SIST EN ISO 20166-3:2019

Molecular in vitro diagnostic examinations - Specifications for pre-examination processes for formalin-fixed and paraffin-embedded (FFPE) tissue - Part 3: Isolated DNA (ISO 20166-3:2018)

This International Standard recommends the handling, documentation, storage and processing of formalin fixed and paraffin embedded (FFPE) tissue specimens intended for DNA examination during the pre-examination phase before a molecular assay is performed. This International Standard is applicable to molecular in vitro diagnostic examinations including laboratory developed tests performed by medical laboratories and molecular pathology laboratories. It is also intended to be used by laboratory customers, in vitro diagnostics developers and manufacturers, but also pertains institutions and commercial organisations performing biomedical research, biobanks, and regulatory authorities.
DNA integrity in tissues can change before and during formalin fixation, processing and storage. Chemical modifications introduced into DNA during tissue fixation might lead to fragmentation and sequence alterations, changes in the methylation status or even structural changes which can lead to e.g., spurious copy number changes in array-CGH profiles. These modifications of the DNA molecules can impact the validity and reliability of the examination test results. Therefore, it is essential to take special measures to minimize the described modifications for subsequent DNA examination.
NOTE   International, national or regional regulations or requirements may also apply to specific topics covered in this International Standard.

Molekularanalytische in-vitro-diagnostische Verfahren - Spezifikationen für präanalytische Prozesse für formalinfixierte und paraffineingebettete (FFPE)-Gewebeproben - Teil 3: Isolierte DNS (ISO 20166-3:2018)

Diese Internationale Norm empfiehlt die Handhabung, Dokumentation, Lagerung und Verarbeitung von aus FFPE-Gewebe bestehendem und für die DNS-Untersuchung vorgesehenem Untersuchungsmaterial während der präanalytischen Phase vor Beginn der molekularen Analyse. Diese Internationale Norm gilt für molekulare in-vitro-diagnostische Untersuchungen, wozu auch im Labor entwickelte Prüfungen zählen, die von medizinischen Laboratorien und Laboratorien der molekularen Pathologie durchgeführt werden. Sie soll auch von Laborkunden, Entwicklern und Herstellern von In-vitro-Diagnostika sowie Einrichtungen und kommerziellen Organisationen, die in der biomedizinischen Forschung tätig sind, Biobanken und Aufsichtsbehörden eingesetzt werden.
ANMERKUNG   Zu bestimmten Bereichen, die in dieser Internationalen Norm behandelt werden, gelten möglicherweise lokale, nationale oder regionale Bestimmungen oder Anforderungen.

Analyses de diagnostic moléculaire in vitro - Spécifications relatives aux processus préanalytiques pour les tissus fixés au formol et inclus en paraffine (FFPE) - Partie 3: ADN extrait (ISO 20166-3:2018)

Le présent document fournit des lignes directrices concernant la manipulation, la documentation, le stockage et le traitement de prélèvements de tissus fixés au formol et inclus en paraffine (FFPE) destinés à l'analyse de l'ADN durant la phase préanalytique précédant la réalisation d'une analyse moléculaire.
Le présent document s'applique aux analyses de diagnostic moléculaire in vitro, y compris les analyses développées en laboratoire, réalisées par des laboratoires de biologie médicale et des laboratoires de pathologie moléculaire. Il est également destiné à être utilisé par des clients de laboratoires, des développeurs et fabricants de l'industrie du diagnostic in vitro, ainsi que par des biobanques, des institutions et des organismes commerciaux spécialisés en recherche biomédicale, de même que des autorités de réglementation.
NOTE       Des réglementations ou exigences internationales, nationales ou régionales peuvent également s'appliquer à des sujets spécifiques traités dans le présent document.

