Practical Guide to Specimen Handling in Surgical Pathology

Authors: Robert Lott, Janet Tunnicliffe, Elizabeth Sheppard, Jerry Santiago, Christa Hladik, Mansoor Nasim, Konnie Zeitner, Thomas Haas,
Shane Kohl, Saeid Movahedi-Lankarani

Practical Guide to Specimen Handling in Surgical Pathology
AUTHORS:
Robert Lott, Janet Tunnicliffe, Elizabeth Sheppard, Jerry Santiago, Christa Hladik, Mansoor Nasim, Konnie Zeitner, Thomas Haas, Shane Kohl, Saeid Movahedi-Lankarani

INTRODUCTION
In spite of the abundant guidelines and recommendations published for specimen handling and testing in a clinical pathology laboratory, relatively little literature is available for guidance of specimen handlingin a surgical pathology laboratory. This document does not relate to cytologic or clinical pathology samples.
The following comprehensive table is intended to serve as a general guideline for proper specimen handling from the time it is taken from the patient to the time a completed slide of the specimen is given to apathologist for interpretation.
DISCLAIMER:
This document was created by members of the CAP/NSH Histotechnology Committee and is intended to serve as a guideline ONLY and NOT AN absolute recommendation for specimen handling. Each laboratory
is advised to use these guidelines as a starting point and modify certain parameters to fit state and local institutional requirements, as appropriate. Regulatory references, standards, and CAP checklist items citedin the guideline are current at the time of publication of this version of the guideline. It is recommended that the user confirm all references used are the latest version available. The use of the information
contained in this guideline does not guarantee compliance with the CAP accreditation requirements or regulations from other accrediting organizations. Some information may be different or more stringent
than the published CAP Checklists.
It is the intent of the CAP/NSH Histotechnology Committee to update this document every 2 years or when required and have the updated version of the document available to members on the College ofAmerican Pathologists (CAP) and National Society for Histotechnology (NSH) websites.

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Table of Contents:
Part I – Specimen Collection and Handling
A. Patient Identification …………………………………………………………………
B. Proper Labeling ……………………………………………………………………...
C. Transport Media ……………………………………………………………………..
D. Completion of Requisition ………………………………………………………….
E. Recommendations for Tissue Collection and Handling …………………………
F. Accessioning ………………………………………………………………………...
G. Handling prior to Gross Examination ……………………………………………...
H. Intra-operative Consultation ………………………………………………………..

pg. 4
pg. 5
pg. 7
pg. 10
pg. 17pg. 26
pg. 27pg. 29Part II – Laboratory Processes
A. Guidelines ……………………………………………………………………………
B. Tissue Cassette Identification ……………………………………………………..
C. Fixation Parameters ………………………………………………………………..
D. Processing …………………………………………………………………………..
E. Embedding …………………………………………………………………………..
F. Microtomy ……………………………………………………………………………
G. Staining ………………………………………………………………………………
H. Coverslipping ………………………………………………………………………..

pg. 32
pg. 38pg. 39pg. 46
pg. 52
pg. 53
pg. 58
pg. 70

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VERSIONREVISION DATEREVISION

2.0

November, 2013

3.0

November, 2014

1. Revised per comments received from CAP Chair review

4.0

January, 2015

1. Updated references – CAP Checklists: ANP, COM, GEN, 4-21-2014
2. All references reviewed
3. Table of contents added

5.0

September, 2015

1. Updated to reflect LAP Committee 2015 Checklist changes

6.0

November, 2015

1. Updated to reflect corrected formalin solution to tissue ratio with references

7.0

September, 2017

1. Updated to reflect August 21, 2017 CAP Checklist edition changes

8.0

September, 2018

1. Updated to reflect August 22, 2018 CAP Checklist edition changes
2. Updated to reflect review of all references

9.0

April, 2020

1. Updated to reflect June 4, 2020 CAP Checklist edition Changes
2. Updated to reflect review of all references
3. Updated Table of Contents
4. Updated Title Page and organizational logos

10.0

October, 2020

1. Updated organizational logos
2. Updated to reflect September 22, 2021 CAP Checklist edition changes

11.0

September, 2023

1. Updated references
2. Updated to reflect August 24, 2023 CAP Checklist edition changes

1. Addition of disclaimer on cover page
2. Addition of version control

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PART I

I. SPECIMEN COLLECTION and HANDLING

Guideline Section
Collection and Handling
A. Patient Identification

Statement

Related CAP Checklist Requirements
2023 Edition•

Patient is to be identified in a manner that respects patient privacy with respect totheir medical records and medical data.

Laboratory General Checklist, GEN.41303(Patient Confidentiality)•

Patient's identity must be verified at the time of specimen collection.

Laboratory General Checklist, GEN.40490(Patient Identification)•

At least two acceptable patient-specific identifiers are required for patient
identification:
o
o
o
o
o

Full name
Assigned identification number (e.g. health record / master index number)
Date of birth
Photo on government issued or other photo ID card, such as driver's license
Other specific personal identifiers

Laboratory General Checklist, GEN.40491(Primary Specimen Container Labeling)

Additional References

Health Insurance and Portability andAccountability Act (HIPAA).
Clinical Laboratory Standards Institute
CLSI - GP33A, Accuracy in Patient andSample Identification; 2019.
International Standard ISO 15189:2012- Medical Laboratories; section 5.4 Pre-examination Processes

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Collection and Handling
B. Proper Labelling•

Specimen is labeled in the presence of the patient.

•

Specimen label must contain at least two patient-specific identifiers:

•

•

•

o

Full patient nameoAssigned identification number (e.g. health record / master index number)oDate of BirthLaboratory General Checklist, GEN.40490(Patient Identification)

Laboratory General Checklist, GEN.40115(Specimen Collection Manual Elements –Surgical Pathology and Cytopathology
Specimens)

Customizable label elements – additional identifiers that are acceptable:oPatient genderoAccession or requisition numberoOrdering physicianoSource of specimen (e.g. skin)oSite of specimen (e.g. left side of chest)

Laboratory General Checklist, GEN. 40491(Primary Specimen Container Labeling)

Clinical Laboratory Standards Institute
CLSI – Auto12-A Specimen Labels:Content and Location, Fonts and Label
Orientation: 2011.

Standardized format for label information should be implemented.
oLast name, first nameoDate of Birth DD–MM–YYYY (i.e. 12 MAR 1968)

oGender M, F, U (unknown), T (Transgender), I (Intersex)

Written documentation developed for the correct positioning of the label on thecollection container.
o

All Common Checklist, COM.06100(Primary Specimen Container Labeling)
All Common Checklist, COM.06200(Secondary Specimen Container Labeling)

Brown RW, Della Speranza V, Alvarez
JO, et al. Uniform labeling of blocks andslides in surgical pathology: Guidelinefrom the College of American
Pathologists Pathology and Laboratory
Quality Center and the National Societyfor Histotechnology. Arch Pathol Lab
Med. 2015;139(12):1515-24.

Do not attach label to the container lid (in whole or part)

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o
•

Do not overlap label resulting in patient data being covered

Written documentation for the correction of labelling errors – to be followed when
specimens cannot be replaced.
Laboratory General Checklist, GEN.40492(Specimen Label Correction)•

All subsequent labelling of patient samples (blocks and slides) must follow same
patient-specific identifying process.

•Submitted slides may be labeled with a single patient-specific identifier but two
are preferred.

•

All parameters used for standard specimen labelling are to be followed.

•

The unique specimen bar code or RFID label must be consistent across all
applications: specimen container, requisition label, cassette and slide labels.

•

Barcode and RIFD specifications within a failure rate established by your facility
for patient care.

•

Barcode label stock or RFID chip validated to withstand chemicals andprocessing used for anatomic pathology specimens.

Laboratory General Checklist, GEN.40825(Specimen ID)

Laboratory General Checklist, GEN.40491(Primary Specimen Container Labeling)

Collection and Handling
B. Proper Labellingi. Barcoding and/orRadio Frequency
Identification(RFID)

Laboratory General Checklist, GEN.40825(Specimen ID)

Zarbo RJ, Tuthill JM, D’Angelo R, et al.
The Henry Ford Production System:
reduction of surgical pathology inprocess misidentification defects by bar
code-specified work process
standardization. Am J Clin Pathol. 2009;
131:469-477.
Clinical Laboratory Standards Institute
CSLI – Auto02-A2 Laboratory
Automation: Bar Codes for Specimen
Container Identification: 2006.

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•Bar coding and/or RFID documentation must be validated and maintained.

•

Automatic identification scanning equipment is validated for accuracy andresistant to chemicals used for anatomic pathology handing.

•If used for specimen chain of custody tracking, the barcode or RFID tracking
system must have intelligent location capabilities.

•

Collection, handling and submission procedures must be made available to all
health care workers involved in the collection, labeling, submission and transportof specimens to the pathology laboratory.

•

All specimens must be placed in leak proof container.

•

Specimens should be transported to the laboratory immediately after collection.

Collection and Handling
C. Transport Mediai. No media / saline

Laboratory General Checklist, GEN.40115(Specimen Collection Manual Elements –Surgical Pathology and Cytopathology
Specimens)

All Common Checklist, COM.06000(Specimen Collection Manual)

Clinical Laboratory Standards Institute
CLSI – GP33A, Accuracy in Patient andSample Identification; 2019.

International Standard ISO 15189:2012- Medical Laboratories; section 16 Preexamination.

Laboratory General Checklist, GEN.74500(Specimen Transport Procedures)•

Specimens that cannot be immediately transferred must be refrigerated until
transferred to the Pathology laboratory.
••

For specimens submitted to the laboratory from remote sites, there is adocumented tracking system to ensure that all specimens are actually
received.

Specimens transferred from distant referral site to pathology lab should be
shipped under temperature-controlled conditions to avoid over heating or
freezing.

Laboratory General Checklist, GEN.40125(Handling of Referred Specimens)

Laboratory General Checklist, GEN.40511(Specimen Tracking/Labeling)

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•

Policies regarding courier service should be established.

•

All specimens must be properly packaged and labelled, indicating materials to betransported prior to shipping to a centralized or referral laboratory.

Laboratory General Checklist, GEN.40535(Specimen Transport QM)

Laboratory General Checklist, GEN.40530(Specimen Tracking)•Bancroft J, Gamble M. Theory andPractice of Histological Techniques, 6thed. New York, NY: Churchill Livingston;
2008.

Carson F, Hladik C. Histotechnology A
Self-Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020.

To avoid drying of tissues that are not immediately placed into formalin at time ofprocurement:oWrap solid tissue masses (i.e. lymph node or breast lump) in saline
dampened gauze prior to placement in labelled container (certain biopsies
may need special handling).

o

Add a small volume of saline to tissue with insufficient naturally occurring
fluids (i.e. conceptus for embryopathology/genetic studies).

Laboratory General Checklist, GEN.40535(Specimen Transport QM)

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Collection and Handling
C. Transport Media•ii. Different fixatives

Collection, handling and submission procedures must be made available to all
health care workers involved in the collection, labelling, submission and transportof Specimens to the pathology laboratory.

Laboratory General Checklist, GEN.40115(Specimen Collection Manual Elements –Surgical Pathology and Cytopathology
Specimens)

All Common Checklist, COM.06000(Specimen Collection Manual)•

All specimens must be placed in leak proof container.
Laboratory General Checklist, GEN.74500(Specimen Transport Procedures)

•

•

•

Specimens must be placed in appropriate fixative as specified incollection/handling and submission procedure.

Bancroft J, Gamble M. Theory andPractice of Histological Techniques, 6thed. New York, NY: Churchill Livingston;
2008.

Carson F, Hladik-Cappellano C.
Histotechnology A Self- Instructional
Text, 5th ed. Chicago, IL: ASCP Press;
2020.

Brown RW. et. al., Histologic
Preparations Common Problems andTheir Solutions. College of American
Pathologists, 2009.

Volume of fixative to tissue ratio must be included in the collection/handling andsubmission procedures. i.e. 10% neutral buffered formalin volume should be 1520 times the volume of the specimen.

Safety Data Sheets (SDS) must be made available to all staff handling fixatives.

International Standard ISO 15189:2012- Medical Laboratories; section 5.4 Pre-examination Processes.

Laboratory General Checklist, GEN.76100(Chemical Safety Document Access)Clinical Laboratory Standards Institute
CLSI – GP 17-A3, Clinical Laboratory
Safety, 3rd edition; 2012.

Occupational Health and Safety
Administration. Occupational Safety &Health Standards 1910.1200 toxic andHazardous Substances.

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•

•

All specimen containers containing fixatives must have appropriate OSHA
Chemical labels attached.

Specimens transferred from distant referral site to Pathology lab should be
shipped under temperature-controlled conditions to avoid over heating or
freezing.

•

Specimen containers should be shipped following appropriate regulations for the
shipping and handling of formalin i.e. hard sided container with absorbent packing
material.

•

Written procedures on how to properly complete a pathology requisition must be
made available to all health care workers involved in the collection, labelling,
submission, and transport of specimens to the pathology laboratory.

Laboratory General Checklist, GEN.40125(Handling of Referred Specimens)

http://www.osha.gov/dsg/hazcom/index.
html

Laboratory General Checklist, GEN.40511(Specimen Tracking/Labeling)

Laboratory General Checklist, GEN.40535(Specimen Transport QM)

Collection and Handling
D. Completion ofrequisition
i. Patient identifiers•

Written or electronic request for patient testing from authorized person.

Laboratory General Checklist, GEN.40750(Requisition Elements)

Laboratory General Checklist, GEN.40930(Authorized Requestor)

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10•Clinical Laboratory Standards Institute
CLSI - GP33A, Accuracy in Patient andSample Identification; 2019.

Required patient identifiers to be included on the requisition / test order:o Patient’s nameo Unique identifier i.e. health record or master index numbero Date of BirthInternational Standard ISO 15189:2012- Medical Laboratories; section 5.4- Preexamination Processes.

o Sex
Collection and Handling
D. Completion ofrequisition
ii. Specimen
name/type/site• Written or electronic request for patient testing to include:
o
o
o
o
o
o
o
o
o

Patient identifiers as listed above
Name and address or other suitable identifiers of the authorized person
requesting the test
Name and address or other suitable identifier for the individual
responsible for receiving the test results
Name and address of the laboratory submitting the specimen
Test and or tests to be performed
Procedure performed
Specimen site – if more than one specimen is collected during a single
procedure; each specimen should be individually identified by anatomic
site and or specimen type
Date and time of procedure or specimen collection
Date specimen received

Laboratory General Checklist, GEN.40930(Authorized Requestor)

Clinical Laboratory Standards Institute
CLSI - GP33A, Accuracy in Patient andSample Identification; 2019.

Laboratory General Checklist, GEN.40750(Requisition Elements)

International Standard ISO 15189:2012- Medical Laboratories; section 5.4 Pre-examination Processes

Laboratory General Checklist, GEN.40900(Specimen Date Received)

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Collection and Handling
D. Completion of•

Written or electronic request for patient testing to include:orequisitioniii. Pertinent clinical

Clinical history – any additional information relevant or necessary for a
specific test to ensure accurate and timely testing and reporting of results,
including interpretation if required.

Laboratory General Checklist, GEN.40750(Requisition Elements)

Health Insurance and Portability andAccountability Act (HIPAA).

Clinical Laboratory Standards Institute
CLSI - GP33A, Accuracy in Patient andSample Identification; 2019.

history

International Standard ISO 15189:2012- Medical Laboratories; section 5.4- Preexamination Processes

Collection and Handling
D. Completion of•

The procedure date should be indicated on the requisition following standardized
format DD–MM–YYYY (i.e. 04 JAN 2012).

