Radioimmunoassays for the detection of antibodies to treponemal polypeptide antigens in serum

Abstract

Cross-reacting treponemal antigens are potentially important candidates for serodiagnostic assays in syphilitic infections. Based on the idea that the organelles for locomotion in virulent and avirulent treponemes might be composed of similar subunits, we attempted to purify the flagellar antigens of Treponema phagedenis biotype Reiter and Treponema refringens for use in radioimmunoassays. With a combination of physical and chemical methods, the major protein subunit of purified flagellar preparations exhibited a mass of approximately 37 kilodaltons (kd) on sodium dodecyl sulfate-polyacrylamide gels. These 37-kd materials, with weight estimates comparable to those of other flagellin molecules, were further purified by preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis and electroelution. Human and rabbit sera, alone or subjected to DEAE Affi-Gel blue chromatography, were subsequently tested in radioimmunoassays employing each of the purified preparations. Even though sera from patients with secondary syphilis and from experimentally infected animals at 3 to 4 weeks postinfection were reactive in radioimmunoassays employing the 37-kd flagellar antigens, the assays were relatively insensitive for detection of immunoglobulin G responses in the early stages of human infection. Detection of immunoglobulin G antibodies in sera obtained early in the course of natural or experimental infection was possible with electroeluted 33- to 64-kd materials from both avirulent and virulent treponemes.