An improved method for home-made one step HBsAg ELISA reagents and its confirmatory test reagents

Objective To improve the sensitivity and precision of home-made one step hepatitis B surface antigen(HBsAg) enzyme-linked immunosorbent assay(ELISA) reagent by prolonging the enzyme reaction time.Methods The enzyme reaction time of home-made one step HBsAg ELISA reagent was prolonged from 30 min to 120 min,and the change of sensitivity and specificity as well as the influence of relative HBsAg confirmatory test reagent were observed.Murex confirmatory test was run in parallel to verify the sensitivity and specificity of the improved method.Results The prolongation of the enzyme reaction time from 30 min to 120 min did not alter the cut-off value of the HBsAg ELISA test.HBsAg negative samples primarily tested by Murex HBsAg test also showed negative results.The prolongation of the enzyme reaction time to 120 min increased the absorbance value(A).The A value was almost doubled for the samples which were dropped in the grey zone.The result of the clinical trial of improved 120 min home-made confirmatory test comparatively with Murex confirmatory test revealed that among the samples of s/co value from 0.5 to 1.0 by routine HBsAg ELISA,4 out of 22 samples were confirmed as positive by the improved home-made test and the Murex test.Among the samples of s/co value from 1.0 to 2.0 in the positive grey zone,50 out of 79 samples were confirmed as positive by the improved for prolonging 120 min home-made test and Murex test.There was no statistical significance between the two tests(χ 2 =0.078,P0.05).Conclusions Prolonging the enzyme reaction time from 30 min to 120 min can improve the sensitivity of home-made one step HBsAg ELISA reagent,and is also suitable to the relative confirmatory test.