•

The requisition must have a space for the documentation of the warm ischemic
time by the physician obtaining the specimen or designate.

•

Warm ischemic time:
The time measured from the interruption of the blood supply to the tissue/tumor
by the surgeon to the excision time of the tissue specimen.

requisition
iv. Procedure
time/datea. Time removed
from patient(Warm ischemic time)•

Collection and Handling

Information should be available in the laboratory for review and/or appear on thepatient accession.

Laboratory General Checklist, GEN.40750(Requisition Elements)

Allison KH, Hammond EH, Dowsett M,
McKernin SE et al. Estrogen andProgesterone Receptor Testing in
Breast Cancer American Society ofClinical Oncology/College of American
Pathologists Guideline Update. ArchPath Lab Med. Early Online Release.
doi: 10.5858/arpa.2019-0904-SA
International Standard ISO 201664:2020 - Molecular in vitro diagnostic
examinations – Specifications for preexamination processes for formalinfixed and paraffin-embedded (FFPE)
tissue for – Part 4: In situ detection
techniques: section 6 – Inside thelaboratory.

Allison KH, Hammond EH, Dowsett M,
McKernin SE et al. Estrogen andVersion: 11.0
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D. Completion of•

The requisition should have a space for the documentation of the cold ischemic
time by the physician obtaining the specimen or designate.

•

Cold ischemic time:
The time from excision of the specimen from the surgical field to the time thetissue is placed in fixative.

requisition
iv. Procedure
time/dateb. Time fixative•

Information should be available in the laboratory for review and/or appear on thepatient accession.

•

The requisition should have a space for the documentation of the date and time
the specimen is placed in fixative by the physician obtaining the specimen or
designate.

added(if required)(cold ischemic time)

Anatomic Pathology Checklist, ANP.22983(Fixation – HER2 and ER Breast Cancer
Predictive Marker Testing)Laboratory General Checklist, GEN.40125(Handling of Referred Specimens)

Progesterone Receptor Testing in
Breast Cancer American Society ofClinical Oncology/College of American
Pathologists Guideline Update. ArchPath Lab Med. Early Online Release.
doi: 10.5858/arpa.2019-0904-SA
Clinical Laboratory Standards Institute
CLSI – MM13, Collection, Transport,
Preparation, and Storage of Specimens
for Molecular Methods: 2020.
Compton CC, Robb JA, Anderson
MW, Berry AB, et.al. Preanalytics andPrecision Pathology: Pathology
Practices to Ensure Molecular Integrityof Cancer Patient Biospecimens for
Precision Medicine. Arch Path Lab
Med. Nov 2019, Vol. 143, No. 11(November 2019) pp. 1346-1363.
International Standard ISO 201664:2020 - Molecular in vitro diagnostic
examinations – Specifications for preexamination processes for formalinfixed and paraffin-embedded (FFPE)
tissue for – Part 4: In situ detection
techniques: section 6 – Inside thelaboratory.

Collection and Handling

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D. Completion of•

The requisition must have a space for documentation of the date and time ofarrival of the specimen in the AP laboratory to allow for calculation of thetransport time.

Laboratory General Checklist, GEN.40535(Specimen Transport QM)•

Transport time:
The time tissue specimen was collected in the operating room/doctor’s
office/clinic until it is received in the pathology laboratory for processing (this is
the time point when the specimen is going to be grossly assessed).

Laboratory General Checklist, GEN.40530(Specimen Tracking)•Information must be available in the laboratory for review and/or appear on thepatient accession.

requisition
iv. Procedure
time/datec. Time received inlab(Transport time)

Clinical Laboratory Standards Institute
CLSI – MM13, Collection, Transport,
Preparation, and Storage of Specimens
for Molecular Methods: 2020.

Collection and Handling
D. Completion ofrequisition
iv. Procedure
time/dated. Calculation of
total fixation time•

The laboratory has the responsibility to calculate and document total time thespecimen was kept in fixative for required specimens (i.e. breast).
To include:oTime specimen held in the operating roomoTransport time from remote site to AP laboTime the specimen was kept in fixative while in the lab (i.e. large specimens
like colon, breast mastectomy were opened/cut to allow for penetration of
fixative)oTime the specimen(s) are kept in cassettes after grossingoTime in fixative onboard the tissue processor

Allison KH, Hammond EH, Dowsett M,
McKernin SE et al. Estrogen andProgesterone Receptor Testing in
Breast Cancer American Society ofClinical Oncology/College of American
Pathologists Guideline Update. ArchPath Lab Med. Early Online Release.
doi: 10.5858/arpa.2019-0904-SA

Anatomic Pathology Checklist, ANP.22983(Fixation – HER2 and ER Breast Cancer
Predictive Marker Testing)

Allison KH, Hammond EH, Dowsett M,
McKernin SE et al. Estrogen andProgesterone Receptor Testing in
Breast Cancer American Society ofClinical Oncology/College of American
Pathologists Guideline Update. ArchPath Lab Med. Early Online Release.
doi: 10.5858/arpa.2019-0904-SA
Wolff AC, Hammond EH, Hicks DG,
Dowsett M, et al: American Society ofClinical Oncology/College of American
Pathologists Guideline Update
Recommendations for Human
Epidermal Growth Factor Receptor 2
Testing in Breast Cancer. Journal of
Clinical Oncology. Vol 31, No. 31, Nov1
2013: pp. 3997-4013.

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Collection and Handling•Tissue handling requirements should be standardized and reported on every
specimen.

•

10% neutral buffered formalin is the recommended fixative.

•

All samples must receive a minimum of six (6) hours of 10% neutral bufferedformalin fixation.

•

Recommended fixation time is 6 to 72 hours for estrogen and progesterone
receptors.

D. Completion ofrequisition
iv. Procedure
time/datee. Fixation time forbreast tissue
specimens•

Recommended fixation time is 6 to 72 hours for Her2neu receptors.

•

Fixation time must be documented, and the following is an example of how thedata could be recorded on the requisition:

Time frame

Warm ischemic time
Cold ischemic time

Transport time from OR /physician office /clinic tolaboratory to time of primary examination

Time whole specimen held for additional fixation prior
to placing in cassettes
Time cassettes are held prior to loading onto tissue
processor
Fixation time on tissue processor (delay time plus
processing time)
Total Fixation time

Minutes

Hours

Laboratory General Checklist, GEN.40115(Specimen Collection Manual Elements –Surgical Pathology and Cytopathology
Specimens)

Laboratory General Checklist, GEN.40125(Handling of Referred Specimens)

Anatomic Pathology Checklist, ANP.22983(Fixation - HER2 and ER Breast Cancer
Predictive Marker Testing)Anatomic Pathology Checklist, ANP.23004(Digital Imaging – Preanalytic Testing Phase
Validation)

Wolff AC, Hammond ME, Allison KH,
Harvey BE, et al. Human Epidermal
Growth Factor Receptor 2 Testing in
Breast Cancer: American Society ofClinical Oncology/College of American
Pathologists Clinical Practice Guideline
Focused Update. Arch Path Lab Med.
Nov 2018, Vol. 142, No. 11. pp. 13641382.
Allison KH, Hammond EH, Dowsett M,
McKernin SE et al. Estrogen andProgesterone Receptor Testing in
Breast Cancer American Society ofClinical Oncology/College of American
Pathologists Guideline Update. ArchPath Lab Med. Early Online Release.
doi: 10.5858/arpa.2019-0904-SA
Clinical Laboratory Standards Institute
CLSI – MM13, Collection, Transport,
Preparation, and Storage of Specimens
for Molecular Methods: 2020.
Werner M, Chott A, Fabiano A, Battifora
H. Effect of Formalin Tissue Fixation
and Processing on
Immunohistochemistry. American
Journal of Surgical Pathology. 24. July
2000:1016-1019.
Spruessel A, Steimann G, Jung M, Lee
SA, Carr T, Fentz AK, Spangenberg J,
Zornig C, Juhl HH, David KA. Tissue
ischemia time affects gene and protein
expression patterns within minutes
following surgical tumor excision
BioTechniques. Vol. 36, No. 6, June
2004:1030–1037.

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Petersen BL, Sorensen MC, Pedersen
S, Rasmussen M. Fluorescence In-situ
Hybridization on Formalin-fixed andParaffin-Embedded Tissue: Optimizing
the Method. Appl Immunohistochem
Mol Morphol. 12(3) September
2004:259-265.
Tanney A, Kennedy RD. Developing
mRNA-based biomarkers from formalinfixed paraffin-embedded tissue. Per
Med (2010) 7(2), 205–211.
Compton CC, Robb JA, Anderson
MW, Berry AB, et.al. Preanalytics andPrecision Pathology: Pathology
Practices to Ensure Molecular Integrityof Cancer Patient Biospecimens for
Precision Medicine. Arch Path Lab
Med. Nov 2019, Vol. 143, No. 11 pp.
1346-1363.
Collection and Handling
D. Completion ofrequisition
iv. Procedure
time/datef. Fixation time for
NON-breast
specimens•

Establish standardized fixation times for all routine and specialized biopsies.

•

Document the recommended fixative for routine and specialized biopsies.

•

Establish specimen acceptance and rejection policies related to specimen
fixation.

Laboratory General Checklist, GEN.40115(Specimen Collection Manual Elements Surgical Pathology and Cytopathology
Specimens)
All Common Checklist, COM.06300(Specimen Rejection Criteria)

Department of Health and Human
Services, Centers for Medicare andMedicaid Services. Clinical laboratory
improvement amendments of 1988;
final rule. Fed Register. 2003(Jan 24):
[42CFR493.1283(a)(3)]Compton CC, Robb JA, Anderson
MW, Berry AB, et.al. Preanalytics andPrecision Pathology: Pathology
Practices to Ensure Molecular Integrityof Cancer Patient Biospecimens for
Precision Medicine. Arch Path Lab
Med. Nov 2019, Vol. 143, No. 11(November 2019) pp. 1346-1363.

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Collection and Handling
D. Completion of•requisition

When alternate identifier is used for authorized person requesting test or
receiving test results (medical billing number, hospital ID number), the number
must be unique and traceable in the LIS.

v. Requesting

Laboratory General Checklist, GEN.40750(Requisition Elements)

Health Insurance and Portability andAccountability Act (HIPAA).

Clinical Laboratory Standards Institute
CLSI - GP33A, Accuracy in Patient andSample Identification; 2019: Vol 30
No7.

physiciana. contact information
available in LIS

International Standard ISO 15189:2012- Medical Laboratories; section 5.4 Pre-examination Processes.
Collection and Handling
E. Recommendations for•

The use of surgical instruments driven by heat should be avoided or limited whenpossible.

•

Thermal injury has been known to interfere with diagnosis.

•

The use of surgical instruments should be avoided or limited as much as
possible when handing the specimen to prevent crushing or damaging the tissue.

Tissue Collection and
Handling
i. Limiting Artifacts

Association of Surgical Technologists(AST) Recommended Standards ofPractice for Handling and Care ofSurgical Specimens. www.ast.org

a. Thermal injury
Collection and Handling
E. Recommendations for
Tissue Collection and
Handling

Association of Surgical Technologists(AST) Recommended Standards ofPractice for Handling and Care ofSurgical Specimens. http://www.ast.org

i. Limiting Artifacts
b. Crush injury

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Collection and Handling
E. Recommendations for•

All tissue should be placed in fixative as soon as possible after removal from thebody, unless special studies are ordered that might be affected by the available
fixative.

•

If fixative cannot be added in a timely manner, the specimen should be placed ina sterile basin and kept moist with sterile saline or wrapped in saline-dampened
sponges until the specimen can be properly placed in fixative.

•

All unfixed specimens should be transported to the pathology laboratory as soon
as possible and refrigerated until placed into appropriate fixative.

Tissue Collection andAnatomic Pathology Checklist, ANP.11250(Adequate Storage)

Handling
i. Limiting Artifactsc. Drying artifact

Association of Surgical Technologists(AST) Recommended Standards ofPractice for Handling and Care ofSurgical Specimens. www.ast.org
Makary MA, Epstein J, Pronovost PJ,
Millman EA, Hartmann EC, Freischlag
JA. Surgical specimen identification
errors: A new measure of quality insurgical care. Surgery. 2007.141:450455.

Laboratory General Checklist, GEN.40535(Specimen Transport QM)

Collection and Handling
E. Recommendations for•

Health care facility policy and procedure should be followed for the propercollection, labeling, and transportation of the specimen to the pathology
department.

•

All fresh specimens are to be submitted to the pathology department as soon
as possible with instructions for special testing or processes.

•

All unfixed specimens should be transported to the pathology laboratory assoon as possible and refrigerated until placed into appropriate fixative.

•

Specimens not in fixative should be placed in a sterile basin and kept moist
with sterile saline or wrapped in saline-soaked sponges until the specimen
can be properly placed in fixative.

Tissue Collection and
Handling
ii. Tissue Transport
a. All fresh
specimens•

Confirmation with surgeon on other types of diagnostic studies to be
performed, including Gram stain, acid fast and mycological studies.

Laboratory General Checklist, GEN.40115(Specimen Collection Manual Elements Surgical Pathology and Cytopathology
Specimens)

Clinical Laboratory Standards Institute
CLSI – MM13, Collection, Transport,
Preparation, and Storage of Specimens
for Molecular Methods: 2020.
Makary MA, Epstein J, Pronovost PJ,
Millman EA, Hartmann EC, Freischlag
JA. Surgical specimen identification
errors: A new measure of quality insurgical care. Surgery. 2007.141:450455.
Slavin L, Best MA, Aron DC. Gone but
not forgotten: The search for the lost
surgical specimens: Application ofquality improvement techniques for
reducing medical error. Quality
Management in Health Care. 2001.
10(1): 45-53.

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18

•The Joint Commission. (2022) 2022
National Patient Safety Goals Hospital
Program.

Exceptions to immediate delivery of tissue specimen must be clearly
described in the policies and procedures. (Example: Placentas must be
refrigerated until delivery).

US Dept of Health and Human
Services. Summary of the HIPAA
privacy rule. 2003.
World Health Organization. Guidelines
for the safe transport of infectious
substances and diagnostic specimens.
1997.
Carson F, Hladik C. Histotechnology A
Self-Instructional Text, 5th ed. Chicago,
IL: ASCP Press 2020.
Bancroft J, Gamble M. Theory andPractice of Histological Techniques, 6thed. New York, NY: Churchill Livingston;
2008.

Collection and Handling
E. Recommendations for•Specimen in fixative must be delivered to the pathology laboratory according to
the Health care facility policies and procedures.

Laboratory General Checklist, GEN.40535(Specimen Transport QM)•

Special guidelines are required for the handling of breast tissues to ensure
fixation guidelines are met (please see section D, iv, e for specific fixation times).

Anatomic Pathology Checklist, ANP.22983(Fixation - HER2 and ER Breast Cancer
Predictive Marker Testing)Tissue Collection and
Handling
ii. Tissue Transport
b. Specimens infixative•

Containers should be rigid, impermeable, unbreakable, and non-reactive tofixative solutions.

Laboratory General Checklist, GEN.40942(Specimen Container Analytic Interference)

Association of Surgical Technologists(AST) Recommended Standards ofPractice for Handling and Care ofSurgical Specimens. www.ast.org

World Health Organization. Guidelines
for the safe transport of infectious
substances and diagnostic specimens.
1997.

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Collection and Handling
E. Recommendations for•

Tissue Collection andDocumentation of fixation time for Breast specimens is required as outlined in
section C.

Anatomic Pathology Checklist, ANP.22983(Fixation – HER2 and ER Breast Cancer
Predictive Marker Testing)Handling
ii. Tissue Transportc. Monitoring of time•and environmental
parameters during

All specimens are received in the pathology laboratory according to the policies
and procedures approved, to include the acceptance of specimen protocol astime received, accessioned, and grossed.

