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22739 Immunohistochemistry for Semi-Quantitative HER2

Immunohistochemistry for Semi-Quantitative HER2
Test Code: HMISC
Synonyms/Keywords
Histology, HER2/Neu, Surgical Pathology, Immunohistochemistry, C-erbB2, HER2, Gastric, Breast, Esophageal ​
Useful For
​Determining over-expression of HER2 protein in formalin-fixed, paraffin-embedded tissue sections. Tissues frequently evaluated for HER2 overexpression are breast, gastric, and esophageal invasive cancers.
Specimen Requirements
Specimen Type Preferred Container/Tube Acceptable Container/Tube Specimen Volume Specimen Minimum Volume
(allows for 1 repeat)		 Pediatric Minimum Volume
(no repeat)
​Paraffin- Embedded Tissue Block or Unstained Sections on Charged Slides ​ Transport Container for Block and/or Slide Holders​ ​ ​ ​
​Completed Pathology Report​​​​​
Collection Processing Instructions
A. BREAST
1. Ischemic Time - removal to initiation of fixation be less than or equal to one hour.
2. Fixative type - 10% Neutral Buffered Formalin.
3. Fixed in 10% NBF for at least 6 hours and less than 72 hours
The above handling and fixative times must be documented on the accession slip or the report or both according to ASCO/CAP guidelines as delayed, under, or over fixation may affect test results.
 
Handling of Surgical Breast Specimens Obtained Remotely -
The following procedure, for breast specimens when cancer is suspected/known. The goal is to expose breast tumor to formalin within one hour of removal.
- Breast lumpectomy
- Pat the specimen dry
- Ink the margins - Inking: Unoriented specimens are inked one color. Oriented specimens are inked six colors. You can ink however you want but write on the requisition slip/histology order how you have inked it. One suggestion for inking:
·         Superior – blue
·         Inferior – green
·         Medial – red
·         Lateral – orange
·         Anterior/Superficial – yellow
·         Posterior/Deep - black
- Dry again and dip in vinegar (helps the ink stick)
- Make an incision in the specimen as close as possible to the lesion. Do not cut all the way through the specimen - keep it intact. More than one incision can be made on larger specimens.
- Cut opposite the margin closest to the tumor.
 
Remember the specimen needs to be inked before it is incised in order to maintain integrity of the margin. Inking after the margin has been cut is not recommended. Specimen should be immersed in a sufficient volume of 10% neutral buffered formalin (10X volume of specimen).
Requirements:
Completed histology requisition form or Authorization for Test Requests form.
Surgical or cytology pathology report containing pertinent patient history, tissue source, and diagnosis.
One paraffin tissue/cell block with corresponding H&E slide, otherwise send 2 (4-5 micron) unstained, air-dried, charged slides with corresponding H&E slide.
 
NOTE: For HER2 FISH testing, send 2 (4-5 micron) unstained, air-dried, charged slides with corresponding H & E slide. 
 
 Contact Histology Pathology Assistants for more information
 
B. GASTRIC/ESOPHAGEAL RESECTIONS OR BIOPSY requirements
- Fixative type - 10% Neutral Buffered Formalin. It is recommended (not required) that since proper fixation is critical for quality staining, resection and biopsy specimens be handled similar to that of breast specimens.
 
Requirements:
Completed histology requisition form or Authorization for Test Requests form and include: Surgical or cytology pathology report containing pertinent patient history, tissue source, and diagnosis.
One paraffin tissue/cell block with corresponding H&E slide, otherwise send 2 (4-5 micron) unstained, air-dried, charged slides with corresponding H&E slide.
 
NOTE: For HER2 FISH testing, send 2 (4-5 micron) unstained, air-dried, charged slides with corresponding H & E slide. 
 
HER2 FISH testing is a send out test. 
 
