An Enzyme Immunoassay for Neopterin and Biopterin Using a Microtitre Plate Linked with N2-(3-Aminopropyl) Derivatives of Pteridines

Abstract

A novel and sensitive enzyme immunoassay for neopterin and biopterin using a microtitre plate has been developed. The antiserum against neopterin or biopterin was incubated with samples prepared from human urine or rat tissues, and was placed in a well of microtitre plate, where N2-(3-aminopropyl) derivative of neopterin or biopterin was kinked by poly-L-Iysine and glutaraldehyde. The antibody, which did not form the immunocomplex with the hapten in the sample, was bound to the fixed hapten on the plate, and then was measured with anti rabbit IgG and peroxidase anti-peroxidase complex. The detection limits of these pterins were both 0.1 pmol. The specificity of the assay was so high that this assay system with employing anti-neopterin antiserum completely distinguished neopterin from biopterin, as judged from the cross reactivity (0.002%) and vice versa. The amounts of neopterin and biopterin in human urine and rat tissues measured by the present assay showed a good agreement with those determined by high performance liquid chromatography.