Laboratory General Checklist, GEN.40115(Specimen Collection Manual Elements –Surgical Pathology and Cytopathology
Specimens)

transport

Laboratory General Checklist, GEN.40125(Handling of Referred Specimens)•

Specimen placed in different environment (i.e. dry ice) must be recorded anddelivered with specimen.

Laboratory General Checklist, GEN.40535(Specimen Transport QM)

Compton CC, Robb JA, Anderson
MW, Berry AB, et.al. Preanalytics andPrecision Pathology: Pathology
Practices to Ensure Molecular Integrityof Cancer Patient Biospecimens for
Precision Medicine. Arch Path Lab
Med. Nov 2019, Vol. 143, No. 11 pp.
1346-1363.
Allison KH, Hammond EH, Dowsett M,
McKernin SE et al. Estrogen andProgesterone Receptor Testing in
Breast Cancer American Society ofClinical Oncology/College of American
Pathologists Guideline Update. ArchPath Lab Med. Early Online Release.
doi: 10.5858/arpa.2019-0904-SA
Wolff AC, Hammond ME, Allison
KH, Harvey BE, et al. Human Epidermal
Growth Factor Receptor 2 Testing in
Breast Cancer: American Society ofClinical Oncology/College of American
Pathologists Clinical
Practice Guideline Focused Update.
Arch Path Lab Med. Nov 2018, Vol.
142, No. 11. pp. 1364-1382.
AST Recommended Standards ofPractice for Handling and Care ofSurgical Specimens.
The Joint Commission. (2022) 2022
National Patient Safety Goals Hospital
Program.

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Collection and Handling
E. Recommendation fortissue collection and•Chain of custody ensures continuity of quality care for the patient and provides amethod to retrieve needed information.

•

All specimens must be recorded on a chain of custody form or log that includes
dates and times, patient identification, specimen number, specimen description,and purpose for specimen delivery to the pathology department.

handling
ii. Tissue Transport
d. Chain of custody
1. Specimen
removal fromThe Joint Commission. (2022) 2022
National Patient Safety Goals Hospital
Program.

US Dept of Health and Human
Services. Summary of the HIPAA
privacy rule. 2003.

World Health Organization. Guidelines
for the safe transport of infectious
substances and diagnostic specimens.
1997.

origin ofCollection(time/date)

Collection and Handling
E. Recommendation fortissue collection andhandling
ii. Tissue Transport
d. Chain of custody
2. Personnel
transporting•

It is advisable that chain of custody include the personnel involved in thehandling and transportation of the specimen to the pathology lab and within thepathology lab during testing procedures.
o Name of transportero Title (i.e. RN, Surgical Tech, MD)o Dates: Collection, transported and received

The Joint Commission. (2022) 2022
National Patient Safety Goals Hospital
Program.
US Dept of Health and Human
Services. Summary of the HIPAA
privacy rule. 2003.
World Health Organization. Guidelines
for the safe transport of infectious
substances and diagnostic specimens.
1997.

specimen(name/title/date)

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Collection and Handling
E. Recommendation fortissue collection andhandling•

Specimen receipt procedure must be available to all personnel in the pathology
department.

•

All specimens must be signed off on the chain of custody form carried by thetransporter and logged into the LIS system of the pathology department foraccessioning.

•

The pathology lab must have a logging system that identifies the person
receiving the specimen, the date and time received.

ii. Tissue Transport
d. Chain of custody
3. Specimen
receipt bylaboratory•(date/time/name)

Laboratory General Checklist, GEN.40115(Specimen Collection Manual Elements –Surgical Pathology and Cytopathology
Specimens)

Laboratory General Checklist, GEN.40900(Specimen Date Received)

The Joint Commission. (2022) 2022
National Patient Safety Goals Hospital
Program.

US Dept of Health and Human
Services. Summary of the HIPAA
privacy rule. 2003.

World Health Organization. Guidelines
for the safe transport of infectious
substances and diagnostic specimens.
1997.

The pathology lab must have a process for documenting who handles theoriginal specimen and all sub-specimens throughout the entire examination,
testing and reporting process.

Association of Surgical Technologists(AST) Recommended Standards ofPractice for Handling and Care ofSurgical Specimens. www.ast.org
Collection and Handling
E. Recommendation for•

A policy and procedure must be made available that identify the process to follow
for labeling discrepancies.

•

In some instances, the specimen can be considered to be a rejection specimen
and only the originator should be making the appropriate labeling changes.

All Common Checklist, COM.06200(Secondary Specimen Container Labeling)•

Label and requisition must be a match. Common mistakes are gender or site.

Laboratory General Checklist, GEN.40492(Specimen Labeling Correction)•

Records of all errors should be maintained.

tissue collection andhandling
ii. Tissue Transporte. Quality Assurance
Monitors
1. Labeling

All Common Checklist, COM.06100(Primary Specimen Container Labeling)

Association of Surgical Technologists(AST) Recommended Standards ofPractice for Handling and Care ofSurgical Specimens. www.ast.org

All Common Checklist, COM.06300(Specimen Rejection Criteria)

discrepancies

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Collection and Handling
E. Recommendation for•

The pathology department must have a policy and procedure that handles
specimen acceptance and rejection.

•

The information on the specimen container must match the information submitted
on the requisition form.

•

Grounds for rejection may include:o Wrong nameo Wrong siteo Wrong identifierso State of specimen

tissue collection andhandling
ii. Tissue Transporte. Quality Assurance
Monitors
2. Specimen

All Common Checklist, COM.06300(Specimen Rejection Criteria)

The Joint Commission. (2022) 2022
National Patient Safety Goals Hospital
Program.
US Dept of Health and Human
Services. Summary of the HIPAA
privacy rule. 2003.
Department of Health and Human
Services, Centers for Medicare andMedicaid Services. Clinical laboratory
improvement amendments of 1988;
final rule. Fed Register. 2003(Jan 24):
[42CFR493.1283(a)(3)]

rejection criteria

World Health Organization. Guidelines
for the safe transport of infectious
substances and diagnostic specimens.
1997.
Association of Surgical Technologists(AST) Recommended Standards ofPractice for Handling and Care ofSurgical Specimens. www.ast.org
Collection and Handling
E. Recommendation fortissue collection andhandling
ii. Tissue Transporte. Quality Assurance
Monitors
3. Tissue•

The specimen collection and handling procedures should include the parameters
for specimens deemed acceptable.
o Identification of the patient sample (labeling)o Completion of the requisition to include all required demographic and clinicaldatao Specimen container to be usedo Type and volume of fixationo Transport packing, temperature, and methodo Additional specialized instructions

All Common Checklist, COM.06300(Specimen Rejection Criteria)International Standard ISO 201664:2020 - Molecular in vitro diagnostic
examinations – Specifications for preexamination processes for formalinfixed and paraffin-embedded (FFPE)
tissue for – Part 4: In situ detection
techniques: section 6 – Inside thelaboratory. The Joint Commission.
(2022) 2022 National Patient Safety
Goals Hospital Program.

Acceptance

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Department of Health and Human
Services, Centers for Medicare andMedicaid Services. Clinical laboratory
improvement amendments of 1988;
final rule. Fed Register. 2003(Jan 24):
[42CFR493.1283(a)(3)]
Association of Surgical Technologists(AST) Recommended Standards ofPractice for Handling and Care ofSurgical Specimens. www.ast.org
Carson F, Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020.
Collection and Handling
E. Recommendation for•tissue collection andhandlingiii. Specimen specificrecommendations

A policy and procedure should be made available that identify the process tofollow for different types of specimens/biopsies:o Muscle - enzyme studies
o Renal/Skin - Immunofluorescenceo Nerve/CNS
o Cardiaco Lymphatic tissue - mercuric fixative; thinner sections, etc.
o Specimens that contain radioactive implants

Anatomic Pathology Checklist, ANP.11670(Specimen- Gross Examination)

Anatomic Pathology Checklist, ANP.11275(Radioactive Material Handling)

1. Specialized

Clinical Laboratory Standards Institute
CLSI MM13-A: Collection, Transport,
Preparation, and Storage of Specimens
for Molecular Methods; Approved
Guideline; 2020.
Carson F, Hladik C. Histotechnology A
Self-Instructional Text, 5th ed. Chicago,
IL: ASCP Press 2020
AFIP, Laboratory Methods in
Histotechnology.

biopsies
Collection and Handling
E. Recommendation for•tissue collection andhandlingiii. Specimen specificrecommendations•

Health care facility policy and procedure should be followed for the propercollection and handling of general biopsies. Procedures to include:
o Type of collection containero Type and volume of fixativeo Transport and holding instructions
All fresh biopsies not needing special handling are to be submitted to thepathology department immediately for processing.

Laboratory General Checklist, GEN.40115(Specimen Collection Manual Elements –Surgical Pathology and Cytopathology
Specimens)

Carson F, Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020.
Bancroft J, Gamble M. Theory andPractice of Histological Techniques, 6thed. New York, NY: Churchill Livingston;
2008.

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2. General biopsies••

Collection and HandlingIf this cannot be completed in a timely manner, the biopsy should be placed in asterile container and kept moist with sterile saline or wrapped in salinedampened sponges until the biopsy can be properly placed in fixative.
Specimens must be placed in appropriate fixative as specified incollection/handling and submission procedure.

•

Health care facility policy and procedure should be followed for the propercollection and handling of bone marrow cores and aspirates.

•

Bone marrow cores/aspirates should be placed in fixative immediately after theprocedure.

iii. Specimen specific•Bone marrow cores/aspirates should be stored at room temperature.

recommendations•

Cores/aspirates must be received in the laboratory, as soon as possible, for
immediate handling according to written protocols.

•

Health care facility policy and procedure should be followed for the propercollection and handling of specimens. Procedures to include:
o Type of collection containero Type and volume of fixative or no fixativeo Transport and holding instructions•

All fresh specimens are to be submitted to the pathology department immediately
with instructions for special testing or processes.

E. Recommendation fortissue collection andhandling

3. Bone marrows

Collection and Handling
E. Recommendation fortissue collection andhandlingiii. Specimen specificrecommendations
4. Large specimen(s)The Joint Commission. (2022). 2022
National Patient Safety Goals Hospital
Program.

•

Large specimens require a longer amount of time for tissue to be properly fixed(e.g. uterus, spleen, lung, liver, etc.).

•

Breast tissue must follow the ASCO guidelines for strict fixation timing andprocessing. (please see section D, iv, e for specific fixation times).

Makary MA, Epstein J, Pronovost PJ,
Millman EA, Hartmann EC, Freischlag
JA. Surgical specimen identification
errors: A new measure of quality insurgical care. Surgery. 2007.141:450455.

Laboratory General Checklist, GEN.40115(Specimen Collection Manual Elements –Surgical Pathology and Cytopathology
Specimens)

Carson F, Hladik C. Histotechnology A
Self-Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020.

Foucar, KM, Bone Marrow Pathology.
2nd ed. Chicago, IL, ASCP Press: 2001.

Laboratory General Checklist, GEN.40115(Specimen Collection Manual Elements –Surgical Pathology and Cytopathology
Specimens)

Anatomic Pathology Checklist, ANP.22983(Fixation – HER2 and ER Breast Cancer
Predictive Marker Testing)

American Society of Clinical Oncology.
(2013). ASCO Guidelines. Retrieved
December 18, 2013, from American
Society of Clinical Oncology (ASCO):
http://www.asco.org/Guidelines/
Lester, SC. Manual of Surgical
Pathology. 3rd ed. Saunders: 2010.

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•

HANDLING PRIOR TO
GROSSGuideline Section
Collection and Handling
F. Accessioning

CAP Checklist

Reference

All Common Checklist, COM.06100(Primary Specimen Container Labeling)

Clinical Laboratory Standards Institute
CLSI - GP33A, Accuracy in Patient andSample Identification; 2019:Vol 30 No7.

HANDLING PRIOR TO GROSSStatement••i. Specimen
Identifiers andLabelling

Anatomic Pathology Checklist, ANP.11250(Adequate Storage)

Wolff AC, Hammond ME, Allison
KH, Harvey BE, et al. Human Epidermal
Growth Factor Receptor 2 Testing in
Breast Cancer: American Society ofClinical Oncology/College of American
Pathologists Clinical Practice Guideline
Focused Update. Arch Path Lab Med.
Nov 2018, Vol. 142, No. 11. pp. 13641382.

Placentas should be refrigerated until delivery to the pathology department.

•

Specimen must be identified/labeled following parameters identified in section B.
Each specimen container received must be compared to the requisition to ensure
correct match of at least 2 patient-specific identifiers:o Full patient nameo Assigned identification number (e.g. health record / master index number)o Date of Birth
Additional requisition information to be checked:o Number of specimen containers
o Type of specimens submitted
o Complete clinical historyo Name of requesting physician to return report too Collection data related to fixation (section D)

All Common Checklist, COM.06200(Secondary Specimen Container Labeling)
Laboratory General Checklist, GEN.40490(Patient Identification)

International Standard ISO 15189:2012- Medical Laboratories; section 5.4 Pre-examination Processes
Zarbo RJ, Tuthill JM, D’Angelo R, et al.
The Henry Ford Production System:
reduction of surgical pathology inprocess misidentification defects by bar
code-specified work process
standardization. Am J Clin Pathol. 2009;
131:469-477.

Collection and Handling
F. Accessioning
ii. Accessioning ordera. Avoiding Error•

It is good laboratory practice to avoid accessioning like-specimens back-to-back.

•If like specimens must be accessioned in sequence it is suggested to separate
by size (e.g. skin punch biopsy followed by skin excision followed by skin punch

All Common Checklist, COM.06100(Primary Specimen Container Labeling)

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biopsy) or to be identified by use of multi colored inks (punch one black ink,
punch two is green ink, punch three blue ink etc.).

All Common Checklist, COM.06200(Secondary Specimen Container Labeling)
Laboratory General Checklist, GEN.40115(Specimen Collection Manual Elements –Surgical Pathology and Cytopathology
Specimens)Collection and Handling
G. Handling prior to•

Gross Examination

There should be sufficient space available in the surgical pathology suite to store
surgical specimens in an orderly fashion after accessioning, and prior to grossexamination:o Space for the containers and accompanying paperwork/request slips.
oStorage area should be clean, free of clutter, and well ventilated.

Laboratory General Checklist, GEN.60000(Adequate Space)
Laboratory General Checklist, GEN.60100(Adequate Space)
Anatomic Pathology Checklist, ANP.11250(Adequate Storage)

Collection and Handling
G. Handling prior to•

Gross Examination
i. Immediate Gross

Site specific documentation on how to handle specimens requiring immediategross examination (i.e. microbiological cultures, electron microscopy,
cytogenetics, flow cytometry, or other special studies) must be available to all
staff handling the specimens and should include:

Examination andoSpecialized grossing techniques (i.e. sterile procedures)

HandlingoSample collection for submission into specialized media (i.e. cytogenetic orEM)oRequisition completion for further testing (i.e. microbiology or pathology
referral lab)oLabeling procedure for sub-specimensoHolding and transport instructions for specialized testing (i.e. refrigerate)oSpecimen cross contamination•

Specimens submitted fresh for immediate gross examination (i.e., frozen
sections, margin determination, etc.) should be kept in their labeledcontainers at room temperature.

