Guidance for Industry
Guidance for Manufacturers
Seeking Marketing Clearance
of Ear, Nose, and Throat
Endoscope Sheaths Used as
Protective Barriers
Document issued on: March 12, 2000
This document supersedes the Guidance for the Content of Premarket Notifications for
Disposable, Sterile, Ear, Nose and Throat Endoscope Sheaths with Protective Barrier
Claims, which was issued on October 21, 1996
U.S. Department Of Health And Human Services
Food and Drug Administration
Center for Devices and Radiological Health
Ear, Nose, and Throat Devices Branch
   Division of Ophthalmic and ENT Devices Office of Device Evaluation
Preface
Public Comment
Comments and suggestions may be submitted at any time for Agency consideration to
Harry Sauberman, P.E., HFZ 460, 9200 Corporate Blvd. Rockville, MD 20850.
Comments may not be acted upon by the Agency until the document is next revised or
updated.  For questions regarding the use or interpretation of this guidance contact Karen
Baker, MSN at 240-276-4242 or by electronic mail at karen.baker@fda.hhs.gov.
Additional Copies
Additional copies of this guidance document may be obtained via the World Wide
Web/CDRH home page: http://www.fda.gov/cdrh/ode/guidance/954.pdf or CDRH Facts
on Demand at 1-800-899-0381 or 301-827-0111, specify number 954 when prompted
for the document shelf number.
2 Table of Contents
Foreword_______________________________________________________________ 1
References______________________________________________________________ 1
Definitions ______________________________________________________________ 1
List of Symbols __________________________________________________________ 2
I.  Introduction___________________________________________________________ 2
II.  Classification_________________________________________________________ 4
III.   Possible Predicate Devices ____________________________________________ 4
IV.  Required 510(k) Information____________________________________________ 4
Sampling Procedure________________________________________________________________ 13
Detection Limit ___________________________________________________________________ 13
Presentation of Results _____________________________________________________________ 13
Report Forms_____________________________________________________________________ 13
Control to test challenge virus stability _________________________________________________ 14
3 1
Guidance  for Manufacturers Seeking
Marketing Clearance of Ear, Nose And Throat
Endoscope Sheaths Used As Protective
Barriers
Foreword
This guidance document provides detailed information to assist manufacturers in the preparation of 510(k)
premarket notification submissions for endoscope sheaths used as protective barriers.  This guidance replaces
all previous 510(k) guidance documents that relate to the subject device.
References
Regulatory Requirements for Medical Gloves, A Workshop Manual. HHS Publication  (FDA) 93-4257,  May 1993. 
Testing Guidance for Male Condoms Made From New Materials - Division of Reproductive,  Abdominal, Ear, 
Nose and Throat and Radiological Devices, Obstetrics and Gynecology Devices Branch, June 29, 1995. 
Labeling of reusable Medical Devices for Reprocessing in Heath Care Facilities: FDA Reviewer Guidance. ODE 
Bluebook Memo K90-1 "510(k) Sterility Review Guidance (2/12/90)
MIL-STD-105E called Sampling Procedure and Tables for Inspection by Attributes, dated May 10, 1989.
FDA-modified ISO 10993, Part 1 Matrix
ASTM F1671-97a Standard Test Method for Resistance of Materials Used in Protective Clothing to Penetration 
by Blood-Borne Pathogens Using Phi-X174 Bacteriophage Penetration as a Test System
ODE Bluebook Memo G91-1 "Device Labeling Guidance  (3/18/91)
Definitions
Diopter ring - section on the distal end of the device that permits individual adjustments for precise focusing
Optical lens  the distal end of the endoscope through which provides vision, magnification and light.
1 This document is intended to provide guidance.  It represents the Agencys current thinking on the above.   It
does not create or confer any rights for or on any person and does not operate to bind FDA or the public.  An
alternative approach may be used if such approach satisfies the requirements of the applicable statute,
regulations, or both. Rod lens - fiberoptic glass bundles that are encased in a non-flexible cylinder.
Surrogate microbe  for the purpose of this guidance, the surrogate microbe is Phi-X 174, a bacteriophage that is
not pathogenic to humans but serves to simulate viruses that are pathogenic to humans.
Bacteriophage  a type of virus that infects bacteria.
Titer  quantity of a substance required to react with, or to correspond to, a given amount of another substance
Viscosity  thickness of consistency of a liquid, resistance to flow.
List of Symbols
pfu - plaque forming units
mL - milliliter
(L - microliter
pfu/10(L - plaque forming units per 10 microliters
pfu/100mL - plaque forming units per 100 microliters
I.  Introduction
A.  Introduction and General Information
This document reflects the current review guidance for Ear, Nose and Throat (ENT) endoscope sheath devices.
