High-sensitivity rapid detection method for aflatoxin B1 (AFB1) in cassava flour and based products

【Objective】Based on the principle of enzyme-linked immunoassay(ELISA), to establish high-sensitive direct competitive ELISA for detecting aflatoxin B₁(AFB₁) in cassava flour and based products samples, provide technical support for cassava food processing and utilization.【Method】Antigen and antibody were obtained through structural modification of AFB₁ antigen, immunization, cell fusion and others protocol. Then the highly sensitive rapid test was established, and it was carries on to test examples comparison with kits from market for verification for cassava flour and based products sample.【Result】The results showed that AFB₁ antigen and antibody were successfully synthesized, whose antibody subtypes were all IgG₁. One of the most sensitive 3F2 cell line had a cross-reactivity to aflatoxins G₁, G₂, and B₂ for 21.4%, 1.9%, 8.8% respectively, and they had no cross-reactivity to other toxins. The linear range of ELISA was 0.02-0.48 μg/kg, half inhibitory concentration(IC₅₀) was 0.047 μg/kg, the standard equation was y=- 3.074x-4.3066(R²=0.9978). With 35% methanol aqueous solution as standard diluent, cassava food raw materials were extracted with 70% methanol aqueous solution and 0.2 g NaCl, the oil-containing cassava pastries were using 70% methanol aqueous solution as the extract, all samples were 10 times diluted. It also found the detection limit was 0.2-4.8 μg/kg and, the recovery rate of cassava flour and based products samples addition was 81.5%-120.0%, whose coefficient of variation was less than 8.5%. While, the test results showed that, the selfbuilt ELISA kit detected 19 positive samples from 96 samples, and the purchased ELISA kit detected 7 positive samples. The detection results of samples more than 2.0 μg/kg were in complete agreement, self-developed ELISA kit was more sensitive than the ELISA kits from market.【Conclusion】The direct competitive ELISA method established in this study has high sensitivity, simple operation to be widely used for rapid determination for AFB₁ in cassava flour and based products.