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BMC Cancer

, 14:226

Translational oncology

Abstract

BackgroundAZD3514 inhibits and down regulates the androgen receptor AR and has undergone clinical trials in prostate cancer. To provide proof-of-mechanism POM in patients, an immunohistochemistry IHC method for determination of AR in circulating tumour cells CTC was developed and validated.

MethodsAfter an assessment of specificity validation focused on intra- and inter-operator reproducibility utilising a novel modification of incurred sample reanalysis ISR. β-Content γ-confidence tolerance intervals BCTI and Cohen’s Kappa κ were employed in statistical analysis of results.

ResultsIn a first set of IHC reproducibility experiments, almost perfect agreement was recorded κ=0.94 when two different operators scored CTC as overall positive or negative for AR. However, BCTI analysis identified a specific bias in scoring staining intensity, where one operator favoured moderate over strong assignments, whereas the reverse was the case with the second operator. After a period of additional training involving deployment of a panel of standardised images, a second set of validation experiments were conducted. These showed correction of the inter-operator bias by BCTI with κ for scoring intensity increasing from 0.59 to 0.81, indicative of almost perfect agreement.

ConclusionsBy application of BCTI to the validation of IHC, operator bias and therefore poor reproducibility can be identified, characterised and corrected to achieve a level of error normally associated with a quantitative biomarker assay, such as an ELISA. The methodological approach described herein can be applied to any generic IHC technique.

KeywordsAZD3514 Immunohistochemistry Method validation Incurred sample reanalysis Cohen’s Kappa β-Content γ-confidence tolerance intervals AbbreviationsARAndrogen receptor

IHCImmunohistochemistry

CTCCirculating tumour cells

BCTIβ-content γ-confidence tolerance intervals

KCohen’s Kappa

CRPCCastration-resistant prostate cancer

POMProof-of-mechanism

ISETIsolation by size of epithelial tumour cells

ISRIncurred sample reanalysis

RECResearch ethics committee.

Electronic supplementary materialThe online version of this article doi:10.1186-1471-2407-14-226 contains supplementary material, which is available to authorized users.

Jeffrey Cummings, Robert Sloane contributed equally to this work.

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Autor: Jeffrey Cummings - Robert Sloane - Karen Morris - Cong Zhou - Matt Lancashire - David Moore - Tony Elliot - Noel Clarke -

Fuente: https://link.springer.com/







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