It is based on 1) current scientific knowledge; 2) clinical experience; 3) previous submissions by manufacturers
to the Food and Drug Administration (FDA);  4) the Safe Medical Devices Act of 1990; the Food and Drug
Administration Modernization Act; and FDA regulations in the Code of Federal Regulations (CFR).  As
advances are made in science and medicine, and changes occur in device regulation, these review criteria will be
re-evaluated and revised as necessary.
This document is an adjunct to the CFR and other FDA guidance documents for the preparation and review of
510(k) submissions. It does not supersede those publications, but provides additional clarification on what is
necessary before the FDA can clear these devices for marketing. The 510(k) submission must provide evidence
that the device is substantially equivalent to a predicate device legally marketed in the United States. In some
cases, the performance of the device can be established by comparison of the device to a standardized reference
method.
The primary reference for the information required in a premarket notification (510(k)) for a medical device is
found in 21 CFR 807.87.  Substantial equivalence to a legally marketed device is to be established with respect to,
but not limited to, intended use, design, energy used/delivered, materials, performance, safety, effectiveness,
labeling, and other applicable characteristics.
B.  Generic Description of ENT Endoscopes
Nasopharyngoscope (flexible or rigid) and accessories, 21 CFR 874.4760, Class II, Procode 77-EOB
Bronchoscope (flexible or rigid) and accessories, 21 CFR 874.4680, Class II, Procode 77-EOQ
Esophagoscope (flexible or rigid) and accessories, 21 CFR 874.4710, Procode 77-EOX
2 Mediastinoscope and accessories, 21 CFR 874.4720, Class II,  Procode 77-EWY
Otoscope, 21 CFR 874.4770, Class I(exempt), Procode 77-ERA
Endoscopes are commonly used in the examination of the ear, nose and throat.  These endoscopes may be
flexible or rigid and made from flexible plastic or stainless steel.  The proximal end of the endoscope most often
includes an eyepiece with a diopter ring that permits individual adjustments for precise focusing.  The
endoscope contains fiberoptic glass bundles for illumination.  The insertion tube of the flexible endoscope
contains the angulation wires controlled by handles at the proximal end of the device for up/down direction of
view.  The degree of flexion may vary, with a maximum of 180.  The rigid scopes may have varying angles of
view from 0 to greater than 100.  The distal end of the endoscope may contain the optical lens or it may be the
rod lens style.
C.  Generic Description of an Endoscope Sheath
The endoscope sheath is most often packaged as a sterile, disposable covering for various types of ENT
endoscopes.  The sheath is tubular in shape, is made from a transparent, flexible polymer or thermoplastic
material and has an optical lens at the distal end.  The sheath may contain air, water or suction channels but most
often is single channel.  To fit the sheath over the endoscope it must first be inflated, rolled, slipped on or
otherwise applied.  Inflation may be accomplished by positive pressure, negative pressure or other means.  The
process is repeated to remove the sheath from the endoscope.  Whatever the method of attachment and removal,
it must be such that the sterility and/or integrity of the sheath is not compromised.  When attached to the
endoscope, the sheath fits snugly and does not add appreciably to the outside diameter of the endoscope and
does not impair articulation or light refraction or cause visual distortion.
The purpose of a sheath is to provide a covering that helps prevent the transmission of pathogens from one
patient to another.  Use of a sheath may also reduce the time needed for disinfection, thereby expediting
endoscope reprocessing.
Manufacturers of endoscope sheaths used as protective barriers will be asked to provide laboratory data that
demonstrates the device to be impermeable to penetration by microorganisms.  It should also be demonstrated
that sheath application and removal processes can be accomplished with aseptic technique.
D.  Regulatory Information
Historically, applications for ear, nose and throat endoscope sheaths have been reviewed under the premarket
notification (510(k)) regulation. For additional information regarding regulatory requirements and testing
guidance, the applicant may wish to review the following documents available from the Division of Small
Manufacturers Assistance (DSMA):
May 1993 - Regulatory Requirements for Medical Gloves, A Workshop Manual. HHS 
Publication (FDA) 93-4257.
June 29, 1995 - Testing Guidance for Male Condoms Made From New Materials - Division of 
Reproductive, Abdominal, Ear, Nose and Throat and Radiological Devices, Obstetrics and
Gynecology Devices Branch.
3 II.  Classification
The appropriate panel is Ear, Nose and Throat.  Endoscope sheaths are also reviewed by the Gastroenterology
Branch of Division of Reproductive, Abdominal and Radiological Devices.
