Comparison of the Binding Affinities of Rabbit IgG Fractions to the Rabbit Fetal Yolk Sac Membrane: Use of 22Na to Facilitate Quantitation of 125I-IgG Binding
Rabbit IgG was purified, Fr-1-(G-200)2, and then separated by DEAE-cellulose column chromatography into three major fractions, Fr-I, -II, and -III. Binding affinities of the 125I-labeled IgG and its fractions to the rabbit fetal yolk sac membrane were studied. At a concentration of 2 mg/ml, rabbit IgG is bound to the extent of 9 µg/cm2 membrane, whereas the values for fractions Fr-I, -II, and -III are 13, 7, and 5 µg/cm2, respectively. The maximal amount of IgG bound appears to be approximately the same, i.e., 23 µg of IgG/cm2 membrane, for Fr-1-(G-200)2 and its three fractions. In contrast, bovine IgG does not bind to the membrane over the range of concentration tested. Binding constants for Fr-1-(G-200)2, fraction Fr-I, -II, and -III are estimated to be 5.4, 8.6, 4.0, and 2.0 × 104 M-1, respectively. The binding affinities of IgG fractions to the yolk sac membrane correlate with the chemical and physicochemical properties of the fractions. Also detailed in the text is the use of 22Na to facilitate quantitation of specific binding of the 125I-IgG to the membrane under equilibrium conditions.