Tests for the detection of antibodies to Treponema pallidum are recommended for the confirmation of reactive nontreponemal test results and the accurate diagnosis of syphilis. The present-day use of Western blot (immunoblot) technology for the diagnosis of retroviruses prompted the development and evaluation of a Western blot assay with whole-cell T. pallidum as the antigen. The assay detected antibodies in syphilitic serum or plasma from dilutions of specimens incubated overnight with test strips. A test was considered positive when at least three of four major antigens having molecular masses of 15.5, 17, 44.5, and 47 kDa were detected. The Western blot assay had 93.8% sensitivity and 100% specificity for clinically defined samples. The Western blot assay was compared with double-staining fluorescent trepoiemal antibody absorption [FTA-ABS (DS)], which had a sensitivity and a specificity of 91.7 and 92.0%, respectively. Dilution series studies of syphilis-positive specimens indicated that the Western blot assay has an endpoint of reactivity at least 3 to 4 serial dilutions greater than that for FTA-ABS (DS). Overall, the >95% agreement between the Western blot assay and FTA-ABS (DS) for clinically defined specimens indicates that the sensitivity of the Western blot assay is equal to or greater than that of FTA-ABS (DS). The Western blot assay demonstrated no false-positive or equivocal reactivities for nonsyphilitic specimens, including normal specimens (both plasma and serum), biological false-positives, and specimens with elevated gamma globulin or antinuclear antibody. We conclude that the high sensitivity and specificity of the T. pallidum Western blot assay, together with its simplicity and objectivity, make it a good confirmatory test for syphilis.

A paramagnetic microparticle (MP) assay for antibody to hepatitis C virus (anti-HCV) was developed, in which the probe for antibody consisted of synthetic peptides corresponding to HCV capsid and nonstructural c-100 regions, as well as a recombinant protein corresponding to the nonstructural c33c region. Assay performance was evaluated by testing serum from 108 geographically diverse patients with non-A, non-B hepatitis (NANBH). The frequency of anti-HCV reactivity detected with the MP assay and with an enzyme-linked immunosorbent assay (ELISA) for c-100 was 91 and 70 percent, respectively. All c-100 HCV ELISA-reactive specimens also reacted on the MP assay. In addition, anti-HCV seroconversion in three plasma donors was detected one to two blood collection dates earlier by the MP assay than by the c-100 HCV ELISA and at similar blood collection dates by the MP assay and a second-generation anti-HCV ELISA. Serologic responses to the three distinct antigenic regions of HCV in NANBH patients varied: reactivity to all three antigens was most common (49%), reactivity to both capsid and c33c (40%) was next most common, and single-antigen reactivity was rare (4%). MP assay reactivity of 825 volunteer donors was 0.1 percent. These results demonstrate both the utility of additional HCV antigens for an effective anti-HCV screening assay and the application of paramagnetic MP technology to serologic testing for HCV infection.

A new rapid serologic enzyme immunoassay for antibodies to hepatitis C virus (HCV) is described. The assay combines synthetic peptide and recombinant antigens representing putative structural and non structural HCV gene products with paramagnetic microparticle assay (MP assay) technology. Assay readout is based upon an enzymatically generated fluorescent product which is quantified with a novel semi-automated washer/reader instrument system. Assay sensitivity and specificity was determined to be greater than the first generation HCV C-100 EIA using a non-A, non-B hepatitis disease panel, an HCV performance panel, an HCV seroconversion panel, dilutions of HCV reactive sera, and random volunteer blood donor specimens.