Sheaths for ENT endoscopes are considered as accessories and are regulated as Class II devices under section
513 of the Federal Food, Drug, and Cosmetic Act.  Although otoscopes are Class I, exempt, sheaths for
otoscopes, with barrier claims will be regulated as Class II devices.
III.   Possible Predicate Devices
It is the manufacturer's responsibility to identify an appropriate predicate device.  Below is a list of several
510(k)s for sheaths that may serve as an appropriate predicate.
K921244, Vision Sciences, Disposable Protective Sheath
K925421, Vision Sciences, Endosheath
K933247, Vision Sciences, Protective Sheaths
K933672, Xomed-Treace, Scope Sheath
K940028, Xomed-Treace, Endoscope Sheath
K963795, Vision Sciences, Endosheath
K990354, Vision Sciences, Endosheath
IV.  Required 510(k) Information
FDA regulations (21 CFR 807.87) prescribe information that must appear in each 510(k) submission.  This
information includes:
A.  Sponsor/Manufacturer Information
The name, contact person, address, telephone number, and (if available) facsimile number of both the sponsor of
the 510(k) and (if different from the sponsor) the device manufacturer.
B.  Proposed Device
The trade name or proprietary name of the device proposed for marketing, as well as the common device name.
C.  Predicate Device
Legally marketed device(s) to which the proposed device is being compared.  To be as specific as possible, the
510(k) should include the following information to identify each predicate device and support the claim of
substantial equivalence.
Trade/proprietary name,
Common/usual name,
Model number,
Manufacturer,
510(k) reference number (if known)
Intended use
4 Technological characteristics/performance specification, and
Labeling
D.  Truthful and Accuracy Statement stating that submitter believes, to the best of his/her
knowledge, that all data  and information submitted are truthful and accurate, and that no material fact has been
omitted as set forth in 21 CFR 807.87(j).
E.  510(k) Summary or Statement
The Safe Medical Devices Act of 1990 (SMDA) requires all persons submitting a premarket notification to
include either (1) a summary of safety and effectiveness information upon which an equivalence determination
could be based (510(k) Summary); OR (2) a statement that safety and effectiveness information will be made
available to interested persons upon request (510(k) Statement).  Safety and effectiveness information refers to
information in the premarket notification submission, including adverse safety and effectiveness information that
is relevant to an assessment of substantial equivalence. The information can be descriptive in content and may
include data about the new and predicate device(s), about safety or performance or about clinical test results
information. The requirements for the content of a 510(k) Summary is found in 21 CFR 807.92.
F.  Device Description
1.  Reason for the Submission
The sponsor should clearly state the reason for the submission of the 510(k), e.g., new device, change
in intended use, or design modification  to an existing sheath.
2.  Intended Use
The 510(k) should provide a clear statement of the proposed devices intended  use.  The intended use
should be identically worded in the physicians labeling, the Indications for Use form, and the 510(k)
Summary (if provided).
3.  Physical Description
The physical description of each sheath to be marketed should be provided.  This should
include a labeled diagram, photograph, schematic, etc., which includes all internal, external, assembled,
unassembled, and interchangeable parts.  The physical description should include the dimensional
specifications, such as length, width, height, diameter, weight, etc., and electrical specifications (i.e.,
power  requirements).  Any parts that are disposable should be identified.
If the sheath is to be sold in a set that includes accessories, such as a pump, air compressor, fixation
device, etc., the accessories are considered part of the medical device.  They should be identified and
described  with the same detail as above.  User instructions must include a clear step-by-step
description of the application and removal of the sheath.  Successful reprocessing (cleaning and
disinfection) of endoscopes devices depends on user knowledge of and compliance with strict aseptic
technique.  Ergonomics should be considered and user instructions  tested to foresee and  minimize
potential user errors.
4.  Sheath Materials
An exact identification of all materials used to fabricate the sheath and its accessories should be
provided, with a statement regarding any differences from pre-Amendment devices or the predicate
device.  If the materials are identical to those used in the pre-Amendment or predicate device and are
identically processed and sterilized, then this should be explicitly stated.  This information should
include all direct and indirect patient contacting materials.  The applicant should consider and address
material wear during maximal stress, such as points of articulation, seams or areas held by adhesives.
5.  Shelf Life
The applicant should also consider the expected shelf life of the sheath based on the life expectancy of
the individual materials.  Accelerated aging testing may be performed to identify whether certain
5 materials or components have a shorter life expectancy than others.  Appropriate expiration date
labeling may be required.
Guidance for barrier testing of materials and finished devices is provided in the Appendix.