Molekularne diagnostične preiskave in vitro - Specifikacije za predpreiskovalne procese za tkiva, ki so fiksirana v formalinu ter položena v parafin - 3. del: Izolirani DNK (ISO 20166-3:2018)

Ta mednarodni standard vsebuje priporočila za obravnavo, dokumentiranje, shranjevanje in obdelavo vzorcev tkiv, ki so fiksirana v formalinu ter položena v parafin (FFPE), namenjenih za analizo DNK med predpreiskovalno fazo, preden se izvede molekularni preskus. Ta mednarodni standard se uporablja za molekularne diagnostične preiskave in vitro, vključno z laboratorijsko razvitimi preskusi, ki jih izvajajo v medicinskih laboratorijih in laboratorijih za molekularno patologijo. Uporabljali naj bi ga tudi uporabniki laboratorijev, razvijalci in proizvajalci diagnostike in vitro, nanaša pa se tudi na institucije in komercialne organizacije, ki izvajajo biomedicinske raziskave, biobanke ter regulativne organe. Celovitost DNK v tkivih se lahko spremeni pred fiksacijo v formalinu, obdelavo in shranjevanjem ter med temi postopki. Kemijske spremembe DNK med fiksacijo tkiva lahko privedejo do fragmentacije in sprememb zaporedja, sprememb v stanju metilacije ali celo strukturnih sprememb, kar lahko vodi do npr. lažnih sprememb števila kopij pri primerjalni genomski hibridizaciji z uporabo mikromrež (aCGH). Te spremembe molekul DNK lahko vplivajo na veljavnost in zanesljivost rezultatov preskusov preiskav. Zato je nujno treba sprejeti posebne ukrepe, da se zmanjšajo opisane spremembe za nadaljnje preiskave DNK. OPOMBA:   Za določene teme, ki so zajete v tem mednarodnem standardu, lahko veljajo tudi mednarodni, nacionalni ali regionalni predpisi ali zahteve.

General Information

• Status: Published
• Public Enquiry End Date: 14-Sep-2016
• Publication Date: 08-Apr-2019
• ICS: 11.100.10 - In vitro diagnostic test systems
• Technical Committee: VAZ - Healthcare
• Current Stage: 6060 - National Implementation/Publication (Adopted Project)
• Start Date: 04-Feb-2019
• Due Date: 11-Apr-2019
• Completion Date: 09-Apr-2019
• Ref Project: EN ISO 20166-3:2019 - Molecular in vitro diagnostic examinations - Specifications for pre-examination processes for formalin-fixed and paraffin-embedded (FFPE) tissue - Part 3: Isolated DNA (ISO 20166-3:2018)

Relations

• Replaces: SIST-TS CEN/TS 16827-3:2015 - Molecular in vitro diagnostic examinations - Specifications for pre-examination processes for FFPE tissue - Part 3: Isolated DNA
• Effective Date: 01-May-2019
• Replaces: SIST-TS CEN/TS 16827-3:2015 - Molecular in vitro diagnostic examinations - Specifications for pre-examination processes for FFPE tissue - Part 3: Isolated DNA
• Effective Date: 08-Jun-2022

Standards Content (Sample)

SLOVENSKI STANDARD
SIST EN ISO 20166-3:2019
01-maj-2019
1DGRPHãþD
SIST-TS CEN/TS 16827-3:2015
0ROHNXODUQHGLDJQRVWLþQHSUHLVNDYHLQYLWUR6SHFLILNDFLMH]DSUHGSUHLVNRYDOQH
SURFHVH]DWNLYDNLVRILNVLUDQDYIRUPDOLQXWHUSRORåHQDYSDUDILQGHO,]ROLUDQL
'1.,62
Molecular in vitro diagnostic examinations - Specifications for pre-examination processes
for formalin-fixed and paraffin-embedded (FFPE) tissue - Part 3: Isolated DNA (ISO
20166-3:2018)
Molekularanalytische in-vitro-diagnostische Verfahren - Spezifikationen für
präanalytische Prozesse für formalinfixierte und paraffineingebettete (FFPE)-
Gewebeproben - Teil 3: Isolierte DNS (ISO 20166-3:2018)
Analyses de diagnostic moléculaire in vitro - Spécifications relatives aux processus
préanalytiques pour les tissus fixés au formol et inclus en paraffine (FFPE) - Partie 3:
ADN extrait (ISO 20166-3:2018)
Ta slovenski standard je istoveten z: EN ISO 20166-3:2019
ICS:
11.100.10 'LDJQRVWLþQLSUHVNXVQL In vitro diagnostic test
VLVWHPLLQYLWUR systems
SIST EN ISO 20166-3:2019 en
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.

