SIST EN ISO 10993-3:2026

SLOVENSKI STANDARD
01-november-2026
Nadomešča:
SIST EN ISO 10993-3:2015
Biološko ovrednotenje medicinskih pripomočkov - 3. del: Ovrednotenje genske
toksičnosti, kancerogenosti, toksičnosti za razmnoževanje in toksičnosti za razvoj
(ISO 10993-3:2026)
Biological evaluation of medical devices - Part 3: Evaluation of genotoxicity,
carcinogenicity, reproductive toxicity and developmental toxicity (ISO 10993-3:2026)
Biologische Beurteilung von Medizinprodukten - Teil 3: Bewertung der Genotoxizität,
Karzinogenität, Reproduktionstoxizität und Entwicklungstoxizität (ISO 10993-3:2026)
Évaluation biologique des dispositifs médicaux - Partie 3: Évaluation de la génotoxicité,
de la cancérogénicité, de la toxicité sur la reproduction et de la toxicité sur le
développement (ISO 10993-3:2026)
Ta slovenski standard je istoveten z: EN ISO 10993-3:2026
ICS:
11.100.20 Biološko ovrednotenje Biological evaluation of
medicinskih pripomočkov medical devices
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.

EN ISO 10993-3
EUROPEAN STANDARD
NORME EUROPÉENNE
September 2026
EUROPÄISCHE NORM
ICS 11.100.20 Supersedes EN ISO 10993-3:2014
English Version
Biological evaluation of medical devices - Part 3:
Evaluation of genotoxicity, carcinogenicity, reproductive
toxicity and developmental toxicity (ISO 10993-3:2026)
Évaluation biologique des dispositifs médicaux - Partie Biologische Beurteilung von Medizinprodukten - Teil 3:
3: Évaluation de la génotoxicité, de la cancérogénicité, Bewertung der Genotoxizität, Karzinogenität,
de la toxicité sur la reproduction et de la toxicité sur le Reproduktionstoxizität und Entwicklungstoxizität (ISO
développement (ISO 10993-3:2026) 10993-3:2026)
This European Standard was approved by CEN on 12 July 2026.

CEN members are bound to comply with the CEN/CENELEC Internal Regulations which stipulate the conditions for giving this
European Standard the status of a national standard without any alteration. Up-to-date lists and bibliographical references
concerning such national standards may be obtained on application to the CEN-CENELEC Management Centre or to any CEN
member.
This European Standard exists in three official versions (English, French, German). A version in any other language made by
translation under the responsibility of a CEN member into its own language and notified to the CEN-CENELEC Management
Centre has the same status as the official versions.

CEN members are the national standards bodies of Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech Republic, Denmark, Estonia,
Finland, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway,
Poland, Portugal, Republic of North Macedonia, Romania, Serbia, Slovakia, Slovenia, Spain, Sweden, Switzerland, Türkiye and
United Kingdom.
EUROPEAN COMMITTEE FOR STANDARDIZATION
COMITÉ EUROPÉEN DE NORMALISATION

EUROPÄISCHES KOMITEE FÜR NORMUNG

CEN-CENELEC Management Centre: Rue de la Science 23, B-1040 Brussels
© 2026 CEN All rights of exploitation in any form and by any means reserved Ref. No. EN ISO 10993-3:2026 E
worldwide for CEN national Members.

Contents Page
European foreword . 3
Annex ZA (informative) Relationship between this European standard and the General
Safety and Performance Requirements of Regulation (EU) 2017/745 aimed to be
covered. 4
European foreword
This document (EN ISO 10993-3:2026) has been prepared by Technical Committee ISO/TC 194
"Biological and clinical evaluation of medical devices" in collaboration with Technical Committee
CEN/TC 206 “Biological and clinical evaluation of medical devices” the secretariat of which is held by
DIN.
This European Standard shall be given the status of a national standard, either by publication of an
identical text or by endorsement, at the latest by March 2027, and conflicting national standards shall
be withdrawn at the latest by March 2027.
Attention is drawn to the possibility that some of the elements of this document may be the subject of
patent rights. CEN shall not be held responsible for identifying any or all such patent rights.