Antibody validation was performed on 10% neutral buffered, formalin-fixed, paraffin embedded tissues/cell blocks.
​
Acceptable Body Sites
​Breast. gastric, and/or their metastases
Acceptable Specimen Types
​Paraffin-embedded tissue blocks and/or unstained charged slides fixed in 10% neutral buffered formalin.  Unstained slides require an H&E slide or an additional unstained slide to be stained for H&E.
Specimen Stability Information
Specimen Type Temperature Time
​Paraffin Tissue/Cell Block, Fixed in 10% NBF Room Temperature/Ambient Indefinite
Unstained, Air-Dried, Charged Slides Room Temperature/​Ambient Ideally slides should be < or = to 6 weeks old
Rejection Criteria
​No Invasive Cancer Present
​Fixative other than 10% Neutral Buffered Formalin
​Decalcified Tissue (if necessary, will require a disclaimer).
​Unstained Slides (no charge on slides)
​Insufficient Data (Incomplete requisition and/or Pathology Report) may delay testing.
Interference
Proper fixation conditions are critical for quality staining. Under or over fixation, or delay to fixative can affect quality results. These tests are not validated on fixatives other than 10% neutral buffered formalin or decalcified tissues.
Performing Laboratory Information
Performing Location Day(s) Test Performed Analytical Time Methodology/Instrumentation
​Marshfield Monday through Friday 36 hours​ Polymer based detection system/Leica Bond III
Reference Range Information
Performing Location Reference Range
​Marshfield Interpretative Report Semi-Quantitative/IHC  ​
Interpretation
 
Test is only available with interpretation.
 
Interpretation: Semi-Quantitative Immunohistochemistry, Manual
 
FISH HER 2 may be ordered and is available separately. All 2+ results will automatically be tested for FISH Her2 at an additional fee.
 
HER2 FISH testing is a send out test.

All blocks and slides will be returned to the requestor.


1. BREAST CANCER Requirements
- Time to fixation < or = 1hr
- Fixed in 10% Neutral Buffered Formalin (NBF)
- Fixed in 10% NBF for at least 6 hours and less than 72 hours

HER2/NEU Scoring System and Criteria
 
IHC
Result
Criteria
0
Negative
No staining is observed* or membrane staining that is incomplete and is faint/barely perceptible and within ≤10% of invasive tumor cells.
1+
Negative
Incomplete membrane staining that is faint/barely perceptible and within >10% of invasive tumor cells*.
2+
Equivocal
Weak to moderate complete membrane staining observed in >10% of tumor cells.
3+
Positive
Circumferential membrane staining that is complete, intense, and within >10% of invasive tumor cells*.
 * See comment
Indeterminate
Technical issues prevent tests from being reported as positive, negative, or equivocal, which may include:
·         Inadequate specimen handling
·         Artifacts that make interpretation difficult
·         Analytic testing failure
·         Sample has strong membrane staining of normal breast ducts
 
Over-expression of HER2 cell membrane receptor protein in breast carcinoma is associated with tumor cell growth, aggressive disease and shortened survival. A positive test result aids in the assessment for possible treatment with Herceptin (trastuzumab) that is targeted against the HER2 receptor protein. The prognostic and therapeutic significance of the results of this assay in other tumors is under investigation. The FDA has approved the reagents used in this immunohistochemistry assay for assessment of HER2 receptor status. The range of over-expression among breast cancers is reportedly between 10% to 20% of primary breast carcinomas.
2. GASTRIC/ESOPHAGEAL RESECTION Requirements
- Fixed in 10% Neutral Buffered Formalin (NBF)
HER2/NEU Scoring System and Criteria
IHC
Result
Criteria
0
Negative
No reactivity or membranous reactivity in < 10% of tumor cells
1+
Negative
Faint/barely perceptible membranous reactivity in >= 10% of tumor cells; cells reactive only in part of their membrane (Reflex to FISH)
2+
Equivocal
Weak to moderate complete, basolateral or lateral membranous reactivity in >= 10% of tumor cells
3+
Positive
Strong, complete, basolateral or lateral membranous reactivity in >=10% of tumor cells
 
