Eu 3+ /Sm 3+ dual‐label time‐resolved fluoroimmunoassay for measurement of hepatitis C virus antibodies

Aim To develop a new immunoassay based on the time‐resolved fluorescence immunoassay ( TRFIA ) system for the simultaneous measurement of IgM and IgG antibodies to HCV . Methods Coated recombinant HCV antigens and Eu 3+ ‐labeled IgM and Sm 3+ ‐labeled IgG antibodies were prepared. HCV ‐IgM/IgG TRFIA was established and optimized, followed by a methodological assessment. Data were expressed asx ¯ + SD and analyzed using the SPSS 13.0 software. The percentile method was used to calculate cutoff values. Results The detection sensitivities of HCV ‐IgM and HCV ‐IgG were 0.06 S/ CO and 0.15 S/ CO , respectively. There was a good linear response from 1:40 to 1:40 960 for HCV ‐IgM and 1:20 to 1:40 960 for HCV ‐IgG, when samples strongly positive for HCV ‐IgM and HCV ‐IgG were serially diluted from 1:10 to 1:81 920. The average intra‐assay coefficients of variation ( CV ) for HCV ‐IgM and ‐IgG were 3.45% and 3.71% and the inter‐assay coefficients of variation ( CV ) were 6.49% and 6.79%, respectively. When HCV ‐negative/positive sera were tested by ELISA and using established kits, the negative, positive, and total conformity rates for HCV ‐IgM were 93.3% (28/30), 100% (25/25), and 96.4% (53/55), and those for HCV ‐IgG were 93.3% (28/30), 100% (35/35), and 96.9% (63/65), respectively. Additionally, the established kit exhibited good stability, with declines in fluorescence values to 11.1% and 9.5%, respectively, after storage at 37°C for 7 days. Conclusion We established a dual‐label HCV ‐IgM/IgG TRFIA assay with a wide detection range, high specificity, high sensitivity, good stability, and good clinical value for the simultaneous measurement of HCV ‐IgM and HCV ‐IgG titers in a single test.