This document supersedes EN ISO 10993-3:2014.
This document has been prepared under a standardization request addressed to CEN by the European
Commission. The Standing Committee of the EFTA States subsequently approves these requests for its
Member States.
For the relationship with EU Legislation, see informative Annex ZA, which is an integral part of this
document.
Any feedback and questions on this document should be directed to the users’ national standards
body/national committee. A complete listing of these bodies can be found on the CEN website.
According to the CEN-CENELEC Internal Regulations, the national standards organizations of the
following countries are bound to implement this European Standard: Austria, Belgium, Bulgaria,
Croatia, Cyprus, Czech Republic, Denmark, Estonia, Finland, France, Germany, Greece, Hungary, Iceland,
Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Republic of
North Macedonia, Romania, Serbia, Slovakia, Slovenia, Spain, Sweden, Switzerland, Türkiye and the
United Kingdom.
Endorsement notice
The text of ISO 10993-3:2026 has been approved by CEN as EN ISO 10993-3:2026 without any
modification.
Annex ZA
(informative)
Relationship between this European standard and the General Safety and
Performance Requirements of Regulation (EU) 2017/745 aimed to be
covered
This European standard has been prepared under M/575 to provide one voluntary means of
conforming to the General Safety and Performance Requirements of Regulation (EU) 2017/745 of 5
April 2017 concerning medical devices [OJ L 117] and to system or process requirements including
those relating to quality management systems, risk management, post-market surveillance systems,
clinical investigations, clinical evaluation or post-market clinical follow-up.
Once this standard is cited in the Official Journal of the European Union under that Regulation,
compliance with the normative clauses of this standard given in Table ZA.1 and application of the
edition of the normatively referenced standards as given in Table ZA.2 confers, within the limits of the
scope of this standard, a presumption of conformity with the corresponding General Safety and
Performance Requirements of that Regulation, and associated EFTA Regulations.
Where a definition in this standard differs from a definition of the same term set out in Regulation (EU)
2017/745, the differences shall be indicated in this Annex Z. For the purpose of using this standard in
support of the requirements set out in Regulation (EU) 2017/745, the definitions set out in this
Regulation prevail.
Where the European standard is an adoption of an International Standard, the scope of this document
can differ from the scope of the European Regulation that it supports. As the scope of the applicable
regulatory requirements differ from nation to nation and region to region the standard can only
support European regulatory requirements to the extent of the scope of the European Regulation for
medical devices ((EU) 2017/745).
NOTE 1 Where a reference from a clause of this standard to the risk management process is made, the risk
management process needs to be in compliance with Regulation (EU) 2017/745. This means that risks have to be
‘reduced as far as possible’, ‘reduced to the lowest possible level’, ‘reduced as far as possible and appropriate’,
‘removed or reduced as far as possible’, ‘eliminated or reduced as far as possible’, ’removed or minimized as far as
possible’, or ‘minimized’, according to the wording of the corresponding General Safety and Performance
Requirement.
NOTE 2 The manufacturer’s policy for determining acceptable risk must be in compliance with General Safety
and Performance Requirements 1, 2, 3, 4, 5, 8, 9, 10, 11, 14, 16, 17, 18, 19, 20, 21 and 22 of the Regulation.
NOTE 3 When a General Safety and Performance Requirement does not appear in Table ZA.1, it means that it is
not addressed by this European Standard.
Table ZA.1— Correspondence between this European Standard and
Annex I of Regulation (EU) 2017/745 [OJ L 117] and to system or process requirements
including those relating to quality management systems, risk management, post-market
surveillance systems, clinical investigations, clinical evaluation, or post-market clinical follow-
up
General Safety and
Performance Requirements Clause(s)/sub-clause(s) of
Remarks/Notes
of Regulation this EN
(EU) 2017/745
This document covers the
assessment of genotoxicity,
carcinogenicity, endocrine
10.1 (a) and (b) 4, 5 and 6
disruptors, and reproductive
toxicity and developmental toxicity
only.
This document covers the
assessment of genotoxicity,
carcinogenicity, endocrine
10.2 4, 5 and 6
disruptors, and reproductive
toxicity and developmental toxicity
only.