G.  Special Controls
Endoscopes and accessories (except otoscopes) are Class II devices and subject to special controls.  Special
controls may include special labeling requirements, mandatory performance standards, patient registries and
post market surveillance.  If microbial barrier statements are made, the sponsor would be expected to submit
appropriate supporting laboratory test data.  An example of an appropriate barrier testing protocol is described in
the appendix of this document.
H.  Sterility Information
Complete information regarding the device and accessories that may be sold sterile should be provided.  This
includes sterilization method; sterilization cycle validation method; packaging materials and a description of the
packaging to ensure sterility is maintained; sterility assurance level (SAL); and radiation dose or the maximum
levels of residuals of ethylene oxide, ethylene chlorohydrin, and ethylene glycol which remain on the device.  If
only parts of the device are sold sterile, labeling should clearly identify those parts that are sterile and non-
pyrogenic.
If the device will be labeled as pyrogen free or non-pyrogenic, a description of the method used to make that
determination (LAL or rabbit test) must be provided.
Accessories that are disposable should be labeled as single use.
Guidance on sterility issues are described in the ODE Bluebook Memo K90-1 "510(k) Sterility Review Guidance
(2/12/90)" and in the ODE Guidance Labeling of reusable Medical Devices for Reprocessing in Heath Care
Facilities: FDA Reviewer Guidance dated April 1996.  Copies of these documents may be obtained from DSMA
by calling (800) 638-2041 or (301) 443-6597.
Endoscopes, as used in ENT practice, are considered to be semi-critical devices as they come in contact with
mucous membranes, which may or may not be intact.  The Center for Disease Control (CDC) and Association of
Practitioners in Infection Control (APIC) recommend high-level disinfection as the minimum acceptable level of
reprocessing for semi-critical medical devices such as endoscopes.  High-level disinfection is the
elimination/killing of all vegetative bacteria, virus and fungal spores and some but not all bacterial endospores.
High level disinfection kills some spores, but under most circumstances that is all that is expected.  Only
sterilization ensures an overkill safety margin.
Reprocessing of the endoscope after removal of the used sheath and before application of a new sheath must be
recommended and described in the user's information manual.  If the applicant sufficiently demonstrates
protective barrier properties of the finished device, a cleaning procedure followed by an intermediate disinfection
step will be required.
The applicant should direct the user to follow the cleaning procedure recommended by the endoscope
manufacturer.  This should include instructions to clean the entire endoscope, including the eyepiece and any
hand attachments.  This process serves two purposes; to clean the endoscope in case of a break in aseptic
technique by the user and to mechanically remove any material that may have contacted the insertion tube of the
endoscope during application or removal of the sheath.  In addition to cleaning, an intermediate disinfection step
such as wiping with a 70% isopropyl alcohol soaked gauze pad should be recommended.  This step is added to
reduce the likelihood that any viable organisms remain on the endoscope prior to application of a new sheath.  If
the applicant wishes to propose a different cleaning regimen a full description and justification will be expected..
6 I.  Biocompatibility Testing
Biocompatibility testing data should be provided on any direct or indirect patient-contacting materials that are
not the same as the pre-Amendment or predicate device, or are differently processed or sterilized.  If data are not
provided, a justification should be included explaining why these data are not needed.  Biocompatibility testing
should follow the FDA-modified ISO 10993, Part 1 matrix.  For example, a single use endoscope sheath may
contact breached or compromised surfaces and have a contact duration of less than 24 hours when used in ENT
procedures.  Therefore, cytotoxicity, sensitization and irritation/intracutaneous reactivity testing may be
necessary.  If it can be documented that the patient-contacting finished device materials are identical to materials
used in a legally marketed predicate with similar body contact, biocompatibility data may not be necessary.
However, the applicant must have determined that the finished device materials have been fabricated utilizing the
same chemical formulations, manufacturing processes and sterilization methodology as the predicate device
materials.
Copies of the FDA-modified ISO 10993, Part 1 matrix may be obtained from DSMA by calling (800) 638-2041 or
(301) 443-6597.
An exact identification of all colorants (inks, dyes, markings, radiopaque materials, etc.) used to fabricate the
device or accessory should be provided.  If the colorants are identical to the pre-Amendment or substantially
equivalent device then this should be explicitly stated.  A statement regarding any colorant changes from the
pre-Amendment or substantially equivalent device should be included.  The sponsor should provide
biocompatibility testing data on any colorant changes that have been implemented that will contact the patient
directly or indirectly.  The information should indicate how the markings are processed (etched, bands, etc.) and
whether the color contacts skin, mucosa, etc.
