Quantification of immunohistochemistry—issues concerning methods, utility and semiquantitative assessment I

Abstract

Immunohistochemistry is no longer a technique used only for research but is employed increasingly for diagnosis and for the assessment of therapeutic biomarkers. The latter, in particular, often require a semiquantitative evaluation of the extent of their presence. There are many factors that can affect this that relate to the method: fixation of tissue, duration and type of antigen retrieval, antibody specificity, antibody dilution and detection systems. Other complexities relate to assessment. Different scoring systems are used for either the same or different antigens. Cut-off levels for assessing whether a tissue is 'positive' or 'negative' can vary for the same antigen. Whilst there are quality assurance schemes for the methodology that have improved standards of staining, there are no similar schemes that relate to interpretation, although errors here can create as many problems. There have been improvements in automated analysis but availability is limited and it is still predominantly a research tool. In order for quantification of immunohistochemistry to be a reliable and reputable tool, there must be easy to use, reproducible, standardized protocols for assessment which are international. Improvements in automated analysis with wider applicability could lead to standardization.

A simple and reproducible scoring system for EGFR in colorectal cancer: application to prognosis and prediction of response to preoperative brachytherapy

“…Several reasons have been suggested for this discrepancy such as non-comparable study populations (Spano et al, 2005a), variability in protocols, fixation and antibodies (Atkins et al, 2004) and the lack of a uniform scoring system (Penault-Llorca et al, 2005;Italiano, 2006;Walker, 2006).…”

Reclassification of tumour origin in resected periampullary adenocarcinomas reveals underestimation of distal bile duct cancer

“…In the study of Juuti 102 , more than 30 years old specimens were included. It is well known that for immunohistochemical staining protocol, aging of the fixed tumour tissue might interfere with staining process 168 . Moreover, the variations in fixation protocols over time might also result in inadequate staining.…”

DNA Aneuploidy in Malignant Salivary Gland Neoplasms is Independent of USP44 Protein Expression

“…Positive controls consisted of normal oral mucosa and negative reaction control was carried out by omitting the primary antibody. Immunohistochemical positivity was evaluated in whole tumor sections and the percentage of positivity was graded as follows: 0-25%; 25-50%; 50-75%; and >75%, regardless of intensity (12). Tumor sections were evaluated independently by two pathologists (VFB and CCG) and they revised together the cases whose results were inconsistent.…”