Rapid and simple purification of human immunodeficiency virus type 1 epitope p24 and its use in ELISA

Abstract

In order to develop a reliable and inexpensive serodiagnostic method, gag protein of human immunodeficiency virus type 1 (HIV-1), p24 (HIV-gag; bp 730-1419)was cloned into an expression vector pCTIO with a sequence encoding a hydroxylamine cleavage site and with a part of lacZ gene (JacZ!f\ 834 bp) as a fusion partner. Overexpression of /acZ’-p24 was induced in E, coli and the p24 fusion protein was purified to homogeneity by centrifugation, hydroxyla-mine cleavage, and two steps of column chromatography (ion-exchange chromatography and gel filtration chromatography). Western bloit analysis and enzyme-linked immunosorbant assay (ELISA) using the purified p24 peptide as an antigen showed high sensitivity and specificity for detecting HIV-1 antibodies in testing human sera when used with a previously purified gp41 epitope which we purifled previously. The mixed antigens showed higher sensitivities and specificities on ELISA test than with the gp41 epitope alone [Sohn, M. J., Cho, S. H.， Jang，W. H.，Chong, Y. H” Nham，S. U.，and Lee, Y. I. (1993) J. Virol. Methods 41, 93-100]. These results suggest that simple and rapid purification of recombinant core antigen will provide a valuable resource of HIV-1 serodiagnostics for screening large groups of blood donors to prevent HIV-1 infection.