3. GASTRIC/ESOPHAGEAL BIOPSY Requirements
- Fixed in 10% Neutral Buffered Formalin (NBF)
HER2/NEU Scoring System and Criteria
IHC
Result
Criteria
0
Negative
No reactivity in any tumor cells
1+
Negative
Tumor cell cluster (>=5 cells) with a faint/barely perceptible membranous reactivity irrespective of tumor cells stained
2+
Equivocal
Tumor cell cluster (>=5 cells) with weak to moderate complete, basolateral or lateral membranous reactivity irrespective of percentage of tumor cells stained (Reflex to FISH)
3+
Positive
Tumor cell cluster (>=5 cells) with a strong complete, basolateral or lateral membranous reactivity irrespective of percentage of tumor cells stained.
 
Depending on the series, the prevalence of HER2 expression in gastric cancer seems to be 15-25%. The level of expression of intestinal type gastric cancer seems to be much higher (~32%) compared with diffuse-type gastric cancer (~6%). Also, tumors that are primarily located at the gastroesophageal junction seem to have higher HER2 positive rates compared to tumors that occur in the rest of the stomach (33% versus 21%).(7)
 

ALL
The clone used is a mouse monoclonal, CB11 and detection system used is a polymer based. This test is validated on paraffin-embedded, 10% neutral formalin fixed tiossues.  Interpretation follows the American Society for Clinical Oncology (ASCO) and College of American Pathologists (CAP) guidelines. 

References:

1. Reference: Wolff AC, Hammond ME, Allison KH, Harvey BE, Mangu PB, Bartlett JM, Bilous M, Ellis IO, Fitzgibbons P, Hanna W, Jenkins RB, Press MF, Spears PA, et al. Human Epidermal Growth Factor Receptor 2 Testing in Breast Cancer: American Society of Clinical Oncology/College of American Pathologists Clinical Practice Guideline Focused Update. J Clin Oncol. 2018;36:2105–22.

2. Walker Ra, Bartlett JMS Dowsett M, Ellis IO, Hanby An, Jasani B, Miller K, and Pinder SE.  HER2 Testing in the UK-Further Update to Recommendations.  Journal of Clinical Pathology 2008. 

3. ​Yaziji H, Goldstein LC, Barry TS, Werling R, Hwang H, Ellis GK, Gralow JR, Livingston RB, Gown AM: HER-2 testing in breast cancer using parallel tissue-based methods. JAMA 2004, 291:1972-1977.

4.Owens MA, Horten BC, Da Silva MM: HER2 amplification ratios by fluorescence in situ hybridization and correlation with immunohistochemistry in a cohort of 6556 breast cancer tissues. Clin Breast Cancer 2004, 5:63-69.

5. Hofmann M et al, Assessment of a HER2 scoring system for gastric cancer: results from a validation study. Histopathol 52:797. 2008.

6. Bang Y et al, Pathologic features of advanced gastric cancer (GC): relationship to human epidermal growth factor receptor 2 (HER2) positivity in the global screening programme of the ToGA trial. J Clin Oncol 25:15s (suppl; abstr 4556).

(7) Reference:

Reference Information:
Questions Relating to Immunohistochemistry for HER2 on Gastric and Gastroesophageal Junction Carcinomas

http://www.cap.org/apps/docs/committees/immunohistochemistry/her2_faqs_gastric.pdf
Outreach CPTs
CPT Modifier
(if needed)		 Quantity Description Comments
88360​ ​ ​1 ​
Classification
This test was developed and its performance characteristics determined by Marshfield Labs. It has not been cleared or approved by the U.S. Food and Drug Administration. The FDA has determined that such clearance or approval is not necessary. This test is for clinical purposes. It should not be regarded as investigational or for research. This laboratory is certified under the Clinical Laboratory Improvement Amendments (CLIA) as qualified to perform high complexity testing.
 
