Radioimmunoassays Employing Double Antibody Techniques

Abstract

Double antibody methods are applicable to all radioimmunoassay systems. At least three approaches exist. The first, in which immunoprecipitating serum is used to precipitate soluble antigen-antibody complexes, is the most commonly used. Since the effects of interfering factors, especially those present in serum, become minimal as equilibrium is approached, care must be taken to insure that the period of incubation is of sufficient length for the immunoprecipitation reaction to reach equilibrium. In general, all efforts should focus on achieving reaction conditions which maximize stability and optimize immunoprecipitation. A second approach involves use of the first antiserum in an insolubilized form after prior precipitation with the immunoprecipitating antiserum. This approach is attractive since most non specific factors cannot interfere with the immunoprecipitation step to a significant extent once the formation of antigen-antibody aggregates is well underway. The third approach, in which immunoprecipitating antiserum is conjugated to a solid matrix, in principle represents a more ideal method of separating bound hormone from free. If this approach is ever to be utilized, a method of conjugating antiserum to a suitable matrix such that the antibody retains most of its immunoreactivity must be devised.

Radioimmunoassays for Human Chorionic Gonadotrophin and Human Hypophysial LH Using the Double Antibody Method

“…It indicates the low degree of reversibility of HCGantiHCG reaction involved in such a radioimmunoassay. Similar observa tions were reported for insulin [7] and gonadotrophins [9,16]. However, for gastrin [4] and oxytocin [2], one could not find such significant differences between equilibrium and non-equilibrium conditions.…”

Synthesis and Secretion of Immunoglobulin, Including IgA, by Human Tonsil and Adenoid Tissue Cultured <i>in vitro</i>

“…IgA was measured by an inhibition assay similar to that described by ReesMidgley et al [14]. Rabbit anti-IgA was diluted in BBS containing 100/ig N/ml of normal rabbit /-globulin (RGG).…”

Response Patterns to <i>DL</i>-Carnitine in Patients on Maintenance Haemodialysis

“…Plasma free fatty acid (FFA) concentrations were determined colorimetrically [9], with the slight modification that a 5% diphenyl-carbazone in diphenyl-carbazide was used as the colour reagent [13]. Plasma immunoreactive insulin levels were measured by the double antibody technique [19] with a commercial kit (Radiochemical Centre. Amersham, England).…”