Analytic variability in immunohistochemistry biomarker studies

Valsamo K. Anagnostou, Allison W. Welsh, Jennifer M. Giltnane, Summar Siddiqui, Camil Lioeaga, Mark Gustavson, Konstantinos N. Syrigos, Jill Reiter, David L. Rimm

Research output: Contribution to journalArticle

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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 r2 = 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 r2 = 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.

Original languageEnglish (US)
Pages (from-to)982-991
Number of pages10
JournalCancer Epidemiology Biomarkers and Prevention
Volume19
Issue number4
DOIs
StatePublished - Apr 2010
Externally publishedYes

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Epidermal Growth Factor Receptor
Biomarkers
Immunohistochemistry
Antibodies
Estrogen Receptors
Western Blotting
MCF-7 Cells
Survival Analysis
Tumor Biomarkers
Quality Control
Fluorescent Antibody Technique
Reading
Lung Neoplasms
Clone Cells
Guidelines
Staining and Labeling
Breast Neoplasms
Survival

ASJC Scopus subject areas

  • Epidemiology
  • Oncology

Cite this

Anagnostou, V. K., Welsh, A. W., Giltnane, J. M., Siddiqui, S., Lioeaga, C., Gustavson, M., ... Rimm, D. L. (2010). Analytic variability in immunohistochemistry biomarker studies. Cancer Epidemiology Biomarkers and Prevention, 19(4), 982-991. https://doi.org/10.1158/1055-9965.EPI-10-0097

Analytic variability in immunohistochemistry biomarker studies. / Anagnostou, Valsamo K.; Welsh, Allison W.; Giltnane, Jennifer M.; Siddiqui, Summar; Lioeaga, Camil; Gustavson, Mark; Syrigos, Konstantinos N.; Reiter, Jill; Rimm, David L.

In: Cancer Epidemiology Biomarkers and Prevention, Vol. 19, No. 4, 04.2010, p. 982-991.

Research output: Contribution to journalArticle

Anagnostou, VK, Welsh, AW, Giltnane, JM, Siddiqui, S, Lioeaga, C, Gustavson, M, Syrigos, KN, Reiter, J & Rimm, DL 2010, 'Analytic variability in immunohistochemistry biomarker studies', Cancer Epidemiology Biomarkers and Prevention, vol. 19, no. 4, pp. 982-991. https://doi.org/10.1158/1055-9965.EPI-10-0097
Anagnostou VK, Welsh AW, Giltnane JM, Siddiqui S, Lioeaga C, Gustavson M et al. Analytic variability in immunohistochemistry biomarker studies. Cancer Epidemiology Biomarkers and Prevention. 2010 Apr;19(4):982-991. https://doi.org/10.1158/1055-9965.EPI-10-0097
Anagnostou, Valsamo K. ; Welsh, Allison W. ; Giltnane, Jennifer M. ; Siddiqui, Summar ; Lioeaga, Camil ; Gustavson, Mark ; Syrigos, Konstantinos N. ; Reiter, Jill ; Rimm, David L. / Analytic variability in immunohistochemistry biomarker studies. In: Cancer Epidemiology Biomarkers and Prevention. 2010 ; Vol. 19, No. 4. pp. 982-991.
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AU - Welsh, Allison W.

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AU - Lioeaga, Camil

AU - Gustavson, Mark

AU - Syrigos, Konstantinos N.

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