 
Synonyms/Keywords
Histology, HER2/Neu, Surgical Pathology, Immunohistochemistry, C-erbB2, HER2, Gastric, Breast, Esophageal ​
Ordering Applications
Ordering Application Description
​Centricity ​None
​Cerner ​Her-2 (c-erbB2) Oncoprotein
If the ordering application you are looking for is not listed, contact your local laboratory for assistance.
Specimen Requirements
Specimen Type Preferred Container/Tube Acceptable Container/Tube Specimen Volume Specimen Minimum Volume
(allows for 1 repeat)		 Pediatric Minimum Volume
(no repeat)
​Paraffin- Embedded Tissue Block or Unstained Sections on Charged Slides ​ Transport Container for Block and/or Slide Holders​ ​ ​ ​
​Completed Pathology Report​​​​​
Collection Processing
A. BREAST
1. Ischemic Time - removal to initiation of fixation be less than or equal to one hour.
2. Fixative type - 10% Neutral Buffered Formalin.
3. Fixed in 10% NBF for at least 6 hours and less than 72 hours
The above handling and fixative times must be documented on the accession slip or the report or both according to ASCO/CAP guidelines as delayed, under, or over fixation may affect test results.
 
Handling of Surgical Breast Specimens Obtained Remotely -
The following procedure, for breast specimens when cancer is suspected/known. The goal is to expose breast tumor to formalin within one hour of removal.
- Breast lumpectomy
- Pat the specimen dry
- Ink the margins - Inking: Unoriented specimens are inked one color. Oriented specimens are inked six colors. You can ink however you want but write on the requisition slip/histology order how you have inked it. One suggestion for inking:
·         Superior – blue
·         Inferior – green
·         Medial – red
·         Lateral – orange
·         Anterior/Superficial – yellow
·         Posterior/Deep - black
- Dry again and dip in vinegar (helps the ink stick)
- Make an incision in the specimen as close as possible to the lesion. Do not cut all the way through the specimen - keep it intact. More than one incision can be made on larger specimens.
- Cut opposite the margin closest to the tumor.
 
Remember the specimen needs to be inked before it is incised in order to maintain integrity of the margin. Inking after the margin has been cut is not recommended. Specimen should be immersed in a sufficient volume of 10% neutral buffered formalin (10X volume of specimen).
Requirements:
Completed histology requisition form or Authorization for Test Requests form.
Surgical or cytology pathology report containing pertinent patient history, tissue source, and diagnosis.
One paraffin tissue/cell block with corresponding H&E slide, otherwise send 2 (4-5 micron) unstained, air-dried, charged slides with corresponding H&E slide.
 
NOTE: For HER2 FISH testing, send 2 (4-5 micron) unstained, air-dried, charged slides with corresponding H & E slide. 
 
 Contact Histology Pathology Assistants for more information
 
B. GASTRIC/ESOPHAGEAL RESECTIONS OR BIOPSY requirements
- Fixative type - 10% Neutral Buffered Formalin. It is recommended (not required) that since proper fixation is critical for quality staining, resection and biopsy specimens be handled similar to that of breast specimens.
 
Requirements:
Completed histology requisition form or Authorization for Test Requests form and include: Surgical or cytology pathology report containing pertinent patient history, tissue source, and diagnosis.
One paraffin tissue/cell block with corresponding H&E slide, otherwise send 2 (4-5 micron) unstained, air-dried, charged slides with corresponding H&E slide.
 
NOTE: For HER2 FISH testing, send 2 (4-5 micron) unstained, air-dried, charged slides with corresponding H & E slide. 
 
HER2 FISH testing is a send out test. 
 
