Prevention of methodological non-specificity

POase activity in tissue or cell preparations is usually eliminated prior to the test. In some cases, endogenous POase is confined to peroxisomes and does not interfere with the test. Fixation in methanol is often accompanied by a decrease in peroxidatic activity. The method of Straus (1971) using 1% sodium nitroferricyanide and 1% acetic acid in methanol, is effective to abolish endogenous POase, but also weakens the immunohistochemical reaction. Streefkerk (1972) 

Prevention of non-immunological adsorption of immuno-reactants to tissue preparations. The frequent use of cross-linkers both in the preparation of conjugates and fixation of the samples may result in a stickiness of the preparations. Most of these crosslinkers react via amino groups. Molin et al. (1978) reduced background staining by the inclusion of 20 mM lysine, with or without 1% BSA, in the incubation steps. 

In cases where Fc receptors in the tissue are a problem (cells of the lymphoid system, central nervous system and some epithelia) the use of Fab may offer a solution. Pretreatment of the samples with 2% normal serum may also obviate this problem. 

One of the most effective means to reduce background staining is to improve the detection limit of the test, so that antisera may be used at higher dilution. Several suitable procedures are listed in Table 17.8. 

The classical method for the prevention of non-specific adsorption of antibodies to the sample, particularly in the PAP method, is the pretreatment of the tissue preparation with normal serum from the species which produced the antiserum (3% normal serum in the pretreatment, 1% normal serum included in the next steps Stern-berger and Joseph, 1979). Grube and Weber (1980), however, reported that IgG or F(ab )2 from non-immune serum was bound non-specifically to gastrin cells (to unidentified anionic constituents) and 

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