The Precipitation Reaction: Experiments on Multiple Zones

Abstract

1. An antiserum containing antibodies for two unrelated antigens, crystalline egg albumin and crystalline horse serum albumin, has been prepared.2. In titrations of the antiserum against mixtures of these antigens it has been possible to produce a single zone or two zones of optimal particulation by varying the relative concentrations of the components in the antigenic mixtures. It is evident, therefore, that whilst (a) a single zone does not necessarily indicate a single antigen-antibody system, (b) multiple zones suggest the presence of more than one antigen-antibody system.

Some observations on the reaction between horse serum and a pool of rabbit antihorse serum. II. An analysis of the antigens concerned in the production of multiple zones

“…The method is based on the identification and correlation of a procedure optima occurring in the parallel titrations of a single antiserum against a mixed antigen and equivalent concentrations of physically or chemically prepared derivatives of it, supplemented by absorption of precipitins at individual and multiple oc procedure optima and retitration of the absorbed sera. In the first place antibody reactions whose optimal proportions are approximately equal, thus confirming the results of Taylor & Adair (1935). However, it seems clear that a single antigen-antibody reaction is manifested by a single zone of rapid particulation, and that the occurrence of two or more zones is due to the use of a mixed system.…”

“…Taylor & Adair (1935) observed that the presence of another antigen-antibody system may cause changes of the proportions of an antigen and antiserum necessary for optimal particulation. Various physical and chemical methods are available for examining the homogeneity of a protein preparation and for defining and differentiating the constituent proteins of a mixture.…”

“…Horse serum is to be regarded as consisting of a number of antigens (Landsteiner, 1946), and Goldsworthy & Rudd concluded that multiple zones of rapid flocculation are due, in part at least, to the independent activity of multiple antigens and their specific antibodies. Taylor & Adair (1935) titrated an antiserum containing antibodies to crystalline egg albumin and crystalline horse serum albumin against mixtures of the two antigens, and found that one or two zones of rapid precipitation appeared, depending on the relative proportions of the two antigens in the mixture. They pointed out that whereas multiple zones suggest the presence of more than one antigenantibody reaction, a single zone does not necessarily indicate a single antigen-antibody system.…”

Observations on the crystallizable albumin fraction of horse serum

“…It will be seen that several of the systems show two clearly marked optimal reacting concentrations of antigen; this may be taken as evidence that, in these at least, more than one antigen-antibody reaction is taking place [cp. Dean et al 1935;Taylor & Adair, 1935;Goldsworthy & Rudd, 1935].…”