Precipitin assays

If an antigen-antibody complex has to be precipitated by a secondary antibody (e.g., antigen bound by mAb precipitated by goat anti-(mouse-IgG) antibody), this range of equivalence must be known. For estimation of the range of equivalence the precipitin assay is used. 

Fig. 4. Detection of immune complexes in supemates from a precipitin curve. Lower panel total protein precipitated by increasing amounts of keyhole limpet hemocyanin (KLH) added to a human anti-KLH serum. Upper panel supemates were analyzed for immune complexes by the conglutinin ( ), monoclonal rheumatoid factor (A), Raji ( ), and staphylococci binding (B) assay, and results expressed as pg/ml aggregated IgG equivalents per ml. Clq binding assay (O) results are expressed as percent Clq precipitated. (Reproduced with permission, J. S. McDougal, M. Hubbard, P. I. Strobel, and F. C. McDuffie, J. Lab. Clin. Med. 100, 705-719, 1982.)...

G. Schiffman, E. A. Kabat, and W. Thompson, Biochemistry 3, 113 (1964). losa Solubility refers to the effects of carrying out the quantitative precipitin assays with the same amounts of antigen and antibody but varying the total volume with saline. 

Structural Insights from Quantitative Precipitin Inhibition Assays... 

The quality control of immunoglobulins includes potency tests and conventional tests for safety and sterility. The potency tests consist of toxin or virus neutralization tests that parallel those used for the potency assay of immune sera, except that for in-process control of some immunoglobulins wider use is made of in vitro assays. In addition to the safety and sterility tests, total protein is determined by nitrogen estimations, the protein composition by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and molecular size by high performance liquid chromatography. The presence of immunoglobulins derived from species other than humans is excluded by precipitin tests. Table 23.4... 

Tg antibodies are directed against the Tg protein, a major constituent of thyroid colloid. Several different techniques have been used to detect and quantify TgAb in peripheral blood. These include passive hemagglutination, the agar gel diffusion precipitin technique, immunofluorescence of tissue sections, enzyme-linked immunosorbent assay (ELISA), radioassay techniques, and chemiluminescence-based immunometric assays. 

Results from Butler et al. (1978). Values for the ElA are based on a standard rat-anti DNP pool ( = 0.82 mg/ml) which was measured by the quantitative precipitin assay. Amplified ElA is an ElA test in which immunologically conjugated enzyme is used (see Section 14.2.1.3). 

Immunolectrophoretic Analysis. The iiranunoelectrophoretic studies were carried out as described by Donini et al. (1964a). Unadsorbed and adsorbed anti-HMG sera were used. The absorption of the antiserum was performed with both HCG (3000 i.u./mg) and Kaolin-acetone extract of children urine, according to the method of Tamada et al. (1967). Any fraction obtained by the last step of purification did not show precipitin line when assayed with antiserum raised against human albumin. 

Assays for Serum Antibodies - Serum antibodies appear to be present in rather low concentrations in drug hypersensitivity in man. For example, in penicillin hypersensitivity, the serum concentrations of BPO-specific antibody protein is under 20-30 pg per ml. and most commonly under 5 pg/ml. Thus the quantitative precipitin analysis, which cannot accurately assay less than about 50 pg/ml, cannot generally be used for drug hypersensitivity. However,there are rare reports of precipitating antibody in drug allergy. 

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