J.   Description of Quality Assurance and Quality Control Programs
The quality assurance and quality control programs should be an integral part of the quality systems
requirements (QSR)as set forth in the Code of Federal Regulations Part 820.  In addition to the statement of
compliance with QSR, the sponsor may want to consider a more detailed description of the quality programs,
such as, finished device, in process or lot testing for device integrity.  Types of testing the sponsor may want to
consider are burst pressure, water leak, helium leak, pressure decay or electrical testing.  Information regarding
conformance to standards may be included.  Test reporting should demonstrate the sensitivity of the method
selected.  The sample
schedule or plan may follow MIL-STD-105E (Sampling Procedure and Tables for Inspection by
Attributes) dated May 10, 1989.
The applicant should provide assurance that QA and QC testing, reporting and corrective action plans are
independent of the manufacturing process.
7 K.  Bench Testing
1.  A comparison of optical qualities of the sheath lens to that of the uncovered endoscope lens should
be included to demonstrate that there is no optical distortion, visual limitations, or impaired light
refraction.
2. Finished device barrier testing may include inoculating the inside of a sheath with a high titer
suspension of challenge organisms, applying the sheath to an endoscope and submerging the
sheath/endoscope combination into a collection fluid, articulating the sheath/endoscope combination a
specified number of times for a specified duration followed by an assay for organisms on the outside of
the sheath and in the collection fluid at the end of the test.  This sample test method is fully described
in Appendix I of this document.  Appendix I provides detail on a suggested test method, elements of
the test, controls and presentation of results.
3.  Alternatively, material qualification testing to measure resistance of the material to ASTM F1671-97a
Standard Test Method for Resistance of Materials Used in Protective Clothing to Penetration by Blood-
Borne Pathogens Using Phi-X174 Bacteriophage Penetration as a Test System.  A copy of this standard
may be obtained by contacting The American Society for Testing and Materials, 100 Barr Harbor Dr.,
West Conshohocken, PA. 19428 (610-832-9500).
The submitter should be aware that the ASTM F1671-97a standard test method is for material qualification only.
Appropriate quality assurance methods for the finished product, sensitive enough to identify defects as small as
5 micron range, may be requested.
Manufacturer's test methods should consider pressure, time, temperature, organism size and real use situations.
The high titer challenge suspension should simulate or exaggerate appropriate body fluid characteristics such as
viscosity, surface tension and pH.
The compliance or noncompliance of the device with any available standards/guidance should be discussed,
including performance, design, and testing provisions.  If available standards/guidance are not used, then an
explanation should be provided.
Other alternative test methods may be used if adequately justified.
L. Clinical Testing
If the application includes statements regarding prevention of disease transmission, clinical testing may be
required.  Data should include microbiological assays of the endoscope surfaces or washings of the sheath
interior after removal from the endoscope.
M.  Labeling
The term "label" includes any identification on the device itself and on the package in which it is stored and
shipped.  If possible, the label on the device should include the device name, corporation name, address, and
phone number.  The package label should include all of the above, as well as sterility status, expiration date,
single use/disposable items, quantity enclosed, size, intended use, etc. and anything specific to the device.
The device label must bear the caution statement as outlined in 21 CFR 801.109(b)(1):  "CAUTION:  Federal law
restricts this device to sale by or on the order of a physician."
8 Device labeling must include all the information required under 21 CFR 801.
1.  The intended use statement should include specific indications, clinical setting, target population,
anatomical sites, etc.
2.  Directions for use should include, but are not necessarily limited to  a) instructions on  statement of
which parts are single use/disposable or reusable, e) functional test procedures for the device prior to
use, and f) clear instructions for aseptic application and removal of the device.  The device is
disposable; however, the endoscope is reprocessed. Therefore, appropriate recommendations for
endoscope cleaning, disinfection or sterilization must be included.
3.  Maintenance and troubleshooting procedures should be maintenance and how often, how and when
to  replace parts, instructions for purchase of replacement parts, and a corporation contact point if
troubleshooting  procedures fail.
4.  Contraindications, precautions, warnings, and adverse effects should be included in the labeling of
the device.
5.  Advertisements or promotional literature that will accompany the device should be provided.
6.  Guidance on labeling issues is described in ODE Bluebook Memo G91-1 "Device Labeling Guidance
(3/18/91)."  A copy may be obtained from the Division of Small Manufacturers Assistance.
V.  Presentation of Data
A.  Tables and Graphs:  Data should be provided in clearly labeled tables.  Any symbols used should
be keyed to a footnote or convenient reference page and described fully.  Graphs may supplement data
tables, but do not replace them.  They also should be clearly labeled.
