Monoclonal antibodies to VRV-PL-VIIIa, a basic multitoxic phospholipase A2 from Vipera russelli venom.
VRV-PL-VIIIa, the most basic phospholipase A2 (PLA2) from the venom of Vipera russelli, induces multiple toxic effects, including neurotoxicity, myotoxicity, edema and hemorrhage. Rabbit polyclonal anti-serum was raised against VRV-PL-VIIIa. The antiserum cross-reacted in enzyme-linked immunosorbant assay (ELISA) with two other PLA2 from the same venom, VRV-PL-V and VRV-PL-VI, and with ammodytoxin A, caudoxin and crotoxin. Twenty-two hybridoma cell lines secreting monoclonal antibodies against VRV-PL-VIIIa were isolated. The monoclonal antibodies exhibited apparent binding affinities in ELISA with VRV-PL-VIIIa ranging over two orders of magnitude. Most of the monoclonal antibodies cross-reacted moderately with VRV-PL-V and weakly with VRV-PL-VI. None of the antibodies cross-reacted with ammodytoxin, caudoxin or crotoxin. Reducing the disulfide bonds of VRV-PL-VIIIa lowered the ELISA signals of each monoclonal antibody to nonspecific levels, suggesting that all the antibodies recognize conformational epitopes. Four of the 22 antibodies neutralized the enzymatic activity of VRV-PL-VIIIa. Interestingly, two of the four exhibited the lowest affinities of the monoclonal antibody library for VRV-PL-VIIIa in ELISA, while the other two exhibited the highest. Each of the monoclonal antibodies was biotinylated and spatial binding relationships were evaluated by competition ELISA.
Gowda
T V
TV
Toxinology Division, United States Army Medical Research Institute of Infectious Diseases, Frederick, MD 21702-5011.
Middlebrook
J L
JL
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000918:Antibody Specificity; D001667:Binding, Competitive; D003429:Cross Reactions; D003439:Crotoxin / Q000276:immunology / Q000378:metabolism / Q000633:toxicity; D004220:Disulfides / Q000378:metabolism; D004487:Edema / Q000139:chemically induced; D004797:Enzyme-Linked Immunosorbent Assay; D054501:Group II Phospholipases A2; D006470:Hemorrhage / Q000139:chemically induced; D007106:Immune Sera / Q000276:immunology; D007114:Immunization; D008297:Male; D009498:Neurotoxins / Q000633:toxicity; D010741:Phospholipases A / Q000276:immunology* / Q000378:metabolism / Q000633:toxicity; D054467:Phospholipases A2; D011817:Rabbits; D030162:Reptilian Proteins; D017840:Daboia; D014757:Viper Venoms / Q000201:enzymology* / Q000276:immunology / Q000378:metabolism / Q000633:toxicity
D000911:Antibodies, Monoclonal; D004220:Disulfides; D007106:Immune Sera; D009498:Neurotoxins; D030162:Reptilian Proteins; D014757:Viper Venoms; D003439:Crotoxin; C068117:ammodytoxin A; D010741:Phospholipases A; D054501:Group II Phospholipases A2; D054467:Phospholipases A2; C034914:caudoxin