GSPR 10.4.1 is only partly covered
by this document, since this
document does not provide
requirements for design and
manufacture, nor does it address
risks associated with particles,
except nanoparticles, from medical
10.4.1 (first paragraph) 4, 5 and 6
devices.
This aspect is addressed
concerning the assessment of
genotoxicity, carcinogenicity,
endocrine disruptors and
reproductive toxicity and
developmental toxicity only.
This aspect is addressed
concerning the assessment of
genotoxicity, carcinogenicity,
endocrine disruptors and
reproductive toxicity and
10.4.2 (a) 4, 5, 6 and 7 developmental toxicity. This
ensures that patients or users are
safeguarded against exposure to
residues, based on the extent and
nature of their interaction with the
medical device.
Table ZA.2— Normative references from clause 2 of this document and their corresponding
European publications
Column 1 Column 2 Column 3 Column 4
Reference in International Title Corresponding European
Clause 2 Standard Edition Standard Edition
ISO 10993- ISO 10993-1:2025 Biological evaluation of medical EN ISO 10993-1:2025
1:2025 devices — Part 1: Evaluation and
testing within a risk management
process
ISO 10993-2 ISO 10993-2:2022 Biological evaluation of medical EN ISO 10993-2:2022
devices — Part 2: Animal welfare
requirements
ISO 10993- ISO 10993- Biological evaluation of medical EN ISO 10993-12:2021
12:2021 12:2021 devices — Part 12: Sample
preparation and reference
materials
ISO 10993-17 ISO 10993- Biological evaluation of medical EN ISO 10993-17:2023
17:2023 devices — Part 17: Establishment
of allowable limits for leachable
substances
ISO 14971:201 ISO 14971:2019 Medical devices - Application of EN ISO 14971:2019
9 risk management to medical EN ISO
devices 14971:2019/A11:2021
OECD 471:202 OECD 471:2020 OECD Guideline for the Testing of For applicable standard
0 Chemicals — Bacterial Reverse edition see Column 2
Mutation Test
OECD 473:201 OECD 473:2016 OECD Guideline for the Testing of For applicable standard
6 Chemicals — In Vitro Mammalian edition see Column 2
Chromosomal Aberration Test
OECD 487:201 OECD 487:2016 OECD Guideline for the Testing of For applicable standard
6 Chemicals — In Vitro Mammalian edition see Column 2
Cell Micronucleus Test
OECD OECD 490:2016 OECD Guideline for the Testing of For applicable standard
490:2016 Chemicals — In Vitro Mammalian edition see Column 2
Cell Gene Mutation Tests Using the
Thymidine Kinase Gene
The documents listed in the Column 1 of table ZA.2, in whole or in part, are normatively referenced in
this document, i.e. are indispensable for its application. The achievement of the presumption of
conformity is subject to the application of the edition of Standards as listed in Column 4 or, if no
European Standard Edition exists, the International Standard Edition given in Column 2 of table ZA.2.
Table ZA.3— Correspondence between this European Standard and Annex II of Regulation (EU)
2017/745 [OJ L 117]
Requirements of Annex II of Clause(s)/sub-clause(s) of
Remarks/Notes
Regulation (EU) 2017/745 this EN
This document covers the
assessment of genotoxicity,
6.2 (c) fourth indent 4, 5 and 6 carcinogenicity, endocrine
disruptors, and reproductive and
developmental toxicity only
WARNING 1 Presumption of conformity stays valid only as long as a reference to this European
standard is maintained in the list published in the Official Journal of the European Union. Users of this
standard should consult frequently the latest list published in the Official Journal of the European
Union.
WARNING 2 Other Union legislation may be applicable to the product(s) falling within the scope of
this standard.