B.  Published Literature:  Published data or methods that are referenced in the submission should be
provided.  Reprints should be appended to the section in which they are referenced.  All referenced
reports and data should be summarized, including an explanation of how they relate to the current
submission.  Reference citations should be complete (e.g., title, author, volume, page, year).
C.  Protocols and Data Analysis:  Reports of any testing conducted with the device should include the
study protocol (objectives, precise description of materials, experimental methods, controls),
data/observations, statistical methods and analysis, results/conclusions and comments.  Raw data
should not be submitted unless requested.
D.  Reference to Submitted Data:  In support of a 510(k) submission, an applicant may refer to
information  submitted to FDA in the past.  If someone other than the applicant submitted the previous
information, then a letter of authorization is required.  The letter may come through the applicant, or
directly from the original submitter.
9 For more information contact:
Office of  Device Evaluation
Ear, Nose and Throat Branch
Division of Ophthalmic Devices
9200 Corporate Blvd.
Rockville, Md. 20850
(301) 594-2080
Division of Small Manufacturers Assistance may be contacted for assistance at (800) 638-2041 or (301) 443-6597.
10 APPENDIX : SUGGESTED METHODS FOR DETERMINING BARRIER PROPERTIES OF ENT ENDOSCOPE
SHEATHS (AND MATERIALS) TO VIRUS PENETRATION
This guidance was developed by the Office of Science and Technology, Center for Devices and Radiological
Health in collaboration with the Office of Device Evaluation.  This section addresses the rationale, methodology
and required test sensitivity for the ability of ENT endoscope sheaths to act as barriers to transmission of
pathogenic organisms, including viruses.  This specific method is not required, but the principles involved
should be considered in devising a different method.
A medical claim for endoscope sheaths, as being effective barriers requires performance of appropriate
laboratory tests.  Since viruses are the smallest etiological agents and include the human immunodefiency virus
(HIV) and hepatitis B virus (HBV), the challenge particle should be a small virus or virus-size particle. Test
conditions should account for as many parameters as possible that are considered to be important in real-life
conditions, i.e., appropriate choice of challenge particle, solution properties, test pressure and duration.  The
barrier properties of a sheath should be determined in a dynamic test, i.e., movement of the sheath (preferably
caused by articulation of an inserted endoscope) during the test.  Choices of parameters that make the in vitro
test more stringent than expected real-life use are encouraged, with appropriate justification.
The choice of challenge particle(s) has several important aspects.  A biological assay may be preferred in general
because there should be no "background" level of confounding "signal," as would be found with viruses or
virus-like particles labeled with radioactivity or other markers.  Surrogate viruses of appropriate size and shape
may substitute for human pathogens.  Such surrogates may be bacterial viruses (bacteriophages), which are
safer, faster and less expensive to use for testing and which can be readily obtained at sufficient titers to provide
an adequate challenge concentration.  However, in order for the test to be used to demonstrate safety with
regard to human pathogens, the test virus should be as small as the hepatitis A virus (30 nm diameter), one of
the smallest human pathogenic viruses.  For these reasons, the following protocol suggests use of a small
bacterial virus as challenge particle.
Preparation of Test Samples
Test sheaths are placed on an endoscope so they can be articulated during the test.  The sheath and endoscope
should be carefully handled so they are not damaged during the test procedure.  Care should be taken to
replicate a worse case application method as closely as possible.
Two proposed Test Methods, Inside Challenge and ASTM F1671-97a
An appropriate test could consist of submerging a sheath containing a challenge virus plus inserted endoscope
into a submersion chamber filled with a submersion buffer. A sheath can be prepared for testing by inserting a
sterile endoscope into a sheath that is partly filled with challenge suspension.  As the endoscope is inserted into
the sheath, the suspension is distributed over the inner surface of the sheath.  The endoscope with sheath
attached, is submerged into an appropriate submersion buffer, articulated a specified number of times and to a
specified degree.  The submersion buffer is assayed for challenge organisms to determine whether any virus
penetrated the sheath during the process.
Alternatively, material qualification testing to measure resistance of the material to penetration by a surrogate
microbe may be performed.  An appropriate test method is the ASTM F1671-97a Standard Test Method for
Resistance of Materials Used in Protective Clothing to Penetration by Blood-Borne Pathogens Using Phi-X174
Bacteriophage Penetration as a Test System.  A copy of this standard may be obtained by contacting The
American Society for Testing and Materials, 100 Barr Harbor Dr., West Conshohocken, PA. 19428 (610-832-9500).
In a previous version of this guidance an alternative method (called Test B) described placing a sterile
sheath/endoscope combination into a chamber containing a high titer suspension of appropriate test organism.
