Culture amplification in human colon adenocarcinoma cell line (CaCo-2) combined with an ELISA as a supplementary assay for accurate diagnosis of rotavirus

Culture amplification in colon adenocarcinoma cell line (CaCo-2) combined with enzyme immunoassay (Pathfinder ELISA) was developed as a supplementary tool for rotavirus diagnosis. One hundred and thirty stools in which results by polyacrylamide gel electrophoresis (PAGE) were in agreement with those obtained by ELISA were amplified in the CaCo-2 cell line. After the first passage 100% specimens were revealed as positive by ELISA. This result was confirmed by PAGE and direct electron microscopy (EM) which increased the rates of rotavirus detection up to 100% after the third and fifth cell passages, respectively. All of the amplified negative stools were confirmed as negative. Among discordant results, three of the eight specimens positive by ELISA but negative by PAGE were confirmed as true positive after the third cell passage. False positive ELISA results could be discarded when the samples were culture amplified and retested by the same ELISA. Using the CaCo-2 amplification-ELISA as supplementary assay, sensitivity and specificity were 1.000 and 0.953 for ELISA and 0.917 and 1.000 for PAGE, respectively. The combined CaCo-2 cell line amplification-immunoassay method proved to be suitable both to evaluate increase in sensitivity of newly developed rotavirus assays and for rotaviral amplification before antigen assays.