Anatomic Pathology Checklist, ANP.11670(Specimen Gross Examination)
Anatomic Pathology Checklist, ANP.11600(Gross Examination – Qualifications)Anatomic Pathology Checklist, ANP.11605(Gross Examination – Supervision)

Department of Health and Human
Services, Centers for Medicare andMedicaid Services. Clinical laboratory
improvement amendments of 1988;
final rule. Fed Register. 1992(Feb28):7183 [42CFR493.1489(b)(6)]

Anatomic Pathology Checklist, ANP.11680(Cross Contamination – Grossing)Anatomic Pathology Checklist, ANP.11810(Intra-operative Preparation Quality)
Anatomic Pathology Checklist, ANP.11670(Specimen - Gross Examination)
All Common Checklist, COM.06100(Primary Specimen Container Labeling)

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•
•

If there is a delay, the fresh specimen should be kept in its labeled container
and refrigerated until it can be examined.
Written procedure to prevent cross contamination.

All Common Checklist, COM.06200(Secondary Specimen Container Labeling)
Anatomic Pathology Checklist, ANP.11250(Adequate Storage)
Anatomic Pathology Checklist, ANP.21397(Cross Contamination – Histology)

Collection and Handling
G. Handling prior to•

Gross Examination

Specimens in fixative requiring gross examination should be assembled/stored in
an orderly fashion after accessioning, with appropriate paperwork/request slips
and labeled cassettes available.

ii. Delayed time to
Gross Examination•

The containers should be sealed to avoid spillage, loss of fixative, loss of
specimen, and to prevent drying of the specimen prior to gross examination.

Anatomic Pathology Checklist, ANP.11600(Gross Examination – Qualifications)Anatomic Pathology Checklist, ANP.11605(Gross Examination – Supervision)

Laboratory General Checklist, GEN.40125(Handling of Referred Specimens)Collection and Handling
G. Handling prior to•

An appropriate room temperature should be maintained, so that specimens areneither frozen nor damaged by excessive heat.

Laboratory General Checklist, GEN.61300(Climate Control)•

Appropriate ventilation should be maintained so that there is adequate air
movement around the specimen containers, without buildup of fixative or othernoxious vapors.

Laboratory General Checklist, GEN.76720(Formaldehyde and Xylene Safety)

Gross Examination
ii. Delayed time to
Gross Examination
a. Monitoring ofEnvironmental
Parameters
Collection and Handling
G. Handling prior to
Gross Examination
ii. Delayed time to•

Adequate fixative should be added to the specimen container as soon as
possible. If insufficient fixative is present when the specimen is received in thelaboratory additional fixative should be added.

Laboratory General Checklist, GEN.40125(Handling of Referred Specimens)

Carson F, Hladik-Cappellano C.
Histotechnology A Self-Instructional
Text, 5th ed. Chicago, IL: ASCP Press
2020.

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Gross Examinationb. Addition of
fixative to
specimen(s)•Brown RW. et. al., Histologic
Preparations Common Problems andTheir Solutions. College of American
Pathologists, 2009

Generally, this should be a volume such that there is a 15-20:1 ratio of fixative totissue specimen. If a large specimen (i.e. uterus, colon, breast, etc.) is
submitted, the specimen should be opened or regularly sliced and covered or
wrapped in an absorptive material (i.e. paper towels, etc.) to maximize surface
exposure to fixative reagents.

•

The specimen container should remain sealed so that drying or other specimen
damage cannot occur.

•

Health care facility policy and procedure should be followed for the propercollection and handling of specimens for intra-operative consultation.
Procedures to include:o Gross examination only.
o Frozen sections
o Touch preps, scrap preps•

All intra-operative consultation results and tissue diagnoses are made and signed
by a pathologist.

Bancroft J, Gamble M. Theory andPractice of Histological Techniques, 6thed. New York, NY: Churchill Livingston;
2008.

Collection and Handling
H. Intra-Operative
Consultation(i.e., Frozen Sections)•

Reagents and slides used for intra-operative consultation are properly labeled.

Anatomic Pathology Checklist, ANP.11670(Specimen – Gross Examination)

Anatomic Pathology Checklist, ANP.11850(Intra-Operative Results)
Anatomic Pathology Checklist, ANP.11660(Surgical Tissue Diagnosis)

Anatomic Pathology Checklist, ANP.11756(Reagents)

Cibull ML. Q&A. Northfield, IL: Collegeof American Pathologists CAP Today.
1997;11(7):112

All Common Checklist, COM.06100(Primary Specimen Container Labeling)
All Common Checklist, COM.06200(Secondary Specimen Container Labeling)•

Intra-operative consultation preparations are adequate for diagnosis.

Anatomic Pathology Checklist, ANP.11810(Intra-operative Preparation Quality)

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•

•

Intra-operative slides are retained and made part of the permanent case.

Residual tissue(s) used for intra-operative examination are processed into
paraffin for comparison with the frozen section interpretation.

Anatomic Pathology Checklist, ANP.12050(Intra-operative Slide Handling)

Zhai Q, Siegal G, eds. College ofAmerican Pathologists. Quality
Management in Anatomic Pathology
Pub 125. Northfield, IL: CAP, 2017.

Anatomic Pathology Checklist, ANP.12075(Residual Frozen Tissue After Frozen
Section Examination)

Rickert RR. Quality assurance goals insurgical pathology. Arch Pathol Lab
Med. 1990;114:1157-1162.

Anatomic Pathology Checklist, ANP.12500(Record and Material Retention – Surgical
Pathology)

Association of Directors of Anatomic
and Surgical Pathology.
Recommendations on quality control
and quality assurance in anatomic
pathology. Am J Surg Pathol.
1991;15:1007-1009.
Gephardt GN, et al. Interinstitutional
comparison of frozen section
consultations. A College of American
Pathologists Q-probes study of 90 538
cases in 461 institutions. Arch Pathol
Lab Med. 1996;120:804-809.
Novis DA, et al. Interinstitutional
comparison of frozen section
consultation in small hospitals. Arch
Pathol Lab Med. 1996;120:1087-1093.

Zhai, Q, Siegal, G, eds. College ofAmerican Pathologists. Quality
Management in Anatomic Pathology
Pub 125. Northfield, IL: CAP, 2017

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Collection and Handling
H. Intra-Operative

When giving a verbal report, the pathologist must be able to speak directly withintra-operative medical/surgical personnel.

•

The patient's identification is checked and confirmed before delivery of any verbal
report.

•All intra-operative consultation reports are made a part of the final surgicalpathology report.

Anatomic Pathology Checklist, ANP.12000(Final Report)•

There is a documented procedure for the routine decontamination of the cryostat
at defined intervals.

Anatomic Pathology Checklist, ANP.23410(Cryostat Decontamination)•

Decontamination of the cryostat is documented, and records are available for
examination.

Clinical Laboratory Standards Institute
CLSI. Protection of Laboratory Workers
from Occupational Acquired Infections,
Approved Guideline M29-A4;
2014;Vol34 No8.
http://www.epa.gov/oppad001/list_b_tub
erculocide.pdf

Laboratory General Checklist, GEN.77800(Hazardous Chemical Waste Disposal)

Lott RL. HQIP: H&E Staining. HQIP - A
Final Critique. Chicago, IL: College ofAmerican Pathologists; 2010.

Consultation
i. Reporting

Anatomic Pathology Checklist, ANP.11900(Verbal Reports)•

Anatomic Pathology Checklist, ANP.11950(Verbal Report/Patient ID)

Collection and Handling
H. Intra-Operative
Consultation
ii. Cryostat
decontamination

Collection and Handling
H. Intra-Operative•

Establish operation procedures for H&E staining:

ConsultationoReagents to be used – concentration and volumesiii. Hematoxylin andoStaining schedule for each staining program

Eosin stain (H&E)

oRotation or change schedule for the reagents

Stain

o

Disposal and or recycle process for reagents•

Establish quality assurance criteria for the staining and evaluation of H&E
staining.

Anatomic Pathology Checklist, Quality
Control, ANP.11756(Reagents)
All Common Checklist, COM.30400(Reagent Expiration Date - Nonwaived
Tests)
Anatomic Pathology Checklist, ANP.11734(Slide Quality)

Brown RW. et. al., Histologic
Preparations Common Problems andTheir Solutions. College of American
Pathologists, 2009.
Carson F, Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020.

Bancroft J, Gamble M. Theory andPractice of Histological Techniques, 6th

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31

ed. New York, NY: Churchill Livingston;2008.
Sheehan DC, Hrapchak BB., Theory
and Practice of Histotechnology, 2nd ed.
Columbus, OH: Battelle Press; 1980.
Horobin RW. Troubleshooting Histology
Stains, 1998, Churchill Livingstone

PART II

II. LABORATORY PROCESSES - Guidelines

Guideline Section

Statement

CAP Checklist

Reference

Anatomic Pathology Checklist, ANP.11250(Adequate Storage)

Clinical Laboratory Standards Institute
CLSI: QMS01-A4: Quality Management
System: A Model for Laboratory
Services; Approved Guideline, 5th ed.
2019.

Laboratory Processes
A. Guidelines•

i. Facility
Requirements

The laboratory has sufficient space and utilities are adequate for grossexamination and specimen storage.

•

Gross examination area has adequate lighting.

•Gross examination area has adequate ventilation system, with policy formonitoring exposure levels to formalin.

•

Laboratory General Checklist, GEN.60150(Adequate Space)
Laboratory General Checklist, GEN.60250(Working Environment)

Laboratory General Checklist, GEN.76720(Formaldehyde and Xylene Safety)

Formalin exposure level of grossing personnel should be examined annually toassure proper ventilation.

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•

Grossing area should have readily available:oPhotographic equipmentoDictation system (unless grossing personnel enters gross dictation directly
into electronic laboratory information system)oAccess to anatomic pathology laboratory information systemoAccess to diagnostic imaging PACS system if located in a clinical hospitalsettingLaboratory Processes
A. Guidelines•

All macroscopic tissue examinations are performed by a pathologist or pathology
resident, or under the supervision of a qualified pathologist.

•

Activities and the nature of supervision is defined in a written protocol.

ii. Personnel

Qualification requirements for non-pathologist or pathology resident personnel who
assist in gross examination of specimens:•

Anatomic Pathology Checklist, ANP.11600(Gross Examination – Qualifications)Anatomic Pathology Checklist, ANP.11605(Gross Examination – Supervision)

Department of Health and Human
Services, Centers for Medicare andMedicaid Services. Clinical Laboratory
Improvement Amendments of 1988;
final rule. Fed Register. 2003(Oct1):1070-1071 [42CFR493.1489], 10711072.

An earned associate degree in laboratory science or medicallaboratory technology, obtained from an accredited institution, OR••

Education/training equivalent to the above that includes at least 60 semester
hours or equivalent from an accredited institution.

This education must include 24 semester hours of medical laboratory
technology courses, OR 24 semester hours of science courses that includes
6 semester hours of chemistry, 6 semester hours of biology, and 12 semester
hours of chemistry, biology or medical laboratory technology in any
combination.

http://www.naacls.org/news/naaclsnews/archives.asp?article_id=599.
Anatomic Pathology Checklist, ANP.11610(Gross Examination Qualifications to Assist
with Grossing)

Department of Health and Human
Services, Centers for Medicare andMedicaid Services. Clinical Laboratory
Improvement Amendments of 1988;
final rule. Fed Register. 2003(Oct1):1070-1071 [42CFR493.1489], 10711072 [42CFR493.1491]

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•

•

In addition, the individual must have laboratory training including either
completion of a clinical laboratory training program approved or accredited by
the NAACLS, ABHES, or other organization approved by HHS (note that thistraining may be included in the 60 semester hours listed above), OR at least 3
months documented laboratory training in each specialty in which theindividual performs high complexity testing.

The laboratory director is responsible in determining whether an individual's
education, training, and experience satisfy the requirements.

Protocols should be in place to specify nature of pathologist supervision of nonpathologist for differing types of specimens.
oProtocol for small simple specimens that do not require knowledge ofanatomy can specify indirect supervision.

oProtocol for more complex specimens can require direct or indirectsupervision based on laboratory director's determination of each grossing
personnel’s ability to properly examine specimen.

•

Pathologist must define in writing the gross activities and the specimen types theindividual is permitted to perform.

•

Performance of non-pathologist who performs gross examination should be
evaluated by a pathologist on a regular basis.
oAnnual review with documentation of errors in grossing, to include specimen
mix-ups, improperly grossed specimens, and other parameters that are felt
to be important by the laboratory director.

Department of Health and Human
Services, Centers for Medicare andMedicaid Services. Clinical Laboratory
Improvement Amendments of 1988;
final rule. Fed Register. 1992(Feb28):7183 [42CFR493.1489(b)(6)]
Cibull ML. Q&A. Northfield, IL: Collegeof American Pathologists CAP Today.
1997;11(7):112

CLIA regulations include exceptions for grandfathered individuals; Refer to
CLIA regulations 42CFR493.1489 and 1491 for details.
••

Anatomic Pathology Checklist, ANP.11670(Specimen– Gross Examination)

Anatomic Pathology Checklist, ANP.11605(Gross Examination – Supervision)

Grzybicki DM, et al. The usefulness ofpathologists' assistants. Am J Clin
Pathol. 1999;112:619-626.
Galvis CO, et al. Pathologists'
assistants practice. A measurement of
performance. Am J Clin Pathol.
2001;116:816-822.
The Joint Commission. Laboratory
Services (CAMLAB) 2012

Anatomic Pathology Checklist, ANP.11640(Competency Assessment of Individuals
Assisting with Grossing)

The Joint Commission. Laboratory
Services (CAMLAB) 2012

Laboratory Processes
A. Guidelines•

Identity of every specimen is maintained at all times during the grossexamination steps.

All Common Checklist, COM.06100(Primary Specimen Container Labeling)

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iii. Specimen Gross
Sectioning•

There are documented instructions or guidelines available for the proper
dissection, description, and histologic sampling of various specimen types (e.g.
gastrointestinal biopsy, mastectomy, colectomy, hysterectomy, renal biopsy,
nerve biopsy, muscle biopsy, etc.).

All Common Checklist, COM.06200(Secondary Specimen Container Labeling)Complex specimens should be dissected, described, and histologicallysampled in a way that:

Anatomic Pathology Checklist, ANP.11670(Specimen– Gross Examination)

o

•



Ensures proper microscopic evaluation and diagnosis can be
performed by the pathologist by following established guidelines for
specimen dissection and histologic sectioning.

All required parameters of CAP Cancer Checklists can be assessedby pathologist.

There are specific policies and procedures for the safe handling, storage, anddisposal of tissues that may contain radioactive material.
Procedures should be developed in conjunction with institutional radiation
safety guidelines and must comply with state regulations for safe handling ofradioactive materials.

Procedures should distinguish policy regarding specimens with low
radioactivity levels (such as sentinel lymph nodes) and high radioactivity
level specimens such as implant devices.



Procedure should specify specific handling details and laboratory should
include specific storage area of higher radioactive material.

Procedure should include institute specific directions for the disposal ofpotentially radioactive tissues.

CAP Cancer Protocols and Checklists.
http://www.cap.org/apps/cap.portal
Barnes CA. False-negative frozen
section results. Am J Clin Pathol.
2000;113(6):900.

Anatomic Pathology Checklist, ANP.11275(Radioactive Material Handling)

Glass EC, et al. Editorial: radiation
safety considerations for sentinel nodetechniques. Ann Surg Oncol. 1999:6:10.
Miner TJ, et al. Guideline for the safeuse of radioactive materials duringlocalization and resection of sentinel
lymph nodes. Ann Surg Oncol.
1999;6:75-82.
Cibull ML. Handling sentinel lymph
node biopsy specimens. A work inprogress. Arch Pathol Lab Med.
1999;123:620-621.

Pfeifer JD. Sentinel lymph node biopsy.
Am J Clin Pathol. 1999;112:599-602.
Fitzgibbons PL, et al.
Recommendations for handling

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•

There is a policy regarding what type of surgical specimens (if any) may be
exempt from submission to the pathology department.