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SIST EN ISO 20166-3:2019

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SIST EN ISO 20166-3:2019


EN ISO 20166-3
EUROPEAN STANDARD

NORME EUROPÉENNE

January 2019
EUROPÄISCHE NORM
ICS 11.100.10 Supersedes CEN/TS 16827-3:2015
English Version

Molecular in vitro diagnostic examinations - Specifications
for pre-examination processes for formalin-fixed and
paraffin-embedded (FFPE) tissue - Part 3: Isolated DNA
(ISO 20166-3:2018)
Analyses de diagnostic moléculaire in vitro - Molekularanalytische in-vitro-diagnostische Verfahren
Spécifications relatives aux processus préanalytiques - Spezifikationen für präanalytische Prozesse für
pour les tissus fixés au formol et inclus en paraffine formalinfixierte und paraffineingebettete (FFPE)-
(FFPE) - Partie 3: ADN extrait (ISO 20166-3:2018) Gewebeproben - Teil 3: Isolierte DNS (ISO 20166-
3:2018)
This European Standard was approved by CEN on 21 December 2018.

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, Former Yugoslav Republic of Macedonia, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania,
Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Romania, Serbia, Slovakia, Slovenia, Spain, Sweden, Switzerland,
Turkey 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
© 2019 CEN All rights of exploitation in any form and by any means reserved Ref. No. EN ISO 20166-3:2019 E
worldwide for CEN national Members.

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SIST EN ISO 20166-3:2019
EN ISO 20166-3:2019 (E)
Contents Page
European foreword . 3

2

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SIST EN ISO 20166-3:2019
EN ISO 20166-3:2019 (E)
European foreword
This document (EN ISO 20166-3:2019) has been prepared by Technical Committee ISO/TC 212 "Clinical
laboratory testing and in vitro diagnostic test systems" in collaboration with Technical Committee
CEN/TC 140 “In vitro diagnostic 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 July 2019, and conflicting national standards shall be
withdrawn at the latest by January 2022.
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 CEN/TS 16827-3:2015.
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, Former Yugoslav Republic of Macedonia,
France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta,
Netherlands, Norway, Poland, Portugal, Romania, Serbia, Slovakia, Slovenia, Spain, Sweden, Switzerland,
Turkey and the United Kingdom.
Endorsement notice
The text of ISO 20166-3:2018 has been approved by CEN as EN ISO 20166-3:2019 without any
modification.


3

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SIST EN ISO 20166-3:2019
INTERNATIONAL ISO
STANDARD 20166-3
First edition
2018-12
Molecular in vitro diagnostic
examinations — Specifications for pre-
examination processes for formalin-
fixed and paraffin-embedded (FFPE)
tissue —
Part 3:
Isolated DNA
Analyses de diagnostic moléculaire in vitro — Spécifications relatives
aux processus préanalytiques pour les tissus fixés au formol et inclus
en paraffine (FFPE) —
Partie 3: ADN extrait
Reference number
ISO 20166-3:2018(E)
©
ISO 2018

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SIST EN ISO 20166-3:2019
ISO 20166-3:2018(E)

COPYRIGHT PROTECTED DOCUMENT
© ISO 2018
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: [phone]
Fax: [phone]
Email: [e-mail]
Website: www.iso.org
Published in Switzerland
ii © ISO 2018 – All rights reserved

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SIST EN ISO 20166-3:2019
ISO 20166-3:2018(E)

Contents Page
Foreword .iv
Introduction .v
1 Scope . 1
2  Normative references . 1
3  Terms and definitions . 1
4  General considerations . 4
5  Outside the laboratory . 5
5.1 Specimen collection . 5
5.1.1 General. 5
5.1.2 Information about the specimen donor/patient . 5
5.1.3 Information about the specimen . 5
5.1.4 Specimen processing . 6
5.2 Transport requirements . 6
6  Inside the laboratory . 7
6.1 Information about the reception of the specimen . 7
6.2 Formalin fixation of the specimen or sample(s) . 7
6.3 Evaluation of the pathology of the specimen and selection of the sample(s) . 8
6.4 Post-fixation of frozen samples . 9
6.5 Decalcification . 9
6.6 Processing and paraffin embedding . 9
6.7 Storage requirements .10
6.8 Isolation of DNA .10
6.8.1 General.10
6.8.2 General information for DNA isolation procedures .10
6.8.3 Using commercial kits .11
6.8.4 Using the laboratories’ own protocols .11
6.9 Quantity and quality assessment of isolated DNA .12
6.10 Storage of isolated DNA .12
Annex A (informative) Impact of the storage temperature on DNA Integrity in FFPE blocks
of tissue .14
Bibliography .16
© ISO 2018 – All rights reserved iii

