Development of monoclonal-antibody-ELISA, -DOT-BLOT and -DIP-STICK immunoassays for Humicola lanuginosa in rice.
Monoclonal antibodies (MAbs) were raised against Humicola lanuginosa, a thermophilic, saprophytic fungus that is commonly isolated from freshly harvested rice grains in Indonesia. Mice were immunized by direct injection into the peritoneum, without prior concentration, of fresh cell-free surface washings from a solid agar slant culture. Hybridoma supernatants were screened by ELISA using wells coated with a dilution of the immunogen. From one fusion 403 hybridoma clones were obtained yielding 52 cell lines secreting antibodies positive for H. lanuginosa. Twelve cell lines were re-cloned, grown in bulk and tested against other storage fungi. Most of the MAbs raised were IgM antibodies that cross-reacted with several of the storage fungi and/or uninfected rice grains. However, the IgM antibody EC6 did not recognize antigens from rice grains and cross-reacted strongly with only one other test fungus, Penicillium variabile, and partially with two others. This MAb was used to develop a highly sensitive DIP-STICK immunoassay to detect the fungus in infected grains. These assays are simple to perform, require no equipment and are suitable for field use by untrained workers.
Dewey
F M
FM
Department of Plant Sciences, University of Oxford, UK.
MacDonald
M M
MM
Phillips
S I
SI
D000911:Antibodies, Monoclonal; D000946:Antigens, Fungal / Q000032:analysis; D005506:Food Contamination; D005516:Food Microbiology; D007118:Immunoassay; D008855:Microscopy, Electron, Scanning; D003904:Mitosporic Fungi / Q000276:immunology* / Q000648:ultrastructure; D012275:Oryza / Q000648:ultrastructure
D000911:Antibodies, Monoclonal; D000946:Antigens, Fungal