•
•

•

Such a policy should be approved by the medical staff or appropriate
health care committee.
Examples of typical exempt specimens include prosthetic devices,
tonsils, and adenoids in children below a certain age, foreskin inchildren, varicose veins, cataracts, and pannus.

There is a complete list of devices required for tracking under the Safe Medical
Devices Act of 1990.
There is a policy for handling sup-optimal specimens (unlabeled specimens,
specimens unaccompanied by adequate requisition information, left unfixed or
unrefrigerated for extended period of time, received in a container/bag with acontaminated outside surface.

There is written procedure for the storage and disposal of all specimens
submitted for examination. The guideline should include:oTime of retention – minimum of two weeks after report issued and results
reported to the referring physicianoApproved disposal method of fixative as per local and state guidelinesoApproved disposal method of solid waste (tissue)

Anatomic Pathology Checklist, ANP.10016(Surgical Pathology Exclusion)

Anatomic Pathology Checklist, ANP.10032(Surgical Pathology Microscopic Exemptions)

Laboratory General Checklist, GEN.20351(Adverse Patient Event Reporting)

All Common Checklist, COM.06300(Specimen Rejection Criteria)

Anatomic Pathology Checklist, ANP.11550(Specimen Retention – Grossing)radioactive specimens obtained bysentinel lymphadenectomy. Am J Surg
Pathol. 2000;24:1549-1551.

Zarbo RJ, Nakleh RE. Surgicalpathology specimens for grossexamination only and exempt from
submission. A College of American
Pathologists Q-Probes study of current
policies in 413 institutions. Arch Pathol
Lab Med. 1999;123:133-139.
Zhai Q, Siegal G, eds. College ofAmerican Pathologists. Quality
Management in Anatomic Pathology
Pub 125. Northfield, IL: CAP, 2017,113114.

Medical devices; device tracking. Fed
Reg. May 29,119;57:22966-22981

College of American Pathologists.
Policies and guidelines manual.
Surgical specimens to be submitted topathology for examination. Northfield,
IL: CAP, 1999:Appendix M

Zhai Q, Siegal G, eds. College ofAmerican Pathologists. Quality
Management in Anatomic Pathology
Pub 125. Northfield, IL: CAP, 2017,113114.

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Laboratory Processes
A. Guidelines

Document physical parameters of sections submitted for histologic examination:

iv. Tissue Submission••

General informationoSample size must be thin (3-4 mm) enough to ensure adequate fixation andprocessing of the tissue.

oSample must small enough to fit in the cassette and allow space forprocessing fluids to enter the cassette on all sides.

oBloody or friable tissues should be wrapped so that the tissue sample iscontained within the cassette to avoid cross contamination with othersamples.

o

The number of biopsies or cores should be limited to enable proper
embedding, all samples flat and within the same plane.

oNumber of cassettes per sample should be recorded.

oNumber of pieces per cassettes should be recorded.

oSpecialized embedding directions should be documented.

Anatomic Pathology Checklist, ANP.11670(Specimen – Gross Examination)
Anatomic Pathology Checklist, ANP.12155(Gross Description Report Elements)

College of American Pathologists.
Policies and guidelines manual.
Surgical specimens to be submitted topathology for examination. Northfield,
IL: CAP, 1999:Appendix M

Zhai Q, Siegal G, eds. College ofAmerican Pathologists. Quality
Management in Anatomic Pathology
Pub 125. Northfield, IL: CAP, 2017,113114.

Small biopsies
o Multiple small pieces for most small biopsies (e.g. stomach, colon,
endometrium) can be submitted in one cassette. For needle core biopsies,
one or at most a few (less than 5) pieces per cassette.

•

Larger tissue fragments or samples from whole organs
o If more than one section is submitted in a block, the combined sections meet
the above-mentioned parameters and that there is sufficient space between
each piece to allow adequate fixation and embedding.

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Laboratory Processes
B. Tissue cassette•

All tissue cassettes must be identified with a unique identifier.

•

The unique identifier must be indelible throughout all subsequent procedures.

identification•

The unique identifier can be applied manually or electronically through the use of
automated printers.

•

Minimum requirements for a unique identifier include:o Accession case identifier – to include year, subsection type (surgical,
cytology etc.)
o Specimen identifier – alpha or numerico Block identifier – alpha or numeric•

Additional identifiers: to be used but not required:ooLaboratory name or identifier
Color coded cassette: tissue type, fixative used, pathologist etc.

•

Barcodes must not be the only identifying mark; a human readable identifier isalso required.

•If a barcode is applied to the cassette, it should be readable by all tracking
modalities used in the laboratory: LIS, Hospital Information system, associatedtesting equipment (slide writers), and third-party tracking software.

All Common Checklist, COM.06100(Primary Specimen Container Labeling)

International Standard ISO 15189:2012- Medical Laboratories; section 5.4 Pre-examination Processes

All Common Checklist, COM.06200(Secondary Specimen Container Labeling)

Clinical Laboratory Standards Institute
CLSI – LIS02A2 – Specifications forTransferring Information Between
Clinical laboratory Instruments andInformation Systems; 2004.

Laboratory General Checklist, GEN.40825(Specimen ID)

Clinical Laboratory Standards Institute
CLSI – Auto07A – Laboratory
Automation; Data Content for Specimen
Identification; 2004.

All Common Checklist, COM.06200(Secondary Specimen Container Labeling)

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FIXATIONLABORATORY PROCESSES – FIXATION

Guideline Section

Statement

CAP Checklist

Reference

Laboratory General Checklist, GEN.40115(Specimen Collection Manual Elements –Surgical Pathology and Cytopathology
Specimens)

Compton CC, Robb JA, Anderson
MW, Berry AB, et.al. Preanalytics andPrecision Pathology: Pathology Practices
to Ensure Molecular Integrity of Cancer
Patient Biospecimens for Precision
Medicine. Arch Path Lab Med. Nov 2019,Vol. 143, No. 11. pp. 1346-1363.

Laboratory Processes
C. Fixation Parameters
i. Type of fixativea. Formalin, types•

Guidelines for the correct fixative to use for each specimen type should be
documented and include:oFixative to be usedoRecommended duration of fixationoRequired documentation of cold and warm ischemia timesoReferences to mandatory fixation guidelines for breast tissuesoSafety precautions and spill clean-up

Anatomic Pathology Checklist, ANP.22983(Fixation – HER2 and ER Breast Cancer
Predictive Marker Testing)

Clinical Laboratory Standards Institute
CLSI – MM13, Collection, Transport,
Preparation, and Storage of Specimens
for Molecular Methods: 2020.
International Standard ISO 201664:2020 - Molecular in vitro diagnostic
examinations – Specifications for preexamination processes for formalin-fixedand paraffin-embedded (FFPE) tissue for– Part 4: In situ detection techniques:
section 6 – Inside the laboratory.
Allison KH, Hammond EH, Dowsett M,
McKernin SE et al. Estrogen andProgesterone Receptor Testing in Breast
Cancer American Society of ClinicalOncology/College of American
Pathologists Guideline Update. ArchPath Lab Med. Early Online Release.
doi: 10.5858/arpa.2019-0904-SA

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39

Wolff AC, Hammond ME, Allison
KH, Harvey BE, et al. Human Epidermal
Growth Factor Receptor 2 Testing in
Breast Cancer: American Society ofClinical Oncology/College of American
Pathologists Clinical
Practice Guideline Focused Update.
Arch Path Lab Med; Nov 2018, Vol. 142,
No. 11. pp. 1364-1382.
Carson F, Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020.
Lott RL. HQIP: H&E Staining. HQIP - A
Final Critique. Chicago, IL: College ofAmerican Pathologists; 2010.
Bancroft J, Gamble M. Theory andPractice of Histological Techniques, 6thed. New York, NY: Churchill Livingston;
2008.
Laboratory Processes
C. Fixation Parameters
i. Type of fixative
b. Recycling formalin
fixatives•oDocumentation of the initial verification of quality of recycled formalin

Section 19 of Occupational Safety and
Health Act (OSHA) 1970 - Public Law
91-596.

oDocumentation of changes and reverification of quality of recycled formalin
after any procedural changes or repairs to equipment used

29 CFR 1910.1000 (OSHA) Toxic andHazardous SubstancesoWhat formalin can be recycled: from tissue samples or tissue processoroRecycled formalin be used with new tissue samples, samples to be stored,
and on tissue processors

29 CFR 1910.1048 (OSHA)
FormaldehydeoProcedure for recycling formalinoProcedure for testing quality of recycled formalinoProcedure for disposal of non-reusable waste

A written policy and procedure for the use of recycled formalin should include:

29 CFR 1910.1200 (OSHA) Hazard
Communication
29 CFR 1910.1048 (OSHA)
Formaldehyde, Irritant and Potential
Cancer Hazard

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oProcedure for cleaning and maintenance of recycling equipmento

Validation studies comparing the filtered/tested solution to new solution are
required.

oDocumentation to show licensing agencies is required.

29 CFR 1910.1450 (OSHA)
Occupational Exposure to Hazardous
Chemicals in Laboratories
40 CFR 262 (EPA) Standards
Applicable to Generators of Hazardous
Wastes49 CFR 172.101 (DOT) Table ofHazardous Materials and Special
Provisions
http://www.osha.gov/dsg/hazcom/index.
html

Laboratory Processes
C. Fixation Parameters
i. Type of fixativec. Non-Formalin,
types

•

Guidelines for the use of specialized fixatives for each specimen type must be
documented and include:oFixative to be usedoRecommended duration of fixationoSpecialized handling requirements (i.e. refrigeration or flammable storage)oSpecialized preparation or usage (i.e. mix before use)oSafety precautions and spill clean-up

Laboratory General Checklist, GEN.40115(Specimen Collection Manual Elements –Surgical Pathology and Cytopathology
Specimens)

Carson F. Hladik C., Histotechnology A
Self- Instructional Text, 3rd ed. Chicago,
IL: ASCP Press; 2009.
Dapson RW: Glyoxal fixation: How it
works and why it only occasionallyneeds antigen retrieval. Biotech
Histochem. 82:161; 2007.
Bancroft J, Gamble M. Theory andPractice of Histological Techniques, 6thed. New York, NY: Churchill Livingston;
2008.
Michel B, et al., Preservation of tissue
fixed immunoglobulins in skin biopsiesof patients with lupus erythematous andbullous diseases: preliminary report. J
Invest Dermato. 59:449; 1972.
Elias JM, et al, New method forshipment of renal biopsies. J
Histotechnol. 1:15; 1977.

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Laboratory Processes
C. Fixation Parameters•

Using 10% neutral buffered formalin (10% NBF), complete fixation of a 4 mm
thick section of tissue is achieved in approximately 24 hours.

•

As a general recommendation, when using 10% NBF, ALL clinical tissue
specimens should be fixed for a minimum of 6 hours and a maximum of 72 hours.

ii. Fixation
Times/Factors
a. Fixative type•

Anatomic Pathology Checklist, ANP.22300(Specimen Modification)

Anatomic Pathology Checklist, ANP.22300(Specimen Modification)

The general recommendations above are fixative dependent and relatespecifically to the use of 10% NBF. Other fixatives, such as alcoholic formalin or
Bouin, may have different guidelines.

Carson F, Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020.

Wolff AC, Hammond ME, Allison
KH, Harvey BE, et al. Human Epidermal
Growth Factor Receptor 2 Testing in
Breast Cancer: American Society ofClinical Oncology/College of American
Pathologists Clinical
Practice Guideline Focused Update.
Arch Path Lab Med. Nov 2018, Vol.
142, No. 11. pp. 1364-1382.
Goldstein NS, Ferkowicz M, Odish E, etal: Minimum formalin fixation time forconsistent estrogen receptor
immunohistochemical staining ofinvasive breast carcinoma. Am J Clin
Pathol. 120:86–92, 2003.

Laboratory Processes
C. Fixation Parametersii. Fixation
Times/Factors
b. Tissue type•

Guidelines for the fixation and handling of specific tissue types must be
documented based on:oAccepted standards – CAP/ASCO guidelines for breast tissuesoTissue anatomy:


Brain



Fatty tissue – requires extended fixation

Dense tissue such as uterus or cervix- requires extended fixation



Lung – requires inflation

Laboratory General Checklist, GEN.40115(Specimen Collection Manual Elements –Surgical Pathology and Cytopathology
Specimens)

Anatomic Pathology Checklist, ANP.11670(Specimen - Gross Examination)

Wolff AC, Hammond ME, Allison KH,
Harvey BE, et al. Human Epidermal
Growth Factor Receptor 2 Testing in
Breast Cancer: American Society ofClinical Oncology/College of American
Pathologists Clinical Practice Guideline
Focused Update. Arch Path Lab Med.
Nov 2018, Vol. 142, No. 11. pp. 13641382.

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

•

Whole organs
Carson F, Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020.

Dense tissues, such as uterus or cervix, and those that are especially fatty or
bloody, like breast, colon, and spleen, usually require extended times in mostroutine fixatives.

Bancroft J, Gamble M. Theory andPractice of Histological Techniques, 6thed. New York, NY: Churchill Livingston;
2008.
Laboratory Processes
C. Fixation Parameters•ii. Fixation
Times/Factors
c. Tissue Size••

Carson F, Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020.

Gross dissection manual should include information about the size and thicknessof the tissue sample – see section A iv.

A gross dissection manual should include specific instructions related to thefixation of the specimen to include:oTotal fixation time required prior to processingoPreparation of large specimen to improve fixation:
Opening / slicing of whole organs

Exchange fixative

Thickness of tissue specimens is especially important because of its effect onreagent penetration. Large specimens should be opened or regularly sliced tomaximize surface exposure to fixative reagents. Gross tissue sections should beno thicker than 3-4 mm and easily fit between the top and bottom of theprocessing cassette.

Anatomic Pathology Checklist, ANP.11670(Specimen - Gross Examination)

Bancroft J, Gamble M. Theory andPractice of Histological Techniques, 6thed. New York, NY: Churchill Livingston;
2008.

Compton CC, Robb JA, Anderson
MW, Berry AB, et.al. Preanalytics andPrecision Pathology: Pathology Practices
to Ensure Molecular Integrity of Cancer
Patient Biospecimens for Precision
Medicine. Arch Path Lab Med. Nov 2019,Vol. 143, No. 11. pp. 1346-1363.

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43

Clinical Laboratory Standards Institute
CLSI – MM13, Collection, Transport,
Preparation, and Storage of Specimens
for Molecular Methods: 2020.
International Standard ISO 201664:2020 - Molecular in vitro diagnostic
examinations – Specifications for preexamination processes for formalinfixed and paraffin-embedded (FFPE)
tissue for – Part 4: In situ detection
techniques: section 6 – Inside thelaboratory.
Laboratory Processes
C. Fixation Parametersii. Fixation••

Times/Factors
d. Total Fixation time

Guidelines for the total fixation of the specimens should be documented.
Total fixation time required prior to processing to include:oTime from placement in fixative to laboTime large specimen is held prior to final dissectionoTime in cassettes prior to processing – hold time and time on processor•

Tissues for clinical assessment should be placed into an appropriate fixative
immediately after surgical removal. Duration of fixation is an important variable inachieving excellent processing, microtomy, staining, and special staining.

•

Total fixation time should be recorded for each specimen and may be dictatedinto the body of the surgical report.

Anatomic Pathology Checklist, ANP.22983(Fixation – HER2 and ER Breast Cancer
Predictive Marker Testing)

Carson F, Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020.