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SIST EN ISO 20166-3:2019
ISO 20166-3:2018(E)

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 documents 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).
Attention is drawn to the possibility that some of the elements of this document may be the subject of
patent rights. ISO shall not be held responsible for identifying any or all such patent rights. Details of
any patent rights identified during the development of the document will be in the Introduction and/or
on the ISO list of patent declarations received (see www .iso .org/patents).
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 ISO/TC 212, Clinical laboratory testing and in vitro diagnostic
test systems.
A list of all parts in the ISO 20166 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 2018 – All rights reserved

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SIST EN ISO 20166-3:2019
ISO 20166-3:2018(E)

Introduction
Molecular in vitro diagnostics, including molecular pathology, has enabled significant progress in
medicine. Further progress is expected with new technologies analysing nucleic acids, proteins, and
metabolites in human tissues and body fluids. However, the profiles and/or integrity of these molecules
can change drastically during specimen collection, transport, storage and processing, thus making
the outcome from diagnostics or research unreliable or even impossible because the subsequent
examination assay will not determine the situation in the patient but an artificial molecular pattern
generated during the pre-examination process. Studies have been undertaken to determine the
influencing factors for the DNA examination from formalin-fixed and paraffin-embedded (FFPE) tissue.
These studies demonstrated that a standardization of the entire process from specimen collection to the
DNA examination is needed. This document draws upon such work to codify and standardize the steps
for FFPE tissue with regard to DNA examination in what is referred to as the pre-examination phase.
DNA integrity in tissues can change before, during and after formalin fixation, processing and storage.
Chemical modifications introduced into DNA during tissue fixation might lead to fragmentation and
sequence alterations, changes in the methylation status or even structural changes which can lead to,
for instance, spurious copy number changes in array-CGH profiles. These modifications of the DNA
molecules can impact the validity and reliability of the examination test results. Therefore, it is essential
to take special measures to minimize the described DNA changes and modifications for subsequent
examination.
In this document, the following verbal forms are used:
— "shall" indicates a requirement;
— "should" indicates a recommendation;
— "may" indicates a permission;
— "can" indicates a possibility or a capability.
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SIST EN ISO 20166-3:2019
INTERNATIONAL STANDARD  ISO 20166-3:2018(E)
Molecular in vitro diagnostic examinations —
Specifications for pre-examination processes for formalin-
fixed and paraffin-embedded (FFPE) tissue —
Part 3:
Isolated DNA
1 Scope
This document gives guidelines on the handling, documentation, storage and processing of formalin-
fixed and paraffin-embedded (FFPE) tissue specimens intended for DNA examination during the pre-
examination phase before a molecular assay is performed.
This document is applicable to molecular in vitro diagnostic examinations including laboratory
developed tests performed by medical laboratories and molecular pathology laboratories. It is also
intended to be used by laboratory customers, in vitro diagnostics developers and manufacturers,
biobanks, institutions and commercial organizations performing biomedical research, and regulatory
authorities.
NOTE International, national or regional regulations or requirements can also apply to specific topics
covered in this document.
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 15189:2012, Medical laboratories — Requirements for quality and competence
ISO 15190, Medical laboratories — Requirements for safety
3  Terms and definitions
For the purposes of this document, the terms and definitions given in ISO 15189 and the following apply.
ISO and IEC maintain terminological databases for use in standardization at the following addresses:
— ISO Online browsing platform: available at https: //www .iso .org/obp
— IEC Electropedia: available at http: //www .electropedia .org/
3.1
aliquot
portion of a larger amount of homogeneous material, assumed to be taken with negligible sampling error
Note 1 to entry: The term is usually applied to fluids. Tissues are heterogeneous and therefore cannot be
aliquoted.
Note 2 to entry: The definition is derived from References [25], [26], and [27].
© ISO 2018 – All rights reserved 1

