Assessment of the specificity of human Iaalloantisera and alloantigens by the use of radioiodinated human Ia antigens

A method for preparing radiolabelled human Ia antigens and a utilization of the labelled antigens in defining the la specificity of antigens and antisera are presented. The standard preparation procedure involves solubilization with Renex 30 (a non-ionic detergent), purification by gel filtration and lectin-affinity chromatography, and radioiodination, and yields an Ia preparation with immunological purity of 48–62%. The basic procedure for defining the Ia specificity of antisera is a direct binding assay: it allows to react a given antiserum with a panel of labelled la preparations of different specificities and measures the extent of binding by the double antibody technique. The major or putative specificity can be deduced from the reaction pattern. The specificity can be further evaluated by a sequential binding test or a binding inhibition test. The Ia tissue typing is performed by the binding inhibition assay: it measures the ability of test tissues to inhibit the binding of a reference labelled Ia preparation with a reference Ia antiserum. This assay permits Ia typing with unseparated blood leukocytes of 2 to 3 × 1057. These radioimmunoassay procedures can offer a solution to problems involved in human Ia serology.