International
Standard
ISO 10993-3
Fourth edition
Biological evaluation of medical
2026-08
devices —
Part 3:
Evaluation of genotoxicity,
carcinogenicity, reproductive
toxicity and developmental toxicity
Évaluation biologique des dispositifs médicaux —
Partie 3: Évaluation de la génotoxicité, de la cancérogénicité,
de la toxicité sur la reproduction et de la toxicité sur le
développement
Reference number
ISO 10993-3:2026(en) © ISO 2026

ISO 10993-3:2026(en)
© ISO 2026
All rights reserved. Unless otherwise specified, or required in the context of its implementation, no part of this publication may
be reproduced or utilized otherwise in any form or by any means, electronic or mechanical, including photocopying, or posting on
the internet or an intranet, without prior written permission. Permission can be requested from either ISO at the address below
or ISO’s member body in the country of the requester.
ISO copyright office
CP 401 • Ch. de Blandonnet 8
CH-1214 Vernier, Geneva
Phone: +41 22 749 01 11
Email: copyright@iso.org
Website: www.iso.org
Published in Switzerland
ii
ISO 10993-3:2026(en)
Contents Page
Foreword .iv
Introduction .v
1 Scope . 1
2 Normative references . 1
3 Terms and definitions . 2
4 Assessment strategies . . 2
4.1 General .2
4.2 Other considerations .3
5 Genotoxicity assessment . 3
5.1 General .3
5.2 Testing strategy .4
5.2.1 General .4
5.2.2 In vitro genotoxic test battery .4
5.2.3 In vivo genotoxicity testing .5
5.2.4 Follow-up evaluation .5
5.3 Test sample preparation .5
6 Carcinogenicity assessments . 6
6.1 General .6
6.2 Evaluation strategy .6
6.2.1 General .6
6.2.2 Genotoxic carcinogens .6
6.2.3 Non-genotoxic carcinogens .7
7 Evaluation of reproductive toxicity and developmental toxicity . 7
7.1 General .7
7.2 Evaluation methods .8
8 Test report . 9
Annex A (informative) Test sample preparation method for genotoxicity testing .10
Annex B (normative) Tests for genotoxicity .16
Annex C (informative) Carcinogenicity tests in animals .27
Annex D (informative) Genotoxicity assessment of nanomaterials .30
Annex E (informative) Recipe for S9 mix .31
Annex F (informative) Endocrine disruptors .32
Annex G (informative) Evaluation of reproductive toxicity and developmental toxicity .33
Bibliography .37

iii
ISO 10993-3:2026(en)
Foreword
ISO (the International Organization for Standardization) is a worldwide federation of national standards
bodies (ISO member bodies). The work of preparing International Standards is normally carried out through
ISO technical committees. Each member body interested in a subject for which a technical committee
has been established has the right to be represented on that committee. International organizations,
governmental and non-governmental, in liaison with ISO, also take part in the work. ISO collaborates closely
with the International Electrotechnical Commission (IEC) on all matters of electrotechnical standardization.
The procedures used to develop this document and those intended for its further maintenance are described
in the ISO/IEC Directives, Part 1. In particular, the different approval criteria needed for the different types
of ISO document should be noted. This document was drafted in accordance with the editorial rules of the
ISO/IEC Directives, Part 2 (see www.iso.org/directives).
ISO draws attention to the possibility that the implementation of this document may involve the use of (a)
patent(s). ISO takes no position concerning the evidence, validity or applicability of any claimed patent
rights in respect thereof. As of the date of publication of this document, ISO had not received notice of (a)
patent(s) which may be required to implement this document. However, implementers are cautioned that
this may not represent the latest information, which may be obtained from the patent database available at
www.iso.org/patents. ISO shall not be held responsible for identifying any or all such patent rights.
Any trade name used in this document is information given for the convenience of users and does not
constitute an endorsement.
For an explanation of the voluntary nature of standards, the meaning of ISO specific terms and expressions
related to conformity assessment, as well as information about ISO's adherence to the World Trade
Organization (WTO) principles in the Technical Barriers to Trade (TBT), see www.iso.org/iso/foreword.html.
This document was prepared by Technical Committee ISO/TC 194, Biological and clinical evaluation of medical
devices, in collaboration with the European Committee for Standardization (CEN) Technical Committee CEN/
TC 206, Biological and clinical evaluation of medical devices, in accordance with the Agreement on technical
cooperation between ISO and CEN (Vienna Agreement).
