AN IMPROVED METHOD FOR SEROLOGICAL DETECTION OF CYMBIDIUM MOSAIC POTEXVIRUS INFECTION IN ORCHIDS

Murine hybridomas secreting monoclonal antibodies to Cymbidium mosaic potexvirus (CyMV) were produced by fusion of FOX-NY myeloma cells with immune splenocytes derived from in vivo or from a combination of in vivo and in vitro immunizations. Forty-five CyMV monoclonal antibodies reacted with the homologous antigen, which was trapped by CyMV rabbit antiserum coated microtiter plates. Twenty-nine of the 45 monoclonal antibodies also reacted with CyMV when the antigen was coated directly on microtiter plates. Detection of CyMV in crude sap of infected orchid leaves by immunosorbent electron microscopy was about twice as sensitive as by enzyme-linked immunosorbent assay (ELISA), whereas the sensitivity of dot blot immunoassay was about eight times that of ELISA. CyMV antigen was detected in direct tissue blots on nitrocellulose membranes in 30 leaf blots of 155 healthy looking orchid plants tested. Of six samples that tested positive by tissue-blot immunoassay, three showed A405nm values of less than 0.05 by ELISA, and three gave ELISA A405nm values between 0.15 and 0.2 when assayed at a 1:20 sap dilution. Ten other positive samples gave ELISA A405nm values between 0.2 and 1.0. The remaining 14 samples, however, had ELISA A405nm values of greater than 1.0. All the plants that tested negative with the tissue-blot immunoassay gave ELISA A405nm values of less than 0.06.