Antibody validation was performed on 10% neutral buffered, formalin-fixed, paraffin embedded tissues/cell blocks.
​
Acceptable Body Sites
​Breast. gastric, and/or their metastases
Acceptable Specimen Types
​Paraffin-embedded tissue blocks and/or unstained charged slides fixed in 10% neutral buffered formalin.  Unstained slides require an H&E slide or an additional unstained slide to be stained for H&E.
Specimen Stability Information
Specimen Type Temperature Time
​Paraffin Tissue/Cell Block, Fixed in 10% NBF Room Temperature/Ambient Indefinite
Unstained, Air-Dried, Charged Slides Room Temperature/​Ambient Ideally slides should be < or = to 6 weeks old
Rejection Criteria
​No Invasive Cancer Present
​Fixative other than 10% Neutral Buffered Formalin
​Decalcified Tissue (if necessary, will require a disclaimer).
​Unstained Slides (no charge on slides)
​Insufficient Data (Incomplete requisition and/or Pathology Report) may delay testing.
Interference
Proper fixation conditions are critical for quality staining. Under or over fixation, or delay to fixative can affect quality results. These tests are not validated on fixatives other than 10% neutral buffered formalin or decalcified tissues.
Useful For
​Determining over-expression of HER2 protein in formalin-fixed, paraffin-embedded tissue sections. Tissues frequently evaluated for HER2 overexpression are breast, gastric, and esophageal invasive cancers.
Reference Range Information
Performing Location Reference Range
​Marshfield Interpretative Report Semi-Quantitative/IHC  ​
Interpretation
 
Test is only available with interpretation.
 
Interpretation: Semi-Quantitative Immunohistochemistry, Manual
 
FISH HER 2 may be ordered and is available separately. All 2+ results will automatically be tested for FISH Her2 at an additional fee.
 
HER2 FISH testing is a send out test.

All blocks and slides will be returned to the requestor.


1. BREAST CANCER Requirements
- Time to fixation < or = 1hr
- Fixed in 10% Neutral Buffered Formalin (NBF)
- Fixed in 10% NBF for at least 6 hours and less than 72 hours

HER2/NEU Scoring System and Criteria
 
IHC
Result
Criteria
0
Negative
No staining is observed* or membrane staining that is incomplete and is faint/barely perceptible and within ≤10% of invasive tumor cells.
1+
Negative
Incomplete membrane staining that is faint/barely perceptible and within >10% of invasive tumor cells*.
2+
Equivocal
Weak to moderate complete membrane staining observed in >10% of tumor cells.
3+
Positive
Circumferential membrane staining that is complete, intense, and within >10% of invasive tumor cells*.
 * See comment
Indeterminate
Technical issues prevent tests from being reported as positive, negative, or equivocal, which may include:
·         Inadequate specimen handling
·         Artifacts that make interpretation difficult
·         Analytic testing failure
·         Sample has strong membrane staining of normal breast ducts
 
Over-expression of HER2 cell membrane receptor protein in breast carcinoma is associated with tumor cell growth, aggressive disease and shortened survival. A positive test result aids in the assessment for possible treatment with Herceptin (trastuzumab) that is targeted against the HER2 receptor protein. The prognostic and therapeutic significance of the results of this assay in other tumors is under investigation. The FDA has approved the reagents used in this immunohistochemistry assay for assessment of HER2 receptor status. The range of over-expression among breast cancers is reportedly between 10% to 20% of primary breast carcinomas.
2. GASTRIC/ESOPHAGEAL RESECTION Requirements
- Fixed in 10% Neutral Buffered Formalin (NBF)
HER2/NEU Scoring System and Criteria
IHC
Result
Criteria
0
Negative
No reactivity or membranous reactivity in < 10% of tumor cells
1+
Negative
Faint/barely perceptible membranous reactivity in >= 10% of tumor cells; cells reactive only in part of their membrane (Reflex to FISH)
2+
Equivocal
Weak to moderate complete, basolateral or lateral membranous reactivity in >= 10% of tumor cells
3+
Positive
Strong, complete, basolateral or lateral membranous reactivity in >=10% of tumor cells
 
3. GASTRIC/ESOPHAGEAL BIOPSY Requirements
- Fixed in 10% Neutral Buffered Formalin (NBF)
HER2/NEU Scoring System and Criteria
IHC
Result
Criteria
0
Negative
No reactivity in any tumor cells
1+
Negative
Tumor cell cluster (>=5 cells) with a faint/barely perceptible membranous reactivity irrespective of tumor cells stained
2+
Equivocal
Tumor cell cluster (>=5 cells) with weak to moderate complete, basolateral or lateral membranous reactivity irrespective of percentage of tumor cells stained (Reflex to FISH)
3+
Positive
Tumor cell cluster (>=5 cells) with a strong complete, basolateral or lateral membranous reactivity irrespective of percentage of tumor cells stained.
 