This fourth edition of ISO 10993-3 cancels and replaces ISO 10993-3:2014 and ISO/TR 10993-33:2015.
The main changes are as follows:
— this document has been entirely revised to emphasize evaluation of genotoxicity, carcinogenicity,
reproductive toxicity and developmental toxicity instead of testing;
— chemical characterization and toxicological risk assessment have been added as an approach to address
genotoxicity, carcinogenicity, reproductive toxicity and developmental toxicity;
— the follow-up evaluation has been changed when an in vitro genotoxicity test is positive and the flowchart
for follow-up evaluation has been removed;
— reproductive toxicity and developmental toxicity have been described separately;
— the annex on cellular transformation has been deleted;
— ISO/TR 10993-33 has been merged into this document as Annex B on genotoxicity test methods.
A list of all parts in the ISO 10993 series can be found on the ISO website.
Any feedback or questions on this document should be directed to the user’s national standards body. A
complete listing of these bodies can be found at www.iso.org/members.html.

iv
ISO 10993-3:2026(en)
Introduction
The basis for biological evaluation of medical devices is often empirical and driven by relevant concerns
for human safety. The risk of serious and irreversible effects, such as cancer or second-generation
abnormalities, is of particular public concern. It is inherent in the provision of safe medical devices that such
risks be minimized to the greatest extent feasible. The assessment of mutagenic, carcinogenic, reproductive
toxicity and developmental toxicity hazards is an essential component of the control of these risks. Not all
test methods for the assessment of genotoxicity, carcinogenicity, reproductive toxicity and developmental
toxicity are equally well developed, nor is their validity well established for the testing of medical devices.
Significant issues with test sample size and preparation, scientific understanding of disease processes and
test validation can be cited as limitations of available methods. Since the previous edition of this document,
many genotoxicity test methods have been updated with revised OECD guidelines. However, these generally
provide clearer recommendations but little alteration in test methods. Scientifically sound alternatives to
the proposed testing can be acceptable insofar as they address relevant matters of safety assessment of the
medical device.
In the selection of tests needed to evaluate a particular medical device, a careful assessment of expected
human uses and potential interactions with biological systems is important, particularly in such areas as
reproductive toxicity and developmental toxicity.
This document presents test methods and evaluation strategies for the identification of specific biological
harms. Testing is not always necessary or appropriate in evaluating biological risks associated with exposure
to medical device materials but, where it is appropriate, it is important that maximum test sensitivity is
achieved.
In view of the multitude of possible outcomes and the importance of factors such as extent of exposure,
species differences and mechanical or physical considerations, risk assessment is typically performed on a
[7]
case-by-case basis. Suggestions for risk consideration and integration of ISO 10993-17 and ISO 10993-18
are provided.
v
International Standard ISO 10993-3:2026(en)
Biological evaluation of medical devices —
Part 3:
Evaluation of genotoxicity, carcinogenicity, reproductive
toxicity and developmental toxicity
1 Scope
This document specifies strategies for risk estimation and evaluation of biological harms with respect to:
— genotoxicity;
— carcinogenicity;
— reproductive toxicity; and
— developmental toxicity.
This document is applicable when the need to evaluate a medical device for potential genotoxicity,
carcinogenicity, reproductive toxicity and developmental toxicity has been established.
This document is not applicable to active pharmaceutical ingredients of device-drug combination products
or biological components of device-biologic combination products which are covered by regulations.
NOTE Guidance on selecting relevant biological effects for medical devices is covered in ISO 10993-1.
2 Normative references
The following documents are referred to in the text in such a way that some or all of their content constitutes
requirements of this document. For dated references, only the edition cited applies. For undated references,
the latest edition of the referenced document (including any amendments) applies.