Anatomic Pathology Checklist, ANP.12155(Gross Description Report Elements)

Wolff AC, Hammond ME, Allison
KH, Harvey BE, et al. Human Epidermal
Growth Factor Receptor 2 Testing in
Breast Cancer: American Society ofClinical Oncology/College of American
Pathologists Clinical
Practice Guideline Focused Update.
Arch Path Lab Med. Nov 2018, Vol. 142,
No. 11. pp. 1364-1382.
Compton CC, Robb JA, Anderson
MW, Berry AB, et.al. Preanalytics andPrecision Pathology: Pathology Practices
to Ensure Molecular Integrity of Cancer
Patient Biospecimens for Precision
Medicine. Arch Path Lab Med. Nov 2019,Vol. 143, No. 11. pp. 1346-1363.

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Revised: September 2023

44

Clinical Laboratory Standards Institute
CLSI – MM13, Collection, Transport,
Preparation, and Storage of Specimens
for Molecular Methods: 2020.
International Standard ISO 201664:2020 - Molecular in vitro diagnostic
examinations – Specifications for preexamination processes for formalin-fixedand paraffin-embedded (FFPE) tissue for– Part 4: In situ detection techniques:
section 6 – Inside the laboratory.
Laboratory Processes
C. Fixation Parameters•ii. Fixation
Times/Factors
e. Environmental

Guidelines for the temperature at which the fixative must be used should be
documented.
o Storage temperature of fixative prior to use
o Temperature the specimen in fixative to be stored at after collectiono Temperature the specimen in fixative to be stored at during transport totesting laboratory.

Parameters• Almost all fixatives are effectively used at room temperature (22-25°C).

1. Temperature• Some fixatives such as acetone are more effective when used cold (4°C).

Laboratory General Checklist, GEN.76720(Formaldehyde and Xylene Safety)

Carson F, Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020.

Bancroft J, Gamble M. Theory andPractice of Histological Techniques, 6thed. New York, NY: Churchill Livingston;
2008.

Laboratory Processes
C. Fixation Parametersii. Fixation
Times/Factors
e. Environmental
Parameters
2. Use ofMicrowaves•

Guidelines for use and operation of specialized microwave equipment used toassist with fixation should include:oSafety instructions to include radiation testing processoWhat solutions can be used in microwaveoType of tissues that can be microwave fixedoSize of tissue that can be microwave fixedoProtocols to be applied

Anatomic Pathology Checklist, ANP.27170(Microwave Usage)

Anatomic Pathology Checklist, ANP.28290(Microwave Monitoring)

Anatomic Pathology Checklist, ANP.28860(Microwave Container Venting)

Carson F, Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020.

Login GR, Giammara B. Rapid
microwave fixation, staining andembedding for light and electron
microscopy. Microscopy Society ofAmerica Workshop; Cincinnati, OH.
1993.

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Anatomic Pathology Checklist, ANP.29430(Microwave Venting)

PROCESSINGLABORATORY PROCESSES – PROCESSING

Guideline Section

Statement

Laboratory Processes•

Procedures must be written and validated for each processing schedule used.

•

Documented processing schedules must include:

D. Processing
i. TimeCAP ChecklistoUnique title that can be related to program on the tissue processoroIdentify what tissue types the schedule can be used foroRush/urgent, biopsies, breast tissue

Indicate any pretreatment of the tissues


Anatomic Pathology Checklist, ANP.23120(Tissue Processing Programs – Validation)
Anatomic Pathology Checklist, ANP.23130(Tissue Processing Programs)

i.e. Tissue must be fully fixed prior to processing as program starts inalcoholoTotal processing timeoSchedule:

Name of reagent

Expiration dateConcentration

Location on processor

Order of application of reagentsEnsure reagents are compatible with each other (i.e. alcohol following
neutral buffered formalin must be 70% or less to stop precipitation ofphosphate salts)

All Common Checklist, COM.30400(Reagent Expiration Date)

ReferenceBancroft JD, Gamble M. Theory andPractice of Histological Techniques.
New York, NY: Churchill Livingstone, 6thed. 2008: 53-92.
Brown RW, et. al., Histologic
Preparations Common Problems andTheir Solutions. College of American
Pathologists, 2009: 4-8.

Carson F. Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020.

Sheehan D, Hrapchak B. Theory andPractice of Histotechnology. Columbus,
OH: Battelle Press, 2nd ed., 1980:59-78.

Llewellyn, B.D., StainsFile,
http://stainsfile.info/StainsFile/prepare/process/auto.htm

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
•



Duration of application

Specialized functions:

Heat – actual temperaturePressure /vacuum – actual levels

Willis, D., Minshew, J., Microwave
Technology in the Histology Laboratory.
Histologic. 2002; 35:1-4.
Login GR, Dvorak AM. The Microwave
Toolbook. A Practical Guide forMicroscopists. Boston, MA: Beth Israel
Hospital; 1994.

Mixing/stirring/agitation – Yes / No

Kok, L.P., Boon, M.E., Microwave
Cookbook of Microscopists. 3rd ed.
Coulomb Press, Leyden, 1992.

Maintenance programs for the processor must be established:ooPreventative maintenance and service contractsCompleted by lab staffCompleted by vendor service

Operational maintenance:

Reagent top up / exchange / rotation schedule based on:•

Number of cassettes processed•

Number of time program run•

Monitored and established by processor softwareEstablish if recycled reagents can be used on processor



Cleaning of reagent reservoir containers

All Common Checklist, COM.30600(Maintenance/Function Checks)
All Common Checklist, COM.30675(Instrument and Equipment Records)

Kok LP, Boon ME. Ultrarapid vacuummicrowave histoprocessing. Histochem
J. 1995;27(5):411-419.

Anatomic Pathology Checklist, ANP.23100(Tissue Processor Solutions)

Clinical Laboratory Standards Institute
CLSI GP31-A Laboratory
Instrumentation, Implementation,
Validation and Maintenance.

Anatomic Pathology Checklist, ANP.23100(Tissue Processor Solutions

Bancroft JD, Gamble M. Theory andPractice of Histological Techniques.

Laboratory Processes
D. Processing•

Establish and document for fixative to be used on the tissue processor:

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ii. Tissue ProcessoroType of fixative to be used

Reagents



10% neural buffered formalin (NBF)i.

Zinc formalin

Alcoholic formalinFormalin substitute or proprietary fixative

Fixativeo

Number of reservoirs of fixative to be usedoDuration of time in fixativeoTemperature / vacuum/ agitationo

Rotation or change schedule•

Verify and document that the fixative used is compatible with the tissues to beprocessed.

•

Establish if recycled fixative can be used on processor.

•

Establish and document procedures for fixative handling that include:oStorageoSafety to include:o

Use of personal protective equipmentSpill control and clean-up

Monitoring of exposure levels

Anatomic Pathology Checklist, ANP.23120(Tissue Processing Programs – Validation

New York, NY: Churchill Livingstone, 6thed. 2008: 53-92.

Anatomic Pathology Checklist, ANP.23130(Tissue Processing Programs)

Brown RW, et. Al., Histologic
Preparations Common Problems andTheir Solutions. College of American
Pathologists, 2009: 4-8.

Carson F, Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020: 31-42.

Sheehan D, Hrapchak B. Theory andPractice of Histotechnology. Columbus,
OH: Battelle Press, 2nd ed., 1980:59-78.

Disposal methods that follow regulatory guidelines

Laboratory Processes
D. Processing•

Develop documentation that establishes the parameters of the dehydrant used on
the tissue processor:

Bancroft JD, Gamble M. Theory andPractice of Histological Techniques.

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ii. Tissue Processor
i.

Reagents fordehydrationoType – alcohol or proprietary productoType of alcohol – ethanol or isopropanolo Concentration – grades alcohols (i.e. 70%, 80%, 95%, 100%)o Number of reservoirs of each alcohol concentration

Anatomic Pathology Checklist, ANP.23100(Tissue Processor Solutions)Anatomic Pathology Checklist, ANP.23120(Tissue Processing Programs – Validation)o Duration of time for each alcohol reservoir and total timeo Temperature / vacuum/ agitationo Rotation or change schedule•

Verify and document that the dehydrant is compatible with the tissues to beprocessed and changed at intervals appropriate for workload.

•

Ensure that dehydrant following fixative is compatible with fixative:

Anatomic Pathology Checklist, ANP.23130(Tissue Processing Programs)

New York, NY: Churchill Livingstone, 6thed. 2008:53-92.

Brown RW, et. Al., Histologic
Preparations Common Problems andTheir Solutions. College of American
Pathologists, 2009:4-8.

Carson F, Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020:31-42.

Sheehan D, Hrapchak B. Theory andPractice of Histotechnology. Columbus,
OH: Battelle Press, 2nd ed., 1980: 5978.

o 10% NBF- the first alcohol in the dehydrating series should be 70% or less toprevent the precipitation of phosphates from the 10% NBF
o Alcoholic formalin – the first alcohol in the dehydrating series can be 95% as
the tissue has already been in 70% alcoholo Formalin substitute or proprietary fixatives – must follow guidelines provided
by the manufacturer•

Validate that the dehydrant is compatible with the reagent that follows in theprocessing cycle; this could be xylene or xylene substitute or paraffin.

•

Develop a documentation process for recording the purchase, use, and disposalof ethanol. Ethanol is strictly controlled by the federal government.

•

Develop procedures for alcohol:
o Storage

Laboratory General Checklist – GEN.76000(Chemical Hygiene Plan)o Safety to include:

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Use of personal protective equipmentSpill control and clean-up

Monitoring of exposure levels

Laboratory General Checklist – GEN.76500(Flammable Storage)o Disposal methods that follow regulatory guidelines
o Recycling procedures:

Testing method to prove quality

What alcohol can be recycledWhen recycled alcohol can be used

Laboratory General Checklist, GEN.77800(Hazardous Chemical Waste Disposal)

Laboratory Processes
D. Processing•

ii. Tissue Processori.

Develop documentation that establishes the parameters of the clearant used on
the tissue processor:oReagents forType – xylene, xylene substitute or proprietary product

clearing

Anatomic Pathology Checklist, ANP.23100(Tissue Processor Solutions)

Verification that clearant is compatible with dehydrants and paraffino Number of reservoirs of clearantAnatomic Pathology Checklist, ANP.23350(Paraffin Baths, Flotation Baths, andEmbedding Stations)o Duration of time for each reservoir of clearant and total timeo Temperature / vacuum/ agitationo Rotation or change schedule••

Laboratory General Checklist, GEN.76000(Chemical Hygiene Plan)o Storageo Safety to include:

Use of personal protective equipment

Brown RW, et. Al., Histologic
Preparations Common Problems andTheir Solutions. College of American
Pathologists, 2009 4-8.

Carson F, Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020:31-42.

Verification that the clearant to be used is compatible with the tissues to beprocessed and changed at intervals appropriate for workload.
Develop procedures for clearant:

Bancroft JD, Gamble M. Theory andPractice of Histological Techniques.
New York, NY: Churchill Livingstone, 6thed. 2008: 53-92.

Sheehan D, Hrapchak B. Theory andPractice of Histotechnology. Columbus,
OH: Battelle Press, 2nd ed., 1980: 5978.

Laboratory General Checklist, GEN.77800(Hazardous Chemical Waste Disposal)

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

Spill control and clean-up

Monitoring of exposure levels

o Disposal methods that follow regulatory guidelines
o Recycling procedures:

Testing method to prove qualityWhen recycled clearant can be used

Laboratory Processes
D. Processing
ii. Tissue Processor
d. Reagents for•Develop documentation that establishes the parameters of the paraffin to be
used on the tissue processor:o Type – with or without additives


infiltrationi.

Paraffin(s)Verification that paraffin is compatible with the dehydrant or clearantusedoMelting point of paraffinoNumber of reservoirs of paraffinoDuration of time for each reservoir of paraffin and total timeoTemperature / vacuum/ agitationo

Rotation or change schedule

Format of wax to be used; melted wax, pellets, solid blockAnatomic Pathology Checklist, ANP.23350(Paraffin Baths, Flotation Baths, andEmbedding Stations)Bancroft JD, Gamble M. Theory andPractice of Histological Techniques.
New York, NY: Churchill Livingstone, 6thed. 2008: 53-92.
Brown RW. Et. Al., Histologic
Preparations Common Problems andTheir Solutions. College of American
Pathologists, 2009: 4-8.

Carson F, Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020: 31-42.
Sheehan D, Hrapchak B. Theory andPractice of Histotechnology. Columbus,
OH: Battelle Press, 2nd ed. 1980:59-78.

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EMBEDDINGLABORATORY PROCESSES – EMBEDDING

Guideline Section

Statement

CAP Checklist

Reference

Laboratory Processes
E. Embedding•i. General
Recommendations••

Develop standardized guidelines for routine embedding and handling of special
biopsies:oOpening of cassettes – one cassette at timeoMold sizeoStorage and temperature of moldsoPlacement of tissue in moldSimilar surfaces in same directionDirection of surface in orientation to block placement on the microtomeoOrientation of the tissue typesoMethod for cooling embedded blocksoMethod for release of blocks from molds and removal of excess paraffinoMethod for cleaning and reuse of molds

Anatomic Pathology Checklist, ANP.23350(Paraffin Baths, Flotation Baths, andEmbedding Stations)

Carson F, Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020.

Bancroft J, Gamble M. Theory andPractice of Histological Techniques, 6thed. New York, NY: Churchill Livingston;
2008.
Luna L. Histopathologic Methods andColor Atlas of Special Stains and tissue
Artifacts; American Histolabs Inc; 1992
(embedding table)

Develop quality assurance procedures:oManual or electronic workload log used to compare recorded number ofcassettes with the actual number of cassettes.

oDocumentation and follow up of discrepancies

Anatomic Pathology Checklist, ANP.21350(Specimen Preparation Records)

Establish guidelines for the order of embedding cassettes:oUrgencyoTissue type; biopsy, routine tissues

Anatomic Pathology Checklist, ANP.23130(Tissue Processing Programs)

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•

Establish guidelines for the use and operation of the embedding center:oTemperature of embedding paraffin – monitored dailyo

Set temperature of other heated elements: holding paraffin, work surface,
and forcepsoCleaning of forceps and work surfacesoAddition of paraffin to reservoir: liquid, pellets, solid blocko

Cleaning of the paraffin reservoir and filter

Anatomic Pathology Checklist, ANP.21397(Cross Contamination – Histology)

Laboratory Processes
E. Embedding•

ii. Paraffin Wax

Establish type of paraffin wax to be used for embedding:oSpecialized paraffin or the same as processing paraffinoAdditives – beeswax, plastic polymers, diethylene glycol distearate, ceresinoMelting point

MICROTOMYLABORATORY PROCESSES - MICROTOMY

Guideline Section

Statement

Anatomic Pathology Checklist, ANP.23350(Paraffin Baths, Flotation Baths, andEmbedding Stations)

Carson F, Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020.
Bancroft J, Gamble M. Theory andPractice of Histological Techniques, 6thed. New York, NY: Churchill Livingston;
2008.

CAP Checklist

Reference

Anatomic Pathology Checklist, ANP.23400(Microtome Maintenance)

Clinical Laboratory Standards Institute
CLSI GP31-A Laboratory
Instrumentation, Implementation,
Validation and Maintenance; 2009.

All Common Checklist, COM.30600(Maintenance/Function Checks)

Carson F, Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020.

Laboratory Processes
F. Microtomyi. Microtome
Maintenance•

Written instructions for the operation of all makes/models of microtomes:oManual vs. automatedoCleaning and maintenanceo

Acceptable cleaning productsoLubrication schedule and reagent

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•

Schedule and document annual preventative maintenance, service, or repair•

Develop technique to standardized position of microtome chuck (block holder) on
all microtomes to ensure blocks can be recut on any microtome.

