RECOMBINANT SINGLE-CHAIN ANTIBODY PEPTIDE CONJUGATES EXPRESSED IN ESCHERICHIA-COLI FOR THE RAPID DIAGNOSIS OF HIV

Recombinant single chain Fv (scFv) antibody fragments can form the bas is of a rapid, whole-brood diagnostic assay. The scFv described in thi s study is derived from a monoclonal antibody which has a high affinit y for grycophorin A, an abundant glycoprotein on the human red brood c ell membrane surface. The prototype reagent built around the scFv was designed to detect, in whole blood samples, the presence of antibodies that have arisen through infection with a foreign organism such as hu man immunodeficiency virus. The scFv was composed of the antibody heav y-chain variable domain (V-h) joined by a 15 residue linker -(GGGGS)(3 -) to the light-chain variable domain (V-1) terminated by either a C-t erminal octapeptide tail (FLAG) or a 35 amino acid segment from the gp 41 surface glycoprotein of HIV-1. Constructs were cloned into a Escher ichia coli expression vector, pHFA, and expressed in a soluble form in to culture supernatant. The product retained anti-glycophorin activity which could be detected directly in culture supernatants by ELISA. Fu rthermore, the scFv-epitope fusion functioned efficiently in the whole blood agglutination assay and was able to distinguish between HIV-1 p ositive and negative sera.