Staining Techniques and Microscopy

Abstract

The analysis of tissues and cells first obtained at forensic autopsy can present particular technical problems. While all established conventional histological staining methods are used, the immunohistochemical visualization of defined antigens has also made progress in forensic histopathology. Tissue that has frequently already undergone autolytic changes or putrefaction can make both conventional and immunohistochemical staining challenging, if not impossible. It is precisely for this reason that the mastery of staining techniques is essential. In the field of immunohistochemistry, the choice of fixative and the duration of fixation play a role, as does the option to pretreat tissues for antigen demasking, which often involves varying incubation times with primary and secondary antibodies. Other histopathological methods are occasionally used, such as in situ hybridization or apoptotic cell detection using the TUNEL technique. The actual practice of microscopy evaluation requires knowledge of possible distortions and artifacts but also depends on selecting the correct stain or the primary antibodies that are appropriate for the immunohistochemical question at hand. Experience in microscopy can significantly reduce problems such as interobserver variability. It is also important not to underestimate the importance of converting microscopic findings into a written expert appraisal. The qualification and microscopic quantification of defined cell types raises additional issues that, in many cases, can only be answered when one also has good knowledge of the relevant literature and its critical review. Highly specialized microscopy investigations, such as confocal laser scanning microscopy and electron microscopy, are used more rarely. However, histopathological findings must always be classified within the overall context of a “case,” taking other findings and information into consideration.