Effect of Formalin Tissue Fixation and Processing on Immunohistochemistry

Abstract

Although immunohistochemistry is routinely performed by many pathology laboratories, its standardization still lags behind. A major cause of variation in the reproducibility of immunohistochemical staining is induced by tissue fixation and, to a lesser degree, tissue processing. This report, stemming from the first meeting of the International Consensus Group on Standardization and Quality Control (ICGSQC) in Nice, France, summarizes the problem and suggests solutions to begin to achieve standardization of fixation and processing. Most laboratories use neutral-buffered formalin (10%) for tissue fixation which introduces cross-links, whereas coagulative fixatives are less popular. Problems with formalin fixation comprise delay of fixation and variations in the duration of the fixation mainly. Solutions to these problems could be to start fixation soon (<30 min) after surgical removal of the tissue and to avoid overfixation (>24-48 hrs). For tissue processing, the most important problem is inadequate tissue dehydration prior to paraffin embedding. This can be prevented by preparing all solutions freshly every week, depending on the volume of tissue processed. If consistently applied, these procedures could eliminate some of the sources of variation in immunohistochemical stains.

Delay to formalin fixation effect on breast biomarkers

“…[32][33][34] Moreover, proteolysis often causes nonspecific binding to unrelated molecules. 35 These factors are most likely the reason behind decreased estrogen and progesterone receptor expression with tumor ischemia. In our study, the estrogen receptor started to decline at a 2 h mark and progesterone receptor at a 1 h mark.…”

A Comparison Study of Histochemical Staining of Various Tissues after Carnoy’s Versus after Formalin Fixation

“…When compared to neutral buffered formalin fixation, RNA was found to be easily extractable from Carnoy's-fixed mouse liver and well preserved with only a slight degradation of high-molecular weight RNA [7]. Although, formalin has better properties on tissues, but its use have many disadvantages, such as health hazards, deterioration of nuclei acids and cross-linking of proteins [14,15,16]. Formalin gives off an unpleasant vapor causes irritation to the respiratory system and eyes [17].…”

Diagnóstico imuno-histopatológico da paratuberculose subclínica em bovinos no estado do Rio de Janeiro

“…O formol é um fixador de baixo custo que inibe a autólise, preserva a arquitetura do tecido e destrói a maioria dos agentes infecciosos. No entanto, uma das suas principais desvantagens é a formação de "cross--linking" das cadeias de aminoácidos (Eltoum et al 2001, Ramos-Vara 2005, ou seja, promove um "entrelaçamento" das moléculas de proteínas, o que pode alterar, fragmentar ou "esconder" os epítopos e inibir as reações no teste de IHQ (Werner et al 2000, Ramos-Vara 2005. Neste estudo, apesar das reativações antigênicas realizadas para induzir a exposição dos epítopos e otimizar a marcação (Shi et al 1998, Ramos-Vara 2005, não foram observados resultados positivos na maioria dos fragmentos de intestino delgado e de linfonodos mesentéricos com infiltração granulomatosa.…”