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3.2
ambient temperature
unregulated temperature of the surrounding air
3.3
analyte
component represented in the name of a measurable quantity
[SOURCE: ISO 17511:2003, 3.2, modified — EXAMPLE has been removed.]
3.4
analytical test performance
accuracy, precision, specificity and sensitivity of a test to measure the analyte (3.3) of interest
Note 1 to entry: Other test performance characteristics such as robustness, repeatability can apply as well.
3.5
cold ischemia
condition after removal of the tissue from the body until stabilization or fixation
3.6
diagnosis
identification of a health or disease state from its signs and/or symptoms, where the diagnostic process
can involve examinations (3.8) and tests for classification of an individual's condition into separate and
distinct categories or subclasses that allow medical decisions about treatment and prognosis to be made
3.7
DNA
deoxyribonucleic acid
polymer of deoxyribonucleotides occurring in a double-stranded (dsDNA) or single-stranded
(ssDNA) form
[SOURCE: ISO 22174:2005, 3.1.2]
3.8
examination
analytical test
set of operations having the object of determining the value or characteristics of a property
Note 1 to entry: Processes that start with the isolated analyte and include all kinds of parameter testing or
chemical manipulation for quantitative or qualitative examination.
[SOURCE: ISO 15189:2012, 3.7, modified — Notes to entry 1 to 3 have been removed, Note 1 to entry has
been added and “analytical test” has been added as a preferred term.]
3.9
formalin
saturated aqueous formaldehyde solution which at 100 % contains 37 % formaldehyde by mass
(corresponding to 40 % by volume)
3.10
formalin fixation
treatment of a sample (3.18) with standard buffered formalin solution (3.20) for stabilization
3.11
grossing
gross examination
inspection of pathology specimens with the bare eye to obtain diagnostic information, while being
processed for further microscopic examination
2 © ISO 2018 – All rights reserved