ISO 10993-1:2025, Biological evaluation of medical devices — Part 1: Requirements and general principles for
the evaluation of biological safety within a risk management process
ISO 10993-2, Biological evaluation of medical devices — Part 2: Animal welfare requirements
ISO 10993-12:2021, Biological evaluation of medical devices — Part 12: Sample preparation and reference
materials
ISO 10993-17, Biological evaluation of medical devices — Part 17: Toxicological risk assessment of medical
device constituents
ISO 14971:2019, Medical devices — Application of risk management to medical devices
OECD 471:2020, OECD Guideline for the Testing of Chemicals — Bacterial Reverse Mutation Test
OECD 473:2016, OECD Guideline for the Testing of Chemicals — In vitro Mammalian Chromosome Aberration
Test
OECD 487:2016, OECD Guideline for the Testing of Chemicals — In vitro Mammalian Cell Micronucleus Test
OECD 490:2016, OECD Guideline for the Testing of Chemicals — In vitro Mammalian Cell Gene Mutation Tests
Using the Thymidine Kinase Gene

ISO 10993-3:2026(en)
3 Terms and definitions
For the purposes of this document, the terms and definitions given in ISO 10993-1, ISO 10993-12 and the
following apply.
ISO and IEC maintain terminology databases for use in standardization at the following addresses:
— ISO Online browsing platform: available at https:// www .iso .org/ obp
— IEC Electropedia: available at https:// www .electropedia .org/
3.1
carcinogenicity test
test to determine cancer outcomes after long term or life-span exposure of animals
Note 1 to entry: Outcomes are typically based on tumorigenic potential and other factors.
3.2
developmental toxicity evaluation
methods to determine adverse effects on the developing organism during prenatal development or
postnatally to the time of sexual maturation
3.3
genotoxicity test
test using cells, bacteria, yeast, fungi, animals or other biological systems to determine whether gene
mutations, chromosomal alterations or other genetic changes are caused by the test sample
3.4
reproductive toxicity evaluation
methods to evaluate the potential effects of test samples on male or female reproductive function during
any stage of development or fertility
3.5
sham-incubated vehicle
vehicle control exposed to the same extraction time and temperature as the test sample extract
3.6
threshold of toxicological concern
TTC
level of exposure for constituents, below which there would be no appreciable risk to human health
Note 1 to entry: The constituent is defined as a chemical or compound present in or on a finished medical device
or material(s) of construction. Constituents can be intentionally present (e.g. an additive such as an antioxidant) or
unintentionally present (e.g. an impurity).
[11]
[SOURCE: ISO/TS 21726:2019, 3.5 , modified — Note 1 to entry has been added.]
4 Assessment strategies
4.1 General
ISO 10993-1 indicates circumstances where the potential for genotoxicity, carcinogenicity, reproductive
toxicity and developmental toxicity are relevant biological harms for consideration in an overall biological
safety evaluation. Evaluation of the biological harms shall consider the following factors:
— whether assessment is necessary based on patient population (e.g. life expectancy, sensitive population)
and the anticipated impact of test results on risk management judgements,
— whether assessment is necessary based on the worst-case quantity, type of contact and duration of
exposure,
ISO 10993-3:2026(en)
NOTE Quantity can be influenced by device size and number used per patient.
— an analysis of the chemical constituents of the device material(s), including manufacturing process
residues and degradation products or metabolites, to identify causes of concern. The identification can
be based on chemical-specific toxicity data if available; otherwise, structure-activity relationships or
previous demonstration of relevant toxicity in the chemical class can be used,
— existing information relevant to the genotoxicity, carcinogenicity, reproductive toxicity and
developmental toxicity evaluation of the medical device, and
— previous use of equivalent (see ISO 10993-1:2025, 6.7) materials and processing in relevant applications.
The decision to waive in vitro genotoxicity, carcinogenicity, reproductive toxicity and developmental toxicity
testing shall be justified. The decision to conduct in vivo testing shall be justified.
[7]
If exposure to constituents identified in accordance with ISO 10993-18 is determined to be within
acceptable levels based on a toxicological risk assessment conducted in accordance with ISO 10993-17, no
additional testing needs to be performed to address the relevant risks.
4.2 Other considerations
Toxicity evaluation can be warranted for additional states of the device, such as wear debris generated
from the device, absorbable materials, or materials that cure in situ (e.g. cements, adhesives, pre-polymer
mixtures) unless toxicological risk assessment based on material information determines no cause for
concern. For guidance on test sample preparation for in situ curing devices, see ISO 10993-12.
