page on 6.16 Pipette tips, sterile.
the ASTM website.
6.17 Rocker, with rocking angle of 8° - 10°, and shaking
Available from International Organization for Standardization (ISO), ISO
Central Secretariat, Chemin de Blandonnet 8, CP 401, 1214 Vernier, Geneva, speed of 30 - 50 rpm.
Switzerland, https://www.iso.org.
6.18 Round stick, 4mm in diameter and min 30mm, or
Available from U.S. Food and Drug Administration (FDA), 10903 New
Hampshire Ave., Silver Spring, MD 20993, http://www.fda.gov. equivalent.
E3371 − 22
involved. The suggested bacteria include Escherichia coli ATCC 8739,
6.19 Sample bag, sterile, with a width of 76mm and a
Pseudomonas aeruginosa ATCC 9027, Staphylococcus aureus ATCC
minimal height of 127mm, or equivalent.
6538P and Staphylococcus epidermidis ATCC 12228.
6.20 Spectrophotometer, set at 600 nm.
NOTE 4—Other microorganisms such as yeast or fungal conidia may
also be tested using this procedure. If other species are used, the species
6.21 Spiral plater (optional).
andthereasonfortheiruseshouldbedocumentedinthetestreport.Ifthe
test specimens are to be challenged with a combination of organisms, no
6.22 Syntheticpolymericcoverfilm,40mm×40mm.Film
more than 3 organisms should be evaluated at one time.
shallnotaffectbacterialgrowthorabsorbwater.(Polyethylene
film that is 0.05mm – 0.1 mm thick is recommended.
9. Procedure
Stomacher bag or Whirl-Pac bag film or (equivalent) has been
9.1 Preparation of Test Specimens:
found to be acceptable).
9.1.1 Testing shall be performed on at least three specimens
6.23 Test materials, sterile if specified by interested parties.
from each untreated and treated test material. Include the three
polyester panels for the reference control material. When
6.24 Ultrasoniccleaner,anycapableofawattdensityoutput
testingaseriesofanti-adherenttreatmentsforasinglepolymer,
of 100 watts per gallon to 133 watts per gallon at a frequency
all anti-adherent specimens must be compared to the untreated
of 42kHz 66%.
reference control specimens. All tests are conducted at the
6.25 Vortex mixer.
same time using the same test inoculum.
7. Reagents and Materials
NOTE 5—An untreated control of the same material is often not
available, or if it is, may have some inherent properties that inhibit
7.1 Purity of Reagents—Reagent grade chemicals shall be
bacterialattachment.Therefore,thereferencecontrollistedinthismethod
used in all tests. Unless otherwise indicated, it is intended that
is required and used in the calculations. If calculations are performed
all reagents shall conform to the specifications of the Commit-
against the submitted untreated control, this should be stated in the test
tee onAnalytical Reagents of theAmerican Chemical Society, report.
where such specifications are available. Other grades may be
9.1.2 Prepare flat 50mm 6 2mm×50mm 6 2mm
used, provided it is first ascertained that the reagent is of
specimens of the treated and untreated test materials. Speci-
sufficiently high purity to permit its use without lessening the
mens should be of uniform thickness. Unless otherwise
accuracy of the determination.
specified, the standard size of the plastic cover film shall be a
square of 40 mm × 40 mm for the 50 mm × 50 mm test
7.2 Purity of Water—Unless otherwise indicated, references
specimen.Ifthetestspecimenisnotofastandardsize,thenthe
towatershallbeunderstoodtomeandistilledwaterorwaterof
size of the plastic cover film shall be reduced in direct
equal purity.
proportion.Donot,however,reducethesizeoffilmtolessthan
7.3 Media:
900 mm and the edges of the cover film shall always be 2.5
7.3.1 Liquid Growth Medium, sterile (See Annex A1).
mmto5.0mminsidetheedgeofthetestspecimenonallsides.
7.3.2 SolidGrowthandPlatingMedium,sterile(SeeAnnex
If the size of the cover film differs from 40 mm × 40 mm, the
A1).
actual size used shall be stated in the test report.
7.3.3 InoculationMedium,1/500NutrientBroth,(pH7.0 6
NOTE 6—If it is difficult or impossible to cut the product into a square
0.2), prepared as described in A1.2. Dilute the nutrient broth
of this size, then test specimens of other sizes and shapes may be used, as
with distilled or deionized water to a 500- fold, dispense into
long as they can be covered with a plastic cover film of surface area
suitable flasks or tubes prior to autoclaving.
2 2
between 900 mm and 1600 mm . It is preferable to prepare test
specimens from the final product itself. However, if the shape of the
NOTE 1—Alternate neutralizing agents may be required and dependent
productpreventsthis,thenthetestspecimensmaybepreparedinaformat
upon the active agents may be present in the specimens being tested.
suitable for testing using the same raw materials and processing methods
NOTE 2—Media formulas are from the FDA’s Bacteriological Analyti-
as are normally used for the product. If the test specimen differs from the
cal Manual (BAM). Other low-nutrient equivalent formulations may be
50 mm × 50 mm square dimensions, the actual dimensions used shall be
used but should be stated in the test report.
stated in the test report.
NOTE 7—When preparing specimens, take care to avoid contamination
8. Test Organism
withmicroorganismsorextraneousorganicdebris.Similarly,donotallow
8.1 Bacteria—Gram-positive bacteria Staphylococcus au-
specimens to contact each other. Test specimens may be cleaned/
disinfected/sterilized prior to testing (for example, by wiping with 70 %
reus ATCC 6538.
ethanol in water or exposure to 30 minutes of UV light) as agreed upon
8.2 Cultures of the test organism shall be maintained using
between all parties involved and stated in the test report.
NOTE8—Cleaningofthetestspecimencancausesoftening;dissolution
appropriate microbiological practices.
of the surface coating or elution of components so should be avoided. If
NOTE 3—Other bacteria may be tested, as agreed upon by all parties
cleaningisrequiredduetogrosscontamination,thecleaningmethodshall
be stated in the test report.
9.2 Preparation of test inoculum:
ACS Reagent Chemicals, Specifications and Procedures for Reagents and
9.2.1 Usingasterileinoculatingloop,transferbacteriafrom
Standard-Grade Reference Materials, American Chemical Society, Washington,
the stock culture to a sterile Petri dish with NutrientAgar and
DC. For suggestions on the testing of reagents not listed by theAmerican Chemical
Society, see Analar Standards for Laboratory Chemicals, BDH Ltd., Poole, Dorset,
incubateat35°C 61°Cfor18hto24h.Fromthisculture,use
U.K., and the United States Pharmacopeia and National Formulary, U.S. Pharma-
a sterile inoculating loop to transfer bacteria onto fresh
copeial Convention, Inc. (USPC), Rockville, MD.
Nutrient Agar and incubate at 35°C 61°Cfor18hto20h.
ATCC is a registered trademark and ATCC 6538 is a trademark of American
Type Culture Collection, Manassas, VA. Using a sterile inoculating loop or cotton swab, suspend the
E3371 − 22
grown organism in 1/500 NB to target5×10 CFU/mL 6 0.5 9.5 Recovery of bacteria from specimens after contact time:
logconcentrationofbacteria.Aspectrophotometeranddilution
9.5.1 Immediately after the contact time, remove the cover
in 1/500 NB may be used. The maximum number of sub-
films from all specimens with sterile forceps. All specimens
passages shall be no greater than 3 from freeze or stock plate.
should be washed as follows: (1) hold them up and fill 20mL
of 0.9% saline into their original P
...