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Analytic variability in immunohistochemistry biomarker studies.

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

SUBMITTER: Anagnostou VK

PROVIDER: S-EPMC3891912 | biostudies-literature | 2010 Apr

REPOSITORIES: biostudies-literature

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Analytic variability in immunohistochemistry biomarker studies.

Anagnostou Valsamo K VK Welsh Allison W AW Giltnane Jennifer M JM Siddiqui Summar S Liceaga Camil C Gustavson Mark M Syrigos Konstantinos N KN Reiter Jill L JL Rimm David L DL

Cancer epidemiology, biomarkers & prevention : a publication of the American Association for Cancer Research, cosponsored by the American Society of Preventive Oncology 20100323 4

<h4>Background</h4>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.<h4>Methods</h4>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, ...[more]

PMID: 20332259

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Project description:SignificanceConcerns about the reproducibility of experimental findings have recently emerged in many disciplines, from psychology to medicine and neuroscience. As NIRS is a relatively recent brain imaging technique, the question of reproducibility has not yet been systematically addressed.AimThe current study seeks to test the replicability of effects observed in NIRS experiments assessing young infants' rule-learning ability.ApproachWe conducted meta-analyses and mixed-effects modeling-based inferential statistics to determine whether effect sizes were replicable and comparable in a sample of 23 NIRS studies investigating infants' abilities to process repetition- and diversity-based regularities in linguistic and nonlinguistic auditory and visual sequences. Additionally, we tested whether effect sizes were modulated by different factors such as the age of participants or the laboratory. We obtained NIRS data from 12 published and 11 unpublished studies. The 23 studies involved a total of 487 infants, aged between 0 and 9 months, tested in four different countries (Canada, France, Italy, and USA).ResultsOur most important finding is that study and laboratory were never significant moderators of variation in effect sizes, indicating that results replicated reliably across the different studies and labs included in the sample. We observed small-to-moderate effect sizes, similar to effect sizes found with other neuroimaging and behavioral techniques in the developmental literature. In line with existing findings, effect sizes were modulated by the participants' age and differed across the different regularities tested, with repetition-based regularities giving rise to the strongest effects; in particular, the overall magnitude of this effect in the left temporal region was 0.27 when analyzing the entire dataset.ConclusionsMeta-analysis is a useful tool for assessing replicability and cross-study variability. Here, we have shown that infant NIRS studies in the language domain replicate robustly across various NIRS machines, testing sites, and developmental populations.

Project description:BackgroundIntestinal metaplasia (IM) is considered a key pivot point in the Correa model of gastric cancer (GC). It is histologically subtyped into the complete and incomplete subtypes, the latter being associated with a greater risk of progression. However, the clinical utility of IM subtyping remains unclear, partially due to the absence of reliable defining biomarkers.MethodsBased on gene expression data and existing literature, we selected CD10 and Das1 as candidate biomarkers to distinguish complete and incomplete IM glands in tissues from patients without GC (IM-GC) and patients with GC (IM + GC). Immunohistochemical staining of individually subtyped IM glands was scored after blinding by two researchers using tissue belonging to both IM-GC and IM + GC patients. Whole tissue Das1 staining was further assessed using digital image quantification (cellSens Dimension, Olympus).ResultsAcross both cohorts CD10 stained the IM brush border and was shown to have a high sensitivity (87.5% and 94.9% in IM-GC and IM + GC patients respectively) and specificity (100.0% and 96.7% respectively) with an overall AUROC of 0.944 for complete IM glands. By contrast Das1 stained mainly goblet cells and the apical membrane of epithelial cells, mostly of incomplete IM glands with a low sensitivity (28.6% and 29.3% in IM-GC and IM + GC patients respectively) but high specificity (98.3% and 85.1% respectively) and an overall AUROC of 0.603 for incomplete IM glands. A combined logistic regression model showed a significant increase in AUROC for detecting complete IM glands (0.955 vs 0.970). Whole tissue digital quantification of Das1 staining showed a significant association with incomplete IM compared to complete IM, both in IM-GC and in IM + GC patients (p = 0.016 and p = 0.009 respectively, Mann-Whitney test and unpaired t test used). Additionally, complete IM in IM + GC patients exhibited significantly more Das1 staining than in IM-GC patients (p = 0.019, Mann-Whitney test).ConclusionsThese findings suggest that CD10 is an outstanding biomarker for complete IM and Das1 may be useful as a secondary biomarker for IM glands at greater risk of progression irrespective of IM subtype. Overall, the clinical use of these biomarkers could lead to improved patient stratification and targeted surveillance.

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Analytic variability in immunohistochemistry biomarker studies.

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Analytic variability in immunohistochemistry biomarker studies.

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