Depending on the series, the prevalence of HER2 expression in gastric cancer seems to be 15-25%. The level of expression of intestinal type gastric cancer seems to be much higher (~32%) compared with diffuse-type gastric cancer (~6%). Also, tumors that are primarily located at the gastroesophageal junction seem to have higher HER2 positive rates compared to tumors that occur in the rest of the stomach (33% versus 21%).(7)
 

ALL
The clone used is a mouse monoclonal, CB11 and detection system used is a polymer based. This test is validated on paraffin-embedded, 10% neutral formalin fixed tiossues.  Interpretation follows the American Society for Clinical Oncology (ASCO) and College of American Pathologists (CAP) guidelines. 

References:

1. Reference: Wolff AC, Hammond ME, Allison KH, Harvey BE, Mangu PB, Bartlett JM, Bilous M, Ellis IO, Fitzgibbons P, Hanna W, Jenkins RB, Press MF, Spears PA, et al. Human Epidermal Growth Factor Receptor 2 Testing in Breast Cancer: American Society of Clinical Oncology/College of American Pathologists Clinical Practice Guideline Focused Update. J Clin Oncol. 2018;36:2105–22.

2. Walker Ra, Bartlett JMS Dowsett M, Ellis IO, Hanby An, Jasani B, Miller K, and Pinder SE.  HER2 Testing in the UK-Further Update to Recommendations.  Journal of Clinical Pathology 2008. 

3. ​Yaziji H, Goldstein LC, Barry TS, Werling R, Hwang H, Ellis GK, Gralow JR, Livingston RB, Gown AM: HER-2 testing in breast cancer using parallel tissue-based methods. JAMA 2004, 291:1972-1977.

4.Owens MA, Horten BC, Da Silva MM: HER2 amplification ratios by fluorescence in situ hybridization and correlation with immunohistochemistry in a cohort of 6556 breast cancer tissues. Clin Breast Cancer 2004, 5:63-69.

5. Hofmann M et al, Assessment of a HER2 scoring system for gastric cancer: results from a validation study. Histopathol 52:797. 2008.

6. Bang Y et al, Pathologic features of advanced gastric cancer (GC): relationship to human epidermal growth factor receptor 2 (HER2) positivity in the global screening programme of the ToGA trial. J Clin Oncol 25:15s (suppl; abstr 4556).

(7) Reference:

Reference Information:
Questions Relating to Immunohistochemistry for HER2 on Gastric and Gastroesophageal Junction Carcinomas

http://www.cap.org/apps/docs/committees/immunohistochemistry/her2_faqs_gastric.pdf
For more information visit:
Performing Laboratory Information
Performing Location Day(s) Test Performed Analytical Time Methodology/Instrumentation
​Marshfield Monday through Friday 36 hours​ Polymer based detection system/Leica Bond III
For billing questions, see Contacts
Outreach CPTs
CPT Modifier
(if needed)		 Quantity Description Comments
88360​ ​ ​1 ​
Classification
This test was developed and its performance characteristics determined by Marshfield Labs. It has not been cleared or approved by the U.S. Food and Drug Administration. The FDA has determined that such clearance or approval is not necessary. This test is for clinical purposes. It should not be regarded as investigational or for research. This laboratory is certified under the Clinical Laboratory Improvement Amendments (CLIA) as qualified to perform high complexity testing.
 
 
For most current information refer to the Marshfield Laboratory online reference manual.