•

Establish guidelines for the orientation of block placement in microtome chuck:

All Common Checklist, COM.30675(Instrument and Equipment Records)

Bancroft J, Gamble M. Theory andPractice of Histological Techniques, 6thed. New York, NY: Churchill Livingston;
2008.

;Laboratory Processes
F. Microtomy
ii. Section preparation

Clinical Laboratory Standards Institute
CLSI - GP33A, Accuracy in Patient andSample Identification; 2019.

a. Block trimmingo

Block identifier to face to the right, left, up, or down.

All Common Checklist, COM.06100(Primary Specimen Container Labeling)
All Common Checklist, COM.06200(Secondary Specimen Container Labeling)•

Establish cutting guidelines:oPlacement of the slide labeloLimiting one patient tissue to a slideoThickness of section
Routine tissues

Specialized tissues (i.e. brain, lymph nodes)

Specialized techniques (i.e. amyloid, immunohistochemistry)

Carson F, Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020.

Bancroft J, Gamble M. Theory andPractice of Histological Techniques, 6thed. New York, NY: Churchill Livingston;
2008.

All Common Checklist, COM.06100(Primary Specimen Container Labeling)

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TissueThickness

Renal Sections

1 to 3 microns

Routine Paraffin
Bone Marrow

Nerve histochemical staining
Amyloid demonstrationo

4 to 5 microns
2 to 3 microns

6 to 15 microns
6 to 12 microns

Number of sections / ribbons per slideSections/ ribbons are same depthEach section / ribbon is a different depthAmount of trim between each section/ribbonoPlacement of sections on the slideoNumber of slides per tissue type (i.e. 2 slides for biopsy blocks)o

Use of specialized slides:oAnatomic Pathology Checklist, ANP.11716(Paraffin Microtomy)Adhesive or no adhesiveControl slides – specialized markings

Addition of additives to water bathAdhesives (i.e. gelatin, agar, Elmer’s glue or proprietary products)

Surfactants (i.e. tween)

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Carson F, Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020.

Laboratory Processes
F. Microtomy•

Establish guidelines for the use and maintenance of flotation/water bath:iii. Flotation BathoTemperature of flotation/water bath – documentation of temperature

a. TemperatureoType of water to be used – tap vs. distilledo

Use of additives – gelatin, agar, Elmer’s glue, proprietary product(s)oCleaning methodFrequencyCleaning products to be used

Laboratory Processes
F. Microtomy•

iv. Slides
a. Labelling

All slides must be clearly labeled to identify the following:o Specimen accession numbero Block identifiero Slide level number
o Patient nameo Stain identifier•

Establish a labeling procedure to be used; It is good laboratory practice to label
slides only as required and to avoid the practice of pre-labeling large numbers ofslides in advance.

•

Establish a quality assurance process of matching slides against the block before
delivery out of the laboratory.

All Common Checklist, COM.30675(Instrument and Equipment Records)

Anatomic Pathology Checklist, ANP.23350(Paraffin Baths, Flotation Baths, andEmbedding Stations)

All Common Checklist, COM.06100(Primary Specimen Container Labeling)
All Common Checklist, COM.06200(Secondary Specimen Container Labeling)

Anatomic Pathology Checklist, ANP.21397(Cross-Contamination – Histology)

Bancroft J, Gamble M. Theory andPractice of Histological Techniques, 6thed. New York, NY: Churchill Livingston;
2008.

Clinical Laboratory Standards Institute
CLSI - GP33A, Accuracy in Patient andSample Identification; 2019.
Carson F, Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020.
Brown RW, Della Speranza V, Alvarez
JO, et al. Uniform labeling of blocks andslides in surgical pathology: Guidelinefrom the College of American
Pathologists Pathology and Laboratory
Quality Center and the National Societyfor Histotechnology. Arch Pathol Lab
Med. 2015;139(12):1515-24.

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Laboratory Processes
F. Microtomy•

iv. Slides
b. Slide Drying•

Clinical Laboratory Standards Institute
CLSI - GP33A, Accuracy in Patient andSample Identification; 2019.

Drying times for slides with paraffin sections should be established and madeavailable to all technical staff. The following recommendations should beconsidered:o

Air drying of cut sections before placing into the drying oveno

Use of a forced air dryer maintained at a temperature just above the melting
point of the paraffinoDrying time and temperature, commonly slides are dried at 58-60°C for 1530 minutes

Carson F, Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020.

Special techniques, such as immunohistochemistry or in-situ hybridization may
require longer drying times. The required drying time should be included in thewritten procedure.

•

Dry slides in an oven for a minimum of 60 minutes at a temperature between 5060°C. Optimal results are achieved at room temperature for 24 hours; however,this is impractical in a clinical laboratory setting (Note: Some molecular testing
protocols require that slides not be oven dried).

•

Guidelines to be established for the retention and disposal of all glass paraffinblocks and slides.

Clinical Laboratory Standards Institute
CLSI – I/L28-A2, Quality Assurance for
Design Control and Implementation of
Immunohistochemistry Assays, 2011.

Laboratory Processes
F. Microtomy
iv. Slides
c. Disposal of
Blocks/Slides

Anatomic Pathology Checklist, ANP.12500(Record and Material Retention – Surgical
Pathology)

Clinical Laboratory Standards Institute
CLSI – GP05-A3 Clinical Laboratory
Waste Management; 2011.

Anatomic Pathology Checklist, ANP.27150(Glass Slide/Block Disposal)

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STAININGLABORATORY PROCESSES – STAINING

Guideline Section

Statement

CAP Checklist

Reference

All Common Checklist, COM.10000(Policy and Procedure Manual)

Clinical Laboratory Standards Institute
CLSI GP31-A Laboratory
Instrumentation, Implementation,
Validation and Maintenance; 2009.

Laboratory Processes
G. Staining•

i. Hematoxylin & Eosin(H&E)

Establish operation procedure for manual or automated staining:oReagents to be used – concentration and volumesoStaining schedule for each specific staining programoRotation or change schedule for the reagentso

Disposal and or recycle process for reagents•

Establish quality assurance criteria for the staining and evaluation of hematoxylin
and Eosin stain.

•

HEMATOXYLIN: When applied correctly, in well-fixed, well-processed tissues,
epithelial cells will demonstrate:o

A well-defined nuclear membraneoClear, open (vesicular) karyoplasm (cytoplasm of the nucleus)oCrisp, fine-spiculed chromatin patternsAlso, in most tissue sections, there are some dense closed(hyperchromatic) nuclear patterns present in lymphoid tissue.

oProminent “eosinophilic” nucleoli (if present)oCartilage and calcium deposits stain dark blueo

The hematoxylin should appear blue to blue-black

Laboratory General Checklist, GEN.77800(Hazardous Chemical Waste Disposal)

Anatomic Pathology Checklist, ANP.10042(Histologic Prep Quality)

Anatomic Pathology Checklist, ANP.10038(Tissue Sample Quality)

Laboratory General Checklist, GEN.30000(Monitoring Analytic Performance)

Anatomic Pathology Checklist, ANP.21395(Special Stains/Studies)

Anatomic Pathology Checklist, ANP.11734(Slide Quality)

Lott RL. HQIP: H&E Staining. HQIP - A
Final Critique. Chicago, IL: College ofAmerican Pathologists; 2010.

Brown RW. et. al., Histologic
Preparations Common Problems andTheir Solutions. College of American
Pathologists, 2009.

Carson F, Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020.

Bancroft J, Gamble M. Theory andPractice of Histological Techniques, 6thed. New York, NY: Churchill Livingston;
2008.

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•

EOSIN: When applied correctly, in well-fixed, well processed tissue, eosin
produces, at least, a “tri-tonal” (three-color) effect.
oMuscle cells (smooth, skeletal, cardiac) and epithelial cell cytoplasm will
stain deep red-pink.

oCollagen will stain a distinct lighter pink.

o

Red blood cells (RBC) will stain a bright orange-red.

oNucleoli (if present) should exhibit a reddish-purple color due to their highprotein and RNA content.



It is essential, when applying eosin, that the smooth muscle/cell cytoplasm andcollagen be differentially stained (different shades of red/pink).

•

Complete and document results of a H&E control prior to staining routine
workload.
o

All Common Checklist, COM.30675(Instrument and Equipment Records)

Anatomic Pathology Checklist, ANP.21360(Automated Stainer)

Prophet EB, Mills B, Arrington JB, Sobin
LH. AFIP Laboratory Methods in
Histotechnology, AFIP; 1992.

Sheehan DC, Hrapchak BB. Theory andPractice of Histotechnology, 2nd ed.
Columbus, OH: Battelle Press; 1980.

Horobin RW. Troubleshooting Histology
Stains, Churchill Livingstone; 1998.

Documentation to include changes or actions taken to correct substandard
staining of the control.

•

Establish a preventative maintenance program that includes annual service andemergency service.

•

Establish written procedures for manual or automated staining procedures toinclude:Laboratory Processes
G. Staining
ii. Histochemical andenzymatic stains(special stains)oSpecial cutting or preparation of tissue sectionoReagents used


Access to material data sheetsConcentration



Storage

DisposaloSpecific steps of staining procedureoQuality assurance process

All Common Checklist, COM.10000(Policy and Procedure Manual)
Laboratory General Checklist, GEN.76100(Chemical Safety Document Access)

Anatomic Pathology Checklist, ANP.21395(Special Stains/Studies)

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•

•

•

•



Define positive control tissue

Define expected stain results

Records of acceptability

Establish operation procedures for automated staining equipment:oValidation processoCleaning and maintenance procedures

Establish a preventative maintenance program that includes annual service andemergency service.

Histochemical stains, or special stains, refer to a group of secondary stains usedin conjunction with H&E staining. They were developed to provide differential
coloration and contrast to cell and tissue constituents with the goal ofunderstanding cell structure and function.

Many are used to identify morphological entities such as bacteria, fungi, nerve
fibers, and for connective tissues including collagen and reticular fibers.

•

Other special histochemical stains are used for specific tissue components andinclude stains for iron, mucins, glycogen, amyloid, and nucleic acids.

•

Enzyme histochemical staining refers to a subclass of histochemistry that
identifies enzymes by employing substrates containing one of a number ofvarious naphthol compounds.

Laboratory General Checklist, GEN.77800(Hazardous Chemical Waste Disposal)

Bancroft J, Gamble M. Theory andPractice of Histological Techniques, 6thed. New York, NY: Churchill Livingston;
2008.

Anatomic Pathology Checklist, ANP.23100(Tissue Processor Solutions)

Carson F, Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020.

Anatomic Pathology Checklist, ANP.23120(Tissue Processing Programs – Validation)

Anatomic Pathology Checklist, ANP.23130(Tissue Processing Programs)

All Common Checklist, COM.30550(Instrument/Equipment Performance
Verification)

All Common Checklist, COM.30600(Maintenance/Function Checks)

Sheehan DC, Hrapchak BB. Theory andPractice of Histotechnology, 2nd ed.
Columbus, OH: Battelle Press; 1980.
Kiernan J. Histological andHistochemical Methods: Theory andPractice 4th ed. Oxfordshire, England;
2008.
Pearse AGE, Stoward PJ.
Histochemistry, Theoretical andApplied, 4th ed. Vol. 2. Analytical
Technique. Edinburgh: ChurchillLivingstone, 1985.
Lillie RD, Fullmer HM. Histopathologic
Technic and Practical Histochemistry.
4th ed. New York: McGraw-Hill; 1976.

All Common Checklist, COM.30675(Instrument and Equipment Records)

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Laboratory Processes
G. Stainingiii.
Immunohistochemical
stains•Establish a procedure for selection and development of antibodies and clones tobe added to menu:oFixation of tissueoCutting of tissue section

Paraffin

Frozeno

Selection and validation of antibody and cloneoSelection, validation, and monitoring of reagentsoValidation of application methodoPretreatment

Antibody dilutionRetrieval method – if required

Detection method
•

DAB

•

Alkaline phosphatase•

FluorescentDocumentation of scoring methodology

Manual or automatedoDocumentation of validation; record test tissue, expected results actual
results and changes to methodoStorage of antibody and reagents

Anatomic Pathology Checklist, ANP.22983(Fixation - HER2 and ER Breast Cancer
Predictive Marker Testing)

Clinical Laboratory Standards Institute
CLSI: ILA28-A2: Quality Assurance for
Design Control and Implementation of
Immunohistochemistry Assays;
Approved Guideline: 2011.

Anatomic Pathology Checklist, ANP.22300(Specimen Modification)

Anatomic Pathology Checklist, ANP.22500(Buffer pH)

Anatomic Pathology Checklist, ANP.22750(Antibody Validation/Verification – NonPredictive Marker)

Anatomic Pathology Checklist, ANP.22978(Predictive Marker Testing –Validation/Verification)

Anatomic Pathology Checklist, ANP.22969(Report Elements)

Fitzgibbons PL, Bradley LA, Fatheree
LA, Alsabeh R, et.al. Principles ofAnalytic Validation of
Immunohistochemical Assays:
Guideline From the College of American
Pathologists Pathology and Laboratory
Quality Center. Arch Path Lab Med;Nov 2014, Vol. 138, No. 11. pp. 14321443.
Troxell ML, Fulton RS, Swanson
PE, Bellizzi AM, Fitzgibbons PL, et.al.
Predictive Markers Require ThoroughAnalytic Validation. Arch Path Lab
Med; Aug 2019, Vol. 143, No. 8. pp.
907-909.
Torlakovic EE. How Validate Predictive
Immunohistochemistry Testing in
Pathology? Arch Path Lab Med; Aug
2019, Vol. 143, No. 8. pp. 907-907.
Validation doc

All Common Checklist, COM.30350(Reagent Storage and Handling –Nonwaived Tests)

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Anatomic Pathology Checklist, ANP.22615(Endogenous Biotin)

Anatomic Pathology Checklist, ANP.22900(Slide Quality)
Anatomic Pathology Checklist, ANP.22760(New Reagent Lot Confirmation of
Acceptability)•

Establish re-validation procedures after change of:oMethodology

oReagentoAntibody

o



Clone



Lot number

Dilution

Equipment

New modelmajor service repair

move or relocation

Anatomic Pathology Checklist, ANP.22780(IHC Assay Performance)

All Common Checklist, COM.30550(Instrument/Equipment Performance
Verification)

Bancroft J, Gamble M. Theory andPractice of Histological Techniques, 6thed. New York, NY: Churchill Livingston;
2008.
Dabbs D. Diagnostic
Immunohistochemistry: Theranostic andGenomic Applications, Expert Consult:
Online and Print , 3rd ed.

Fitzgibbons PL, Bradley LA, Fatheree
LA, Alsabeh R, et.al. Principles ofAnalytic Validation of
Immunohistochemical Assays:
Guideline From the College of American
Pathologists Pathology and Laboratory
Quality Center. Arch Path Lab Med;Nov 2014, Vol. 138, No. 11. pp. 14321443.

Taylor, Cote; Immunomicroscopy
Volume 19 in Major Problems in
Pathology Series, 3rd ed.
Hayat MA. Microscopy,
Immunohistochemistry and Antigen
Retrieval Methods: For Light andElectron Microscopy, Springer Press;
2002.
Elias JM. Immunohistopathology: A
Practical Approach to Diagnosis; 2nd ed.
Chicago, IL: ASCP Press, 2003.

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•

Establish procedures for cleaning and maintenance of equipment:oCalibration of pipettesoMonitoring of refrigerator and freezer temperature


o

o
•

All Common Checklist, COM.30820(Quantitative Pipette Accuracy andReproducibility)

NIST calibration procedure

Ancillary equipment


Microwave ovenSteamer

All Common Checklist, COM.30750(Temperature Checks)

Stainer

Establish a preventative maintenance program that includes annual service andemergency service.