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3.12
interfering substances
endogenous substances of a specimen (3.15)/sample (3.18) or exogenous substances (e.g. stabilization
solution) that can alter an examination result
3.13
paraffin embedding
process in which a tissue sample is placed in paraffin to generate a hard surrounding matrix so that
thin microscopic sections can be cut
3.14
pre-examination process
pre-analytical phase
pre-analytical workflow
process that starts, in chronological order, from the clinician’s request and includes the examination
request, preparation and identification of the patient, collection of the primary sample(s) (3.15),
transportation to and within the medical or pathology laboratory, isolation of analytes (3.3), and ends
when the analytical examination begins
Note 1 to entry: The pre-examination phase includes preparative processes that influence the outcome of the
intended examination.
[SOURCE: ISO 15189:2012, 3.15, modified — “pre-analytical workflow” has been added as a preferred
term, Note 1 to entry has been added and the definition has been extended.]
3.15
primary sample
specimen
discrete portion of a body fluid, breath, hair or tissue taken for examination (3.8), study or analysis of
one or more quantities or properties assumed to apply for the whole
[SOURCE: ISO 15189:2012, 3.16, modified — Notes to entry 1 to 3 have been removed.]
3.16
proficiency test
evaluation of participant performance against pre-established criteria by means of inter-laboratory
comparisons
[SOURCE: ISO 17043:2010, 3.7, modified — Notes to entry 1 and 2 have been removed.]
3.17
room temperature
for the purposes of this document, temperature in the range of 18 °C to 25 °C
Note 1 to entry: Local or national regulations can have different definitions.
3.18
sample
one or more parts taken from a primary sample (3.15)
[SOURCE: ISO 15189:2012, 3.24, modified — EXAMPLE has been removed.]
3.19
stability
ability of a sample material, when stored under specified conditions, to maintain a stated property
value within specified limits for a specified period of time
Note 1 to entry: The analyte for the purpose of this document is isolated DNA.
[SOURCE: ISO Guide 30:2015, 2.1.15, modified — “reference material” has been replaced by “sample
material" and Note 1 to entry has been changed.]
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3.20
standard buffered formalin solution
neutral buffered formalin
NBF
10 % formalin (3.9) solution in water with a mass fraction of 3,7 % (corresponding to a volume fraction
of 4 %) formaldehyde, buffered to pH 6,8 to pH 7,2
Note 1 to entry: Standard buffered formalin solutions often contain small amounts of methanol to inhibit
oxidation and polymerisation of formaldehyde.
3.21
storage
prolonged interruption of the pre-analytical workflow (3.14) of a sample (3.18) or analyte (3.3)
respectively, or of their derivatives, such as stained sections or tissue blocks, under appropriate
conditions in order to preserve their properties
Note 1 to entry: Long-term storage typically occurs in laboratory archives or in biobanks.
3.22
tissue processor
automated instrument where tissue fixation, dehydration, clearing and paraffin infiltration occurs
3.23
validation
confirmation, throughout the provision of objective evidence, that the requirements for a specific
intended use or application have been fulfilled
Note 1 to entry: “Validated” is used to designate the corresponding status.
[SOURCE: ISO 9000:2015, 3.8.13, modified — Notes to entry 1 and 3 have been removed.]
3.24
warm ischemia
condition before the tissue is removed from the body, but where it is deprived of its normal blood supply
3.25
workflow
series of activities necessary to complete a task
3.26
homogeneous
uniform in structure and composition
4  General considerations
For general statements on medical laboratory quality management systems and in particular on
specimen collection and handling (including avoidance of cross contaminations) see ISO 15189:2012,
4.2, 5.4.4, 5.4.6 or ISO/IEC 17020:2012, clause 8 and 7.2. The requirements for laboratory equipment,
reagents, and consumables in accordance with ISO 15189:2012, 5.3 shall be followed; ISO 15189:2012,
5.5.1.2 and 5.5.1.3, and ISO/IEC 17020:2012, 6.2 can also apply.
All steps of a diagnostic workflow can influence the final analytical test result. Thus, the entire
workflow including biomolecule stability and sample storage conditions shall be verified and validated.
Workflow steps which cannot always be controlled (e.g. warm ischemia) shall be documented. A risk
assessment of non-controllable workflow steps including their potential impact on the analytical test
performance shall be performed and mitigation measures shall be established to enable the required
analytical test performance.
In contrast to RNA or proteins, DNA in tissue is relatively stable during warm and cold ischemia.
Changes of DNA sequence or copy numbers [e.g. comparative genomic hybridization (CGH) profiles] due
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[7]
to longer warm and cold ischemia durations are unknown . However, DNA methylation patterns may
[6]
change in response to ischemia . The duration until the specimen is placed into standard buffered
formalin solution should be kept as short as possible in order to avoid enzymatic degradation of DNA.
The duration before fixation shall be documented and the temperature before fixation should be
[5]
documented .
During the fixation, processing and storage, the DNA integrity can change depending on the kind
of fixative, fixation time and temperature, storage or archiving of the fixed paraffin-embedded
tissue as well as the method used for DNA isolation and purification. When using a fixative based on
formaldehyde, temperature and fixation duration have a significant impact on DNA integrity. The longer
the fixation duration and the higher the temperature, the more chemical modifications and crosslinks
[9][10][11][12][13][14]
are introduced, which can lead to degradation or sequence alterations . These effects
can limit the size of amplifiable target DNA and/or influence the target sequence of primers used for
amplification.
Safety instructions on specimen transport and handling shall be considered and followed in accordance
with ISO 15189:2012, 5.2.3 and 5.4.5 and ISO 15190.
During the whole pre-examination process precautions shall be taken to avoid cross contamination
between different specimens/samples, e.g. by using single-use material whenever feasible or
appropriate cleaning procedures between processing of different specimens/samples.
If a commercial product is not used in accordance with the manufacturers' instructions, responsibility
for its use and performance lies with the user.
5  Outside the laboratory
5.1  Specimen collection
5.1.1  General
For the collection of the specimen, the requirements (e.g. disease condition, specimen size) for intended
molecular examination (see also Clause 6) should be considered.
See also ISO 15189:2012, 5.4.4.
5.1.2  Information about the specimen donor/patient
The documentation shall include the ID of the specimen donor/patient, which can be in the form of a code.
The documentation should include, but is not limited to:
a) the relevant health status of the specimen donor/patient [e.g. healthy, disease type, concomitant
disease, demographics (e.g. age and gender)];
b) the information about routine medical treatment and special treatment prior to tissue collection
(e.g. anaesthetics, medications, surgical or diagnostic procedures);
c) the appropriate consent from the specimen donor/patient.
5.1.3  Information about the specimen
The documentation shall include, but is not limited to:
a) the start of ischemia within the body (warm ischemia) by documentation of the ischemia-relevant
vessel ligation/clamping time point (usually arteria
...