Evaluation of micro-ELISA for schistosomiasis japonica using crude egg antigen

Procedures of micro-ELISA for detecting antibody of Schistosoma japonicum infection were improved by using crude egg antigen, peroxidase-labeled antibody and O-phenylenediamine on a micro-ELISA plate (M129A, Dynatech). Reactions were performed with 0.1 ml of reagents in 0.3 ml wells at each step and 0.3 ml of substrate was placed at the final procedure. The endpoint of reaction was defined as the upper limit of 99% critical range of absorbance in negative sera at 1:40 dilution which was approximately twice the absorbance of a pooled negative serum at 1:40. Using this endpoint, appropriate concentrations of antigen and conjugate were determined. Cross-reactions of egg antigen were observed with sera at 1:40 from the infections with other schistosomes, Trichobilharzia brevis, Fasciola hepatica, Echinococcus multilocularis and Trichinella spiralis were diminished at 1:200 serum dilutions except for other schistosomes. Among 177 egg positives, 171 (96.6%) showed the titer of 200 or higher while 67 old cases at Kofu, Japan showed low titers where 22 (31.9%) were lower than 40, 44 (63.7%) were between 40 and 160 and 3 (4.4%) were 320 or higher. The proven non-infected controls of 93 cases from Leyte and Manila, Philippines, Tokyo and Kofu, Japan were all negative. The result of ELISA for schistosomiasis japonica by crude egg antigen was satisfactory after standardization and stabilization of the procedures which were considered to be as important as using defined antigens.