The use of immunological techniques in the study of disease

Traditionally, the basis for this 'investigative immunopathology' is serology. Detection of antibody against a specific pathogen, and of rising (or falling) titers of antibody, has long been the mainstay of the approach. The concept of autoimmunity is heavily based upon the identification of circulating antibodies directed against intrinsic antigens, and there is also reciprocal serology, based upon the use of antisera to detect a given antigen.

There are now many ways to detect and amplify the signal given in serology-based assays. The use of complement fixation to detect the formation of complexes was an early example; and there are now a host of other similar procedures, many of which rely on a second superimposed antigen-antibody reaction, or upon a combination of antibody with biotin, which can then be identified using an avidin-coupled detection system. Enzyme-linked antibody in ELISA assays has been used either alone or in conjunction with biotin-avidin and has proven a useful and efficient alternative to isotope. The most significant development in serology-based studies has undoubtedly been the advent of monoclonal antibodies, which have made a considerable impact upon the specificity that can be achieved.

In a sense, most of the other techniques that are used in investigative immunopathology are extensions of the serological approach. These methods have been used to purify proteins by affinity, and have also been used in more sensitive detection systems such as western blotting. Furthermore, serology is also the basis for studies that are orientated toward cells and tissues. Thus antibodies are used as the detection system for different cell types, both for the immune cells themselves (such as the lymphocyte subsets) and for other cells in epithelial and connective tissues.

Immunofluorescence, where an antibody is coupled to a fluorescent dye can be used in several different ways: for fluorocytometry (including preparative sorting), for detection of antibodies directed against cellular targets in autoimmunity, and for both detection and isotyping of immune complex deposits. In histological examination of tissues, immunostaining methods, where the detection is via either peroxidase or alkaline phosphatase, have formed the basis for re-evaluation of previous tumor classification, and have made quantitative analysis possible. There are difficulties in this approach, in particular those related to the preservation of the antigen (in terms of accessibility, structure and configuration) under fixation conditions; but there-are also unique advantages, since immunopathologic methods can be used in this context not only as cell detectors, but also to monitor cell function, where the antibody may be directed against cell cycle-related proteins, or against growth factors and their receptors.

Recently, immunopathology has also provided powerful adjunct techniques for molecular pathology, where the in vitro translation products of specific genes have been characterized using monoclonal antibodies; and, conversely, expression vector libraries have been generated which can be used to screen for identification of the gene which encodes for a particular protein.

However, investigative immunopathology is not confined to the study of antibody. Analysis of the complement components per se may also be used to monitor disease, either involving the complement system itself or as a marker of complex formation. Quantitative cellular studies are useful, and thus far have proven more reproducible than any of the functional immunopathological assays (such as phagocytosis, chemotaxis, natural killer activity, cytotoxicity, and alloresponses) that have been used as research tools to examine disease. Strictly, blood group serology and HLA typing are not classifiable as immunopathologic methods, since they document physiological markers rather than disease states, but both have been used to determine disease associations.

This armamentarium of procedures has provided the opportunity for investigating disease in many different ways, some of which have been mentioned already. It is probably true today that even if at a superficial level there is no obvious connection between a particular disease process and the immune system, in every instance there will be at least one investigative procedure which falls within the remit of immunopathological analysis.

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