All Common Checklists, COM.30600(Maintenance/Function Checks)

All Common Checklist, COM.30675(Instrument and Equipment Records)•

Establish procedure for the disposal of reagents as per local, state and national
requirements.

•

Immunohistochemistry (IHC) staining refers to the method of localizing specificantigens (e.g. proteins) in cells of a tissue by the principle of an antibody /antigen recognition. This reaction is labelled by a detection technique andvisualized by a chromagen.

•Establish Quality Control and Quality Assurance procedures to include:

Laboratory General Checklist, GEN.77800(Hazardous Chemical Waste Disposal)

Hayat MA. Immunogold-Silver Staining:
Principles, Methods, and Applications,
CRC; 1995.
Javois LC. Immunocytochemical
Methods and Protocols, 3rd ed. :BIOS
Scientific; 2003.
Polack JM. Introduction to
Immunocytochemistry, 3rd ed. :BIOS
Scientific; 2003.
Hayat MA. Microscopy,
Immunohistochemistry and Antigen
Retrieval Methods: For Light andElectron Microscopy, Springer Press;
2002.
Javois LC. Immunocytochemical
Methods and Protocols, 3rd ed. :BIOS
Scientific; 2003.
Shi S, Taylor CR. Antigen Retrieval
Techniques: Immunohistochemistry andMolecular Morphology, Eaton
Publications; 2000.
Immunochemical Staining Methods
Handbook, 3rd ed., Dako Corp,
Carpinteria, CA.

Clinical Laboratory Standards Institute
CLSI – I/L28-A2, Quality Assurance for
Design Control and Implementation of
Immunohistochemistry Assays, 2011.

Laboratory Processes
G. Staining
iv.

Anatomic Pathology Checklist, ANP.21395(Special Stains/Studies)

Fitzgibbons PL, Bradley LA, Fatheree
LA, Alsabeh R, et.al. Principles ofAnalytic Validation of

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ImmunohistochemicaloSelection of appropriate control material

Stainso

Validation of control materiala. Quality ControloDocumentation of test of control at accredited lab

Use and application of controls

Patient and antibody reagent control



Positive and negative

Anatomic Pathology Checklist, ANP.21850(QC – Immunofluorescence)
Anatomic Pathology Checklist, ANP.22550(QC – Antibodies)

Anatomic Pathology Checklist, ANP.22570(QC – Antibodies)•

Establish procedures for the review of controls and release of patient slides forinterpretation.

•

Records of review need to be retained.

•

IHC quality control measures are essential to provide and ensure consistency of
performance and reproducibility of the intended target.

Anatomic Pathology Checklist, ANP.22660(Control Slide Review)

Anatomic Pathology Checklist, ANP.22780(IHC Assay Performance)
Laboratory General Checklist, GEN.30000(Monitoring Analytic Performance)

Immunohistochemical Assays:
Guideline From the College of American
Pathologists Pathology and Laboratory
Quality Center. Arch Path Lab Med.
Nov 2014, Vol. 138, No. 11. pp. 14321443.
Bancroft J, Gamble M. Theory andPractice of Histological Techniques, 6thed. New York, NY: Churchill Livingston;
2008.
Dabbs D. Diagnostic
Immunohistochemistry: Theranostic andGenomic Applications, Expert Consult:
Online and Print, 3rd ed.
Taylor C, Cote RJ. Immunomicroscopy.
Volume 19 in Major Problems in
Pathology Series, 3rd ed.

Hayat MA. Microscopy,
Immunohistochemistry and Antigen
Retrieval Methods: For Light andElectron Microscopy, Springer Press;
2002.
Elias JM. Immunohistopathology: A
Practical Approach to Diagnosis; 2nd ed.
Chicago, IL: ASCP Press; 2003.
Taylor C, Cote RJ. Immunomicroscopy:
A Diagnostic Tool for the Surgical
Pathologist, 3rd ed., WB Saunders;
2005.
Immunochemical Staining Methods
Handbook, 3rd ed., Dako Corp,
Carpinteria, CA.

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Laboratory Processes
G. Staining•

Establish procedure for clinical validation of each antibody:iv.

oNumber of tissue sections to be tested per antibody

ImmunohistochemicaloComparison of results to previous stained slides or duplicate slides stainedby accredited lab

stainsb. Intended Use of
the Antibody

Anatomic Pathology Checklist, ANP.22750(Antibody Validation/Verification – NonPredictive Marker)
Anatomic Pathology Checklist, ANP.22760(New Reagent Lot Confirmation of
Acceptability)

Anatomic Pathology Checklist, ANP.22550(QC – Antibodies)•

Each antibody MUST be clinically validated to be relevant to its intended target

antigen.

Anatomic Pathology Checklist, ANP.22570(QC – Antibodies)

Anatomic Pathology Checklist, ANP.22978(Predictive Marker Testing – Validation/
Verification)

Clinical Laboratory Standards Institute
CLSI – I/L28-A2, Quality Assurance for
Design Control and Implementation of
Immunohistochemistry Assays: 2011.
Fitzgibbons PL, Bradley LA, Fatheree
LA, Alsabeh R, et.al. Principles ofAnalytic Validation of
Immunohistochemical Assays:
Guideline From the College of American
Pathologists Pathology and Laboratory
Quality Center. Arch Path Lab Med.
Nov 2014, Vol. 138, No. 11. pp. 14321443.
Troxell ML, Fulton RS, Swanson
PE, Bellizzi AM, Fitzgibbons PL, et.al.
Predictive Markers Require ThoroughAnalytic Validation. Arch Path Lab
Med. Aug 2019, Vol. 143, No. 8. pp.
907-909.
Torlakovic EE. How to Validate
Predictive Immunohistochemistry
Testing in Pathology? Arch Path Lab
Med. Aug 2019, Vol. 143, No. 8. pp.
907-907.

Laboratory Processes
G. Staining
v. In Situ Hybridization•

Establish a procedure for selection and development of probes to be added tomenu:oPreparation and cutting of tissue sectionoSelection of probeo

Validation of application methodPretreatment

Anatomic Pathology Checklist, ANP.22956(ISH Probe Validation/ Verification)

Clinical Laboratory Standards Institute
CLSI – MM13, Collection, Transport,
Preparation, and Storage of Specimens
for Molecular Methods: 2020.

Anatomic Pathology Checklist, ANP.22978(Predictive Marker Testing – Validation/
Verification)

International Standard ISO 201664:2020 - Molecular in vitro diagnostic
examinations – Specifications for preexamination processes for formalinfixed and paraffin-embedded (FFPE)

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•



Antibody dilutionRetrieval method – if required

Detection method
•

DAB

•

Alkaline phosphatase•

FluorescentoSelection and validation of control materialoInstructions on how to score slide and expected resultsoDocumentation of validation; record test tissue, expected results, actual
results, and changes to methodoStorage of probe and reagentsoRetention and storage of slides and or images

Establish procedures for change of:oMethodology

oReagentoAntibody

o



Clone



Lot number

Dilution

Equipment

New model



Major service repairMove or relocation

Anatomic Pathology Checklist, ANP.22964(ISH Controls)
Anatomic Pathology Checklist, ANP.22959(ISH Assay Performance)

Anatomic Pathology Checklist, ANP.22963(ISH scoring)

Anatomic Pathology Checklist, ANP.22965(Image and Slide Retention – ISH)

Anatomic Pathology Checklist, ANP.22956(ISH Probe Validation/ Verification)

Anatomic Pathology Checklist, ANP.22963(ISH Scoring)

Anatomic Pathology Checklist, ANP.22964(ISH Controls)

All Common Checklist, COM.30450(New Reagent Lot and Shipment
Confirmation of Acceptability – Nonwaived
Tests)
Anatomic Pathology Checklist, ANP.22966(ISH Interpretation)

tissue for – Part 4: In situ detection
techniques: section 6 – Inside thelaboratory.
Clinical Laboratory Standards Institute
CLSI. MM7-A2 Fluorescence In Situ
Hybridization (FISH) Methods forClinical Labs, Approved Guideline, 2nded. 2013.
Bancroft J, Gamble M. Theory andPractice of Histological Techniques, 6thed. New York, NY: Churchill Livingston;
2008.
David J. Dabbs. Diagnostic
Immunohistochemistry: Theranostic andGenomic Applications, 3rd ed.
Philadelphia, PA: Saunders Elsevier;
2010.

Awatif I. AL-Nafussi, 2nd ed. Tumor
Diagnosis, Practical Approach andPattern Analysis. London, Hodde
Arnold; 2005.

American College of Medical Genetics
Laboratory. Standards and guidelines
for clinical genetics laboratories,
Bethesda, MD: ACMG; 2021

Clinical Laboratory Standards Institute
CLSI. MM7-A2 Fluorescence In Situ
Hybridization (FISH) Methods forClinical Labs, Approved Guideline, 2nded. 2013.

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•

Establish procedure for clinical validation of each probe:oNumber of tissue sections to be tested per probeoComparison of results to previous stained slides or duplicate slides stainedby accredited lab•

In Situ Hybridization (ISH) staining refers to a method using probes made up ofcomplementary strands used to target sequences of mRNA, viral DNA or
chromosomal DNA located in tissue cells.

•

Retention of Images and permanent slides

Jennings L, Van Deerlin VM, Gulley ML(2009) Recommended Principles andPractices for Validating ClinicalMolecular Pathology Tests. Arch Path
Lab Med. Vol. 133, No. 5: 743-755.
Wolff AC, Hammond ME, Allison KH,
Harvey BE, et al. Human Epidermal
Growth Factor Receptor 2 Testing in
Breast Cancer: American Society ofClinical Oncology/College of American
Pathologists Clinical Practice Guideline
Focused Update. Arch Path Lab Med.
Nov 2018, Vol. 142, No. 11. pp. 13641382.

Tanner M, Gancberg D, Di Leo A,
Larsimont D, Rouas G, Piccart MJ, etal. Chromogenic in situ hybridization: apractical alternative for fluorescence insitu hybridization to detect HER-2/neu
oncogene amplification in archival
breast cancer samples. Am J Pathol.
2000;157(5):1467-72.
Di Palma S, Collins N, Faulkes C, et al.
Chromogenic in situ hybridisation(CISH) should be an accepted methodin the routine diagnostic evaluation of
HER2 status in breast cancer. J Clin
Pathol. 2007;60(9):1067-8.
Gong Y, Gilcrease M, Sneige N.
Reliability of chromogenic in situ
hybridization for detecting HER-2 genestatus in breast cancer: comparison
with fluorescence in situ hybridization
and assessment of interobserver

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reproducibility. Mod Pathol.
2005;18(8):1015-21.
Hauser-Kronberger C, Dandachi N.
Comparison of chromogenic in situ
hybridization with other methodologies
for HER2 status assessment in breast
cancer. J Mol Histol. 2004;35(6):647-53.
Saez A, Andreu FJ, Segui MA, et al.
HER-2 gene amplification bychromogenic in situ hybridisation(CISH) compared with fluorescence insitu hybridisation (FISH) in breast
cancer-A study of two hundred cases.
Breast. 2006;15(4):519-27.
Bhargava R, Lal P, Chen B.
Chromogenic in situ hybridization forthe detection of HER-2/neu gene
amplification in breast cancer with anemphasis on tumors with borderline andlow-level amplification: does it measure
up to fluorescence in situ hybridization?
Am J Clin Pathol. 2005;123(2):237-43.
Dietel M, Ellis IO, Höfler H, et al.
Comparison of automated silver
enhanced in situ hybridisation (SISH)
and fluorescence ISH (FISH) for thevalidation of HER2 gene status inbreast carcinoma according to theguidelines of the American Society ofClinical Oncology and the College ofAmerican Pathologists. Virchows Arch.
2007;451(1):19-25.
van de Vijver M, Bilous M, Hanna W,
Hofmann M, Kristel P, Penault- Llorca
F, et al. Chromogenic in situ
hybridisation for the assessment ofVersion: 11.0
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68

HER2 status in breast cancer: an
international validation ring study.
Breast Cancer Res. 2007;9(5):R68.
Bilous M, Morey A, Armes J, Cummings
M, Francis G. Chromogenic in situ
hybridisation testing for HER2 gene
amplification in breast cancer produces
highly reproducible results concordant
with fluorescence in situ hybridisationand immunohistochemistry. Pathology.
2006;38(2):120-4.

Di Palma S, Collins N, Bilous M, Sapino
A, Mottolese M, Kapranos N, et al. A
quality assurance exercise to evaluate
the accuracy and reproducibility ofchromogenic in situ hybridisation for
HER2 analysis in breast cancer. J Clin
Pathol. 2008;61(6):757-60.
Laboratory Processes
G. Staining
v.Immunohistochemistry
and In Situ Hybridizationa. Quality assurance• Establish Quality Assurance procedures for IHC and ISH procedures to include:oCompilation of predictive marker results


Total cases

% positive, % negativeComparison to benchmarks



Corrective action taken

Anatomic Pathology Checklist, ANP.22970(Annual Result Comparison – Breast
Carcinoma)

Allison KH, Hammond EH, Dowsett M,
McKernin SE et al. Estrogen andProgesterone Receptor Testing in
Breast Cancer American Society ofClinical Oncology/College of American
Pathologists Guideline Update. ArchPath Lab Med. Early Online Release.
doi: 10.5858/arpa.2019-0904-SA
Wolff AC, Hammond ME, Allison KH,
Harvey BE, et al. Human Epidermal
Growth Factor Receptor 2 Testing in
Breast Cancer: American Society ofVersion: 11.0
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69

• Documented participation in external proficiency testing for HER2 and ER
All Common Checklist, COM.01520(PT and Alternative Performance
Assessment for IHC, ICC, and ISH Predictive
Markers)

Clinical Oncology/College of American
Pathologists Clinical Practice Guideline
Focused Update. Arch Path Lab Med;Nov 2018, Vol. 142, No. 11. pp. 13641382.
Fitzgibbons PL, Bradley LA, Fatheree
LA, Alsabeh R, et.al. Principles ofAnalytic Validation of
Immunohistochemical Assays:
Guideline From the College of American
Pathologists Pathology and Laboratory
Quality Center. Arch Path Lab Med;Nov 2014, Vol. 138, No. 11. pp. 14321443.

COVERSLIPPINGLABORATORY PROCESSES - COVERSLIPPING

Guideline Section

Statement

CAP Checklist

Reference• Establish manual coverslipping procedures that:

Laboratory General Checklist, GEN.77200(Ergonomics)

Bancroft J, Gamble M. Theory andPractice of Histological Techniques, 6thed. New York, NY: Churchill Livingston;
2008.

Laboratory Processes
H. Coverslippingi. Manual/Automated•oInclude ergonomic techniquesoReduce chemical exposure

Use mounting media with an appropriate refractive index for proper resolution:oAqueous vs. non aqueouso

Non fluorescent•

Identify size and weight of coverslip to be used•

Identify drying method of coverslip and slide•

Establish validation and operation procedures for an automated coverslipper:

Carson F, Hladik C. Histotechnology A
Self- Instructional Text, 5th ed. Chicago,
IL: ASCP Press; 2020.

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o
o
o
o
o
o
o
o
•

Speed of operation
Type of mounting media
Size and type of coverslip
Type and volume of transfer fluid (xylene or xylene substitute)
Cleaning and maintenance
Reagent filling or change
Filter change
Drying time

Establish a preventative maintenance program that includes annual service andemergency service.

All Common Checklist, COM.30575(Instrument/Equipment Operation)
All Common Checklists, COM.30600(Maintenance/Function Checks)

All Common Checklist, COM.30675(Instrument and Equipment Records)

END

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