Development of botulinum toxin detection assay by antibody capture ELISA
Keywords:botulinum neurotoxin, Clostridium botulnum, ELISA
Clostridium botulinum is an anaerobic bacteria, gram-positive rod-shaped spore-forming with the ability to produce the neurotoxin botulinum. The toxins are divided into 8 types (A-H).The typing identification of C. botulinum toxin is testing by injection in mice. The alternative toxin detection to reduce the use of laboratory animals is enzyme-linked immunosorbent assay (ELISA) and immunochromatography (IC). The study tested the appropriate concentration of the primary antibody and a secondary antibody were coating plate in the Capture ELISA of 4 types including A, B, E and F. Four type foods (bamboo shoot, fermented soil bean, fermented crab and orange juices) spiked with 4 types botulinum neurotoxin were used for quantitative toxin detection assay. The results showed that type A, B and E were used goat antibody as primary antibody and rabbit complex as secondary antibody. Both antibodies were used concentration type A, B and E as 0.5, 0.5 and 0.5 ug/ml, respectively. For type F was used rabbit complex antibody as primary antibody and goat antibody as secondary antibody. Both antibodies were used concentration type F as 1 ug/ml. The least concentration detection toxin type A, B, E and F were 1, 10, 5 and 1 ng/ml., respectively. The botulinum neurotoxin (A, B, E and F) were detected in spiked 4 type foods (bamboo shoot, fermented soil bean, fermented crab and orange juices) at least about 10-50 ng/ml. The results of this research were developed assays for detection botulinum neurotoxin by ELISA for testing food samples and drinking water. The diagnostic kit is suitable for use in the country and reduces the cost of imports for botulinum neurotoxin test kits from abroad.