Evaluation of the latency-associated nuclear antigen (ORF73) of kaposi's sarcoma-associated herpesvirus by peptide mapping and bacterially expressed recombinant western blot assay

Kaposi's sarcoma (KS)-associated herpesvirus open-reading frame (ORF) 73 en codes a latency-associated nuclear antigen (LANA) that is the basis for sev eral serologic assays. Immunoreactive epitopes were searched for by peptide mapping, and 171 cleavable, biotinylated 17-mer peptides offset by 5 resid ues were synthesized and screened with human serum samples by ELISA, The in itial screen, which used highly reactive serum diluted 1:500, identified 38 immunoreactive peptides, These were subsequently tested on additional seru m samples diluted 1:40. Thirteen peptides were more reactive with serum sam ples from patients with KS than with control serum samples. No single epito pe was recognized by most KS patient serum samples. Combined use of these p eptides did not increase test sensitivity to that of current indirect immun ofluorescence assays for LANA (80%-90%). For comparison, full-length ORF73 was expressed in bacteria and analyzed by Western blot. The overall sensiti vity was 67% (range, 100% among US patients with classic KS to 52% among It alian patients with classic KS). These studies suggest that LANA immunoreac tivity may be due to variations in patient response or conformational epito pes.