Format

Send to:

Choose Destination
See comment in PubMed Commons below
Cancer Epidemiol Biomarkers Prev. 2010 Apr;19(4):982-91. doi: 10.1158/1055-9965.EPI-10-0097. Epub 2010 Mar 23.

Analytic variability in immunohistochemistry biomarker studies.

Author information

  • 1Department of Pathology, Yale Cancer Center, New Haven, CT, USA. valsamo.anagnostou@yale.edu

Abstract

BACKGROUND:

Despite the widespread use of immunohistochemistry (IHC), there are no standardization guidelines that control for antibody probe variability. Here we describe the effect of variable antibody reagents in the assessment of cancer-related biomarkers by IHC.

METHODS:

Estrogen receptor (ER), epidermal growth factor receptor (EGFR) 1, and human epidermal growth factor receptor 3 (HER3) were evaluated by quantitative immunofluorescence. Correlations between ER clones 1D5, SP1, F10, and ER60c, and EGFR monoclonal 31G7, 2-18C9, H11, and 15F8, and polyclonal 2232 antibodies were assessed in 642 breast cancer patients. HER3 was measured by RTJ1, RTJ2, SGP1, M7297, RB-9211, and C-17 antibodies in 42 lung cancer patients. Survival analysis was done with the use of multiple cutoff points to reveal any prognostic classification.

RESULTS:

All ER antibodies were tightly correlated (Pearson's r(2) = 0.94-0.96; P < 0.0001) and western blotting confirmed their specificity in MCF-7 and BT474 cells. All EGFR antibodies but 2232 yielded specific results in western blotting; however, only 31G7 and 2-18C9 were strongly associated (Pearson's r(2) = 0.61; P < 0.0001). HER3 staining was nonspecific and nonreproducible. High EGFR-expressing patients had a worse prognosis when EGFR was measured with H11 or 31G7 (log rank P = 0.015 and P = 0.06). There was no statistically significant correlation between survival and EGFR detected by 2-18C9, 15F8, or polyclonal 2232 antibodies.

CONCLUSIONS:

Antibody validation is a critical analytic factor that regulates IHC readings in biomarker studies. Evaluation of IHC proficiency and quality control are key components toward IHC standardization.

IMPACT:

This work highlights the importance of IHC standardization and could result in the improvement of clinically relevant IHC protocols.

PMID:
20332259
[PubMed - indexed for MEDLINE]
PMCID:
PMC3891912
Free PMC Article
PubMed Commons home

PubMed Commons

0 comments
How to join PubMed Commons

    Supplemental Content

    Full text links

    Icon for HighWire Icon for PubMed Central
    Loading ...
    Write to the Help Desk