NOTE Material information can include chemical composition, degradation products, debris size and morphology,
pre-polymerization, post-polymerization products, etc.
5 Genotoxicity assessment
5.1 General
Assessment of genotoxic risk should be made:
a) by conducting toxicological risk assessment based on chemical characterization according to
[7]
ISO 10993-17 and ISO 10993-18 ; or
b) by performing genotoxicity testing in the standard test battery according to 5.2.2.
In certain cases (e.g. a material which has not been used in a legally marketed device for its intended use),
both toxicological risk assessment of medical device constituents based on chemical characterization and
genotoxicity testing of a medical device can be appropriate.
NOTE 1 Genotoxicity testing can also be appropriate when chemical characterization data is insufficient for a
toxicological risk assessment.
If the device is made with and releases nanomaterials, additional evaluation shall be considered.
[9]
NOTE 2 See ISO/TR 10993-22 and Annex D for more information.
If chemical characterization of the device identifies genotoxicants which are not easily detected using non-
[7]
targeted analytical approaches described in ISO 10993-18 (e.g. glutaraldehyde, formaldehyde), targeted
analysis of genotoxic residuals shall be conducted. Where compositional profiling or analytical chemistry
reveals the presence of chemical constituents with inadequate genotoxicity data and where quantitative
structure activity relationship (QSAR) information is inconclusive (see ISO 10993-17), testing of individual
chemicals or device components containing the chemical should be considered. In either case, the test
sample shall be justified and documented.

ISO 10993-3:2026(en)
5.2 Testing strategy
5.2.1 General
No single test is capable of detecting all types of genotoxic agents. Therefore, the usual approach is to
conduct a battery of in vitro and, under certain circumstances, also in vivo tests.
Genotoxicity tests are designed to detect the two major classes of genetic damage:
— gene mutations (DNA single base pair changes, small deletions or insertions), and
— chromosomal damage [structural aberrations such as translocations, small or large deletions and
insertions, and gain or loss of whole chromosomes (aneuploidy) or parts of chromosomes].
NOTE Bacterial reverse mutation assays have been shown to detect relevant small-scale mutational changes
produced by the majority of genotoxic carcinogens detected by rodent bioassays.
5.2.2 In vitro genotoxic test battery
When genotoxicity testing is performed, it shall be conducted in accordance with Annex B and the test
battery shall include:
a) a test for gene mutations in bacteria (see OECD 471), modified for use with medical devices, see
Clause B.2, and
b) one of the following three validated in vitro mammalian cell tests:
1) an in vitro test with cytogenetic evaluation of chromosomal damage with mammalian cells (see
OECD 473), modified for medical devices, in accordance with Clause B.3 and B.4.2, or
+/- +/-
2) an in vitro mouse lymphoma tk assay using L5178Y (tk ) cells (see OECD 490), modified for
medical devices, including detection of small (slow growing) and large colonies, see Clause B.3 and
B.4.1, or
3) an in vitro mammalian cell micronucleus test for chromosomal damage and aneugenicity (see OECD
487), modified for medical devices, see Clause B.3 and B.4.3.
NOTE 1 If the medical device is known to contain nanomaterials testing according to OECD 471 is not recommended,
see Annex D.
All tests (bacterial and in vitro mammalian tests) shall be performed in the presence and absence of
metabolic activation. Metabolic activation is achieved by addition of a co-factor-supplemented post-
mitochondrial fraction (S9).
NOTE 2 An example for a S9 recipe is given in Annex E.
In vitro mammalian tests shall include both a short-term exposure with and without S9 and a long-term
exposure without S9.
NOTE 3 See B.3.2.3 for descriptions of short-and long-term exposure.
+/-
Results from both chromosomal aberration assays and the in vitro mouse lymphoma tk assay (MLA) have
a relatively high level of congruence for compounds that are regarded as genotoxic but yield negative results
in the bacterial reverse mutation assay. The MLA detects the broadest set of genetic damage, including both
small-scale and large-scale genetic damages. The micronucleus assay detects chromosomal damage in the
form of whole chromosome loss or chromosomal fragments and uniquely detects aneuploidy. The three tests
are equally acceptable as an in vitro mammalian genotoxicity test (see OECD 473, OECD 487 and OECD 490).
