Rapid and sensitive heterologous enzyme immunoassays for densonucleosis virus (Parvoviridae)

There exists a serious lack of rapid and sensitive methods to identify densonucleosis viruses and to discriminate among them. Two different enzyme-linked immunosorbent assays (ELISA) were adapted for this purpose, which were both significantly faster and more sensitive than currently used ELISA procedures. This increase in sensitivity was due to an improvement in the conjugation procedure of peroxidase to antibody, the establishment of the optimum conditions for the various incubations, an optimisation of the substrate (H2O2) concentration, and the use of a new H-donor. The speed of the assay could be considerably shortened by the use of polyethylene glycol-6000 (i.e. the total time of the assay needed for maximum sensitivity of the indirect assay was only 2 hours). The assays using peroxidase conjugates were found considerably more sensitive than those using alkaline phosphatase, which is very probably due to a more efficient and better controlled conjugation procedure for peroxidase. The virus could be detected in the pg to ng range in a large excess of nonspecific antigens and titers for the antisera usually exceeded 10(6). Small differences in the viruses could be detected. Several factors, which may influence the sensitivity and specificity of these densonucleosis virus assays, were further investigated.