Development of monoclonal-antibody-ELISA and -DIP-STICK immunoassays for Penicillium islandicum in rice grains

Monoclonal antibodies (mAbs) were raised against Penicillium islandicum, a fungus commonly isolated from stored rice grains in South-East Asia. Mice were immunized by a direct, simple method; fresh cell-free surface washings from a solid agar slant culture were injected directly into the peritoneum without prior concentration. Hybridoma supernatants were screened by ELISA. Most of the mAbs raised cross-reacted with other storage fungi and/or uninfected rice grains but there were species-specific. One of these, PI01, was used to develop ELISA and DIP-STICK assays for the detection of P. islandicum in individual grains. All inoculated grains and approximately 90% of grains in natural infected samples from Indonesia tested positively, by ELISA, for P. islandicum. This result and those obtained for discoloured grains from both Indonesia and the Philippines, 32% and 14% respectively, are higher than those obtained by direct plating of surface-sterilized grains. Heat and periodate treatment of the PI01 antigen and binding on Western blots indicate that it is a glycoprotein of Mr greater than 90,000. Hyphae of all ages stained uniformly by immunofluorescence using the PI01 antibody but mature conidia stained only weakly.