The hypoxanthine guanine phosphoribosyl transferase gene mutation assay (HGPRT/HPRT) method (see
[84]
OECD 476 ) is not included in the recommended test battery for device testing. This test can be appropriate
for medical devices containing nanomaterials (see Annex D).

ISO 10993-3:2026(en)
5.2.3 In vivo genotoxicity testing
In vivo genotoxicity tests are not part of the recommended genotoxicity test battery because they are
relatively insensitive and not likely to detect genotoxicity at the concentrations of substances generally
[73][74]
found in medical devices or in medical device extracts . These concentrations can be insufficient to
meet the limit dose required by the test methods. However, under certain circumstances, an in vivo test can
be useful.
These circumstances can include:
a) assessment of nanomaterial or other particulates suspected of sequestration in particular organs or
[9]
tissues (see Annex D, Reference [30] and ISO/TR 10993-22 ); or
[6]
b) when additional relevant factors [e.g. toxicokinetics (see ISO 10993-16 ), organ-specific toxicity] need
to be considered.
In vivo tests are valid only with data demonstrating the test sample reached the target cells, organs, or
tissues. Mutational analysis in target organs of rats or mice can be determined using one of several rodent
[27]
transgenic mutation assays (see OECD 488 ); chromosomal damage can be assessed in the erythrocyte
[24] [28]
micronucleus assay (see OECD 474 ), the in vivo comet assay (see OECD 489 ), or the in vivo chromosomal
[25]
aberration assay (see OECD 475 ). See Clause B.5 for additional considerations for in vivo assays.
All animal studies shall be performed in a facility meeting the requirements of ISO 10993-2.
5.2.4 Follow-up evaluation
If in vitro genotoxicity tests are performed in accordance with 5.2.2 and if the tests are negative, the test
sample can be considered non-genotoxic and further testing is not necessary.
Equivocal results (i.e. elevated results that do not meet criteria for a positive result), can require repeating
tests with a modified protocol such as adjusting dilution of the test sample or extract.
When any single in vitro test is positive or equivocal, then the following options shall be considered:
a) Identification of genotoxic constituents by chemical characterization (information gathering or
analytical chemistry testing) or other testing, with appropriate steps taken to eliminate or manage any
identified risks.
b) If confounding factors (e.g. osmolality, pH, cytotoxicity) are suspected to have caused the positive
response or equivocal in an initial genotoxicity test, the dosing or test conditions may be modified with
a justification to address confounding factors.
c) Presume that a genotoxicity risk exists and manage the risk in accordance with ISO 14971.
5.3 Test sample preparation
Test samples should be extracted in vehicles compatible with the test system (see Annex A). Use of extraction
vehicles that are incompatible with the test system requires additional manipulation (e.g. solvent removal)
that can alter the composition of the extract and, therefore, affect the results of the test. Use of extraction
vehicles that are incompatible with the test system shall be justified.
Extraction shall be performed in accordance with ISO 10993-12 unless otherwise justified.
Tests can be performed on solutions (e.g. soluble or liquid devices), suspensions (e.g. Method A in Annex A),
extracts (e.g. Method B in Annex A) of the finished product, medical device components or individual
chemicals from the medical device. If individual chemicals are tested, then the appropriate OECD guidelines
shall be followed.
NOTE Some authorities having jurisdiction prefer Method C in Annex A while others do not accept Method C.

ISO 10993-3:2026(en)
6 Carcinogenicity assessments
6.1 General
The objective of carcinogenicity studies is to determine tumorigenic potential of medical devices or medical
device constituents in animals and to evaluate the relevant risk to patients. Mechanisms of carcinogenesis
can be genotoxic or non-genotoxic. Medical devices shall be evaluated for both types of carcinogenicity,
preferably by alternative approaches, such as chemical characterization and toxicological risk assessment,
that do not use in vivo testing.
Animal testing for carcinogenicity is technically challenging. It requires implantat
...