11 The endoscope/sheath is inserted into the chamber and articulated a specified number of times and to a specified
degree.  Following removal of the endoscope/sheath from the challenge and withdrawal of the endoscope, the
interior surface of the sheath and the surface of the endoscope are rinsed and the wash is assayed for test
organisms.  This method has been eliminated from the guidance as the sensitivity is inadequate and inferior to
both the ASTM F1671-97a and to the inside challenge method described below.
Inside Challenge Method
The elements of the Inside Challenge Method should include:
1.  inoculating the test sheath with challenge virus suspension (in addition to the endoscope) to allow for
submersion of a test portion of the sheath/endoscope (justify the length of the test portion) into the submersion
buffer (sufficient volume of challenge virus suspension is used to cover the inside of the sheath when the
endoscope is in place),
2.  using a submersion vessel that allows for full articulation of the test sheath/endoscope,
3.  utilizing a buffer with appropriate properties (physiological saline, pH approximately 7.0, surface tension less
than 0.05 N/m).  Adding either of two nonionic surfactants, 0.1% Tween 80 or 0.1% triton X-100, will provide the
surface tension desired, while also preventing virus adsorption by nonionic interactions.  This buffer can be
used as the submersion fluid and as the suspending fluid for the challenge virus suspension.
4.  filling the submersion vessel with  100ml of buffer that has appropriate properties (physiological saline, pH
approximately 7.0, surface tension not more than .05 N/m; see #3 above). The test may be performed at room
temperature [68-72F],
5.  inoculating the test sheath with a challenge virus suspension at sufficient titer, even at the end of the test (at
least 108 plaque forming units/mL (pfu/mL), of a small, approximately spherical virus, e.g., the bacteriophage
PhiX174),
6.  articulating the endoscope to simulate the movement of actual use (maximum deflection in both directions and
holding for a predetermined period of time which would simulate a stressed real use scenario.  The period of time
in the unarticulated state should account for not more than 50% of the test time. The test time should be a
minimum of 60 minutes duration.  Increased virus titer to decrease test duration may be considered,
7.  carefully removing the endoscope/sheath combination and sampling the submersion buffer to collect any
virus that penetrated through the sheath (Care is required to prevent contamination of the submersion buffer),
8.  assaying the submersion buffer to determine whether any challenge virus has penetrated the sheath, and
9.  calculating the equivalent volume of challenge virus penetration needed to account for amount of virus found
in submersion buffer.
Controls
It is well known that some viruses can be removed from suspension by certain materials through different forms
of adsorption or that they can be rendered biologically undetectable by chemical inactivation.  Thus controls are
needed to assure that the virus penetration test will yield meaningful data. Positive control experiments are
needed to assure that the overall test is functioning properly.  Sheaths with intentional pinholes may be used.
It must be ascertained whether the challenge virus is stable during the test.  Collection of these data can be a
part of each sheath test.  The titer of the challenge virus suspension in each sheath in the Inside Challenge
Method, (or in the ASTM Test ) at the end of the test is compared to the original titer.  This determines if and
12 how much the challenge virus titer changes during the test because of interaction with the test sample and test
apparatus.
It must also be ascertained whether any virus that penetrates the sheath remains detectable in this test
procedure.  This can be done in the Inside Challenge Method by "spiking" the submersion buffer in the
submersion vessel with a small volume (10 microliters [L]) of virus (at 106 pfu/mL) before a mock test and
assaying the titer of the submersion buffer at the end of the mock test.  This determines if and how much of the
penetrated virus can be recovered.  The ASTM test includes a similar control experiment.
If either (or both) of the above controls indicates loss of virus titer, the starting challenge titer must be increased
to compensate for the loss in order to maintain the overall sensitivity of the test.
Sampling Procedure
A complete data set should include results from at least 21 sheaths (7 from each of 3 lots), in order to provide
assurance that overall quality of each of three lots is satisfactory.
Detection Limit
Detection limit expressed as volume of challenge virus suspension that penetrated the barrier is probably the
most useful measure of test sensitivity.  For example, in a real-life risk assessment the volume of transmitted
virus-containing fluid can be translated into infectious units when the titer of a pathogenic virus (in real life) is
known.
The test procedure should be able to detect 1x10-6 mL penetration of the challenge virus suspension.  This can
be done in the Inside Challenge Method by using a challenge titer of 108 pfu/mL, a submersion buffer volume of
100 mL and assaying 1 mL in triplicate from the submersion buffer (assuming no loss of virus titer in the
challenge buffer nor in the recovery of penetrated virus):  the assay detection limit of 1 pfu/mL is equivalent to
penetration by 100 pfu (1 pfu/mL x 100 mL) or 1x10-6 mL (100 pfu divided by 108 pfu/mL).
