Specialized stains

A combination of specialized staining techniques and high-resolution microscopy has allowed geneticists... 

Mycobacteria are slow-growing organisms in the laboratory, they require special stains, special growth media, and long periods of incubation to isolate and identify... 

In summary - IHC should no longer be regarded a just a special stain but rather as a standardized controlled quantitative tissue based ELISA assay - quantification of IHC is coming, it is just a matter of how and when.11,22... 

Taylor CR, Kledzik G. Immunohistologic techniques in surgical pathology—a spectrum of new special stains. Hum. Pathol. 1981 12 590-596. 

Glyoxal-based fixatives work faster than formalin. Small biopsies may be ready to process after only an hour while properly grossed larger specimens are ready in about 6h. Structural detail is remarkable in its clarity (Fig. 12.9). Red blood cells are lysed, but that rarely presents a problem. Eosinophilic granules are reduced in prominence (see below). Special stains work well, except for tests for iron (the mildly acidic pH is detrimental) and the silver detection methods for Helicobacter pylori. Most notably, glyoxal-fixed tissues retain strong immunoreactivity for most antigens. The chemistry behind most of this is known. 

In addition to the standard set of tissues specified in Table 7.8, observations during the course of the study or in other previous studies may dictate that additional tissues be collected or special examinations (e.g., special stains, polarized light or electron microscopy, immunocytochemistry, or quantitative morphometry) be undertaken to evaluate the relevance of, or understand the mechanisms underlying, certain observations. 

Ambrogi, L. P., ed. (1957) Manual of histologic and special staining techniques. Armed Forces Institute of Pathology, Washington, DC, p. 33. 

Histology. The tissues listed in Table 21.6 plus any lesions, masses, or abnormal tissues are embedded, sectioned, and strained for light microscopy. Parafin embedding and staining with hemotoxylin and eosin are the preferred routine methods, but special stains may be used for particular tissues or for a more specific examination of certain lesions. Electron microscopy may also be used for more specific examination of lesions or cellular changes after their initial localization by more routine methods. 

While it may appear to be a matter of semantics, the author has become convinced that whether IHC is viewed as a stain or as an assay , can play an important role in establishing the proper mind set of the laboratory staff and the pathologists. The IHC method is regarded by many as simply a stain it produces a visible tinctorial reaction within the tissue section. However, IHC should not be regarded as simply another special stain , like a PAS stain or a silver stain. As already noted, IHC is essentially an ELISA method applied to a tissue section. In this respect, when correctly performed, IHC has the potential to perform as a reproducible and quantitative tissue based ELISA assay much more than a simple stain. That the IHC method mostly does not perform to this level, reflects faults in the application of the method, specifically inconsistent sample preparation, lack of reference or calibration standards, and inadequate validation of reagents (7, 8). The use of RTUs does not finally solve these problems, but for reasons that will be discussed below, can lead to increased reproducibility and consistency in a practical... 

The need for deparaffinization of FFPE slides for FISH analysis does not differ from standard preparation of slides for histological staining methods like H E, special stains or IHC. Following mounting by baking, the slides are deparaffinized in xylene (or substitutes) and rehydrated in a series of ethanol. 

After the determination of organ weights and macroscopic examinations the tissues are processed for histo-pathological evaluations. For this they are fixed in formalin or equivalent solutions, or they are frozen (important for the diagnosis of increased fat content in a tissue, e.g. in the liver, organic solvents would dissolved the fat) trimmed to small parts, put into paraffin blocks, cut with a microtome, and stained (hematoxylin-eosin, or special staining for fat and/or collagen etc.). 

S. chartarum. Here, air samples on culture media showed negative results even on cellulose-based nutrient agar, while microscopy yielded 7,500 spores of S. chartarum per cubic meter [20]. Additionally special staining techniques allow differentiation between the sum of all bacteria or mould spores from active ones [21]. 

After more than two decades, advances in IHC have provided a feasible approach to performing immunostain-ing on routinely processed tissues, such that this method is now routine for the performance of IHC special stains in surgical pathology laboratories using EEPE tissues (see Appendix lA). However, demands for improved reproducibility and for quantification have led to a growing recognition that IHC has the potential to be more than just a special stain. If properly controlled in all aspects of its performance, IHC can provide a tissue-based immunoassay with the reproducibility and quantitative characteristics of an ELISA (enzyme-linked immunosorbent assay) test, which not only detects the presence of an analyte (protein or antigen) but also provides an accurate and reliable measure of its relative or real amount (see Quality Control and Standardization section). 

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