Att. Cii - IHC_summaryrecommendations

Attachment Cii

Principles of Analytic Validation of
Immunohistochemical Assays
Summary of Recommendations
Guideline Statement

Strength of Recommendation

1. Laboratories must validate all IHC tests before placing into clinical
service.

Recommendation

Note: Such means include (but are not necessarily limited to):
Correlating the new test’s results with the morphology and expected results;
Comparing the new test’s results with the results of prior testing of the
same tissues with a validated assay in the same laboratory;
Comparing the new test’s results with the results of testing the same tissue
validation set in another laboratory using a validated assay;
Comparing the new test’s results with previously validated
non-immunohistochemical tests; or
Testing previously graded tissue challenges from a formal proficiency
testing program (if available) and comparing the results with the graded
responses.

2. For initial validation of every assay used clinically, with the exception
of HER2/neu, ER, and PgR (for which established validation
guidelines already exist), laboratories should achieve at least 90%
overall concordance between the new test and the comparator test
or expected results. If concordance is less than 90%, laboratories
need to investigate the cause of low concordance.

Recommendation

3. For initial analytic validation of nonpredictive factor assays,
laboratories should test a minimum of 10 positive and 10 negative
tissues. When the laboratory medical director determines that fewer
than 20 validation cases are sufficient for a specific marker (eg, rare
antigen), the rationale for that decision needs to be documented.

Expert Consensus Opinion

Note: The validation set should include high and low expressors for positive
cases when appropriate, and should span the expected range of clinical
results (expression levels) for markers that are reported quantitatively.

4. For initial analytic validation of all laboratory-developed predictive
marker assays (with the exception of HER2/neu, ER and PgR),
laboratories should test a minimum of 20 positive and 20 negative
tissues. When the laboratory medical director determines that fewer
than 40 validation tissues are sufficient for a specific marker, the
rationale for that decision needs to be documented.

Expert Consensus Opinion

Note: Positive cases in the validation set should span the expected range of
clinical results (expression levels). This recommendation does not apply to
any marker for which a separate validation guideline already exists.

5. For a marker with both predictive and nonpredictive applications,
laboratories should validate it as a predictive marker if it is used as
such.

Recommendation

Source: Fitzgibbons PL, Bradley LA, Fatheree LA, et al. Principles of analytic validation of immunohistochemical assays: Guideline from the
College of American Pathologists Pathology and Laboratory Quality Center. Arch Pathol Lab Med. 2014;138(11):1432–1443.
© 2015 College of American Pathologists. All rights reserved.
23685.060215

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Guideline Statement

Strength of Recommendation

6. When possible, laboratories should use validation tissues that have
been processed using the same fixative and processing methods as
cases that will be tested clinically.

Recommendation

7. If IHC is regularly done on cytologic specimens that are not processed Expert Consensus Opinion
in the same manner as the tissues used for assay validation
(eg, alcohol-fixed cell blocks, air-dried smears, formalin postfixed
specimens), laboratories should test a sufficient number of such
cases to ensure that assays consistently achieve expected results.
The laboratory medical director is responsible for determining the
number of positive and negative cases and the number of predictive
and nonpredictive markers to test.
8. If IHC is regularly done on decalcified tissues, laboratories should
test a sufficient number of such tissues to ensure that assays
consistently achieve expected results. The laboratory medical
director is responsible for determining the number of positive and
negative issues and the number of predictive and nonpredictive
markers to test.

Expert Consensus Opinion

9. Laboratories may use whole sections, TMAs and/or MTBs in their
validation sets as appropriate. Whole sections should be used if
TMAs/MTBs are not appropriate for the targeted antigen or if the
laboratory medical director cannot confirm that the fixation and
processing of TMAs/ MTBs is similar to clinical specimens.

Recommendation

10. When a new reagent lot is placed into clinical service for an existing
Expert Consensus Opinion
validated assay, laboratories should confirm the assay’s performance
with at least 1 known positive case and 1 known negative case.
11. Laboratories should confirm assay performance with at least 2
known positive and 2 known negative cases when an existing
validated assay has changed in any one of the following ways:
Antibody dilution;
Antibody vendor (same clone);
Incubation or retrieval times (same method).

Expert Consensus Opinion

12. Laboratories should confirm assay performance by testing a
sufficient number of cases to ensure that assays consistently
achieve expected results when any of the following have changed:
Fixative type;
Antigen retrieval method (eg, change in pH, different buffer,
different heat platform);
Antigen detection system;
Tissue processing or testing equipment;
Environmental conditions of testing (eg, laboratory relocation);
Laboratory water supply.

Expert Consensus Opinion

The laboratory medical director is responsible for determining how
many predictive and nonpredictive markers and how many positive
and negative tissues to test.
13. Laboratories should run a full revalidation (equivalent to initial
analytic validation) when the antibody clone is changed for an
existing validated assay.

Expert Consensus Opinion

14. The laboratory must document all validations and verifications in
compliance with regulatory and accreditation requirements.

Expert Consensus Opinion

Abbreviations: IHC, immunohistochemistry; ER, estrogen receptor; PgR, progesterone receptor; TMA, tissue microarray; MTB, multitissue
block

© 2015 College of American Pathologists. All rights reserved.
23685.060215

cap.org