Presentation of Results
A table of the results for all the test sheaths should be presented that includes: the challenge virus titer, the titer
or amount of virus that penetrated the sheath, any correction factor for loss of virus  (determined in the
controls), and the calculated challenge volume that penetrated (for the sheaths that allowed virus transmission).
The volume of challenge virus suspension needed to account for the virus penetration into the submersion
buffer can be calculated for each sheath by the method presented in the previous section. If some loss of virus
titer occurs either inside the sheath or inside the submersion vessel, the calculation should include the
appropriate correction for such loss.  For sheaths that apparently did not allow virus transmission, the detection
limit of that particular test should be given, e.g., as <1x10-6 mL.
Report Forms
Please report the test results for either the Inside Challenge Method or the ASTM method in the following
manner:
Test results for virus penetration of sheath samples should be presented in tabular form, where the data for each
sheath are individually reported.  The example given is for the Inside Challenge Method. Necessary items for
each test sample are:
i) date test was performed,
ii)titer of challenge virus at the end of the test,
iii) calculated detection limit based on the challenge virus titer, the submersion buffer volume,
and the volume assayed,
13 iv) pfu's detected in the submersion buffer,
vi) calculated titer of penetrated virus in submersion buffer, and volume of challenge virus suspension
needed to account for the amount of virus detected in the rinse buffer.
Information accompanying the table should include:
a) the challenge virus,
b) the challenge and submersion fluids (e.g., buffer and surfactant),
c) how the titer of the challenge virus suspension was determined (dilution, volume assayed, and
number of replicate assays),
d) the submersion depth of the sheath (if variable from one test sample to another, it should be included
in  the table for each sheath),
e) the submersion buffer volume (if variable from one test sample to another, it should be included in
the table for each sheath),
f) any evidence of an equipment or procedural malfunction during any particular test.
Example Table I. Results for virus penetration through sheath samples of Brand X, Lot #34068.
-------------------------------------------------------------
   i       ii  iii iv   v      vi
 Date  Titer,      Detect     Virus in     Titer,          Volume of   
 challenge  limit     submersion   submersion       penetrated virus
 virus buffer    buffer
              (pfu/mL)   (mL)     (pfu)         (pfu/mL)      (mL)
--------------------------------------------------------------
1  10/28  1.15x108      0.87x10-6            29,28,33                 3.0x101                     2.6x10-5
                                             (+0.2)
2  10/28  1.23x108       0.81x10-6  0,0,0                <1               <0.8x10-6
.
.
.               
-------------------------------------------------------------------------------------------------------------------------------------------
Positive Control
Reporting the results of the positive control experiment should be done using the same reporting format as with
virus penetration of test samples.
Control to test challenge virus stability
Results from the test of challenge virus stability should be presented in tabular form, where the data for each
sheath are individually reported.  Necessary items for each test sample are:
     i)  date test was performed,
     ii)  titer of challenge virus placed in the sheath  at the beginning of the test,
     iii) titer of challenge virus at the end of the test, and
     iv) calculated ratio of final to beginning titers.
14 Example Table II.  Results of test for stability of challenge virus in samples of Brand X, Lot #34068.
---------------------------------------------------------------------------------------------------------------------------
Sample Date Beginning Final Ratio
titer titer final/
(pfu/mL) (pfu/mL) begin
---------------------------------------------------------------------------------------------------------------------------
1 10/28/96 1.15x108 1.14x108 0.99
( 0.11) ( 0.13)
.
.
.
-------------------------------------------------------------------------------------------------------------------------------------------
Control to test detectability of virus, which penetrates sheath ("spiking experiment")
Results from tests to determine the delectability of penetrated virus should be in tabular form, where the data for
each sheath are individually reported.  Necessary items for each test sample are:
     i)  date test was performed,
     ii)  number of virus (pfu) in 10 microliters at the beginning of the test,
     iii) total number of virus (pfu) in submersion buffer at the end of the test, and
     iv) calculated ratio of final to beginning numbers.
Example Table III. Results of test for recoverability of penetrated virus in sheath samples of Brand X, Lot #34068.
---------------------------------------------------------------------------------------------------------------------------
Sample    Date   Beginning Final Ratio,
                                                         virus number    virus number   final/
                                                         (pfu/10L)        (pfu/100mL) begin
--------------------------------------------------------------------------------------------------------------------------
1            10/28/96     1.2x104 1.1x104 0.92
       ( 0.1)    0.1)
.
.
.
--------------------------------------------------------------------------------------------------------------------------
15