Development Of Ultrasenitive Immunoassay For Detection Of Deoxynivalenol

Deoxynivalenol (DON) is a mycotoxin commonly found in corn, wheat, bread, barley, as well as other most important sources of human intake that have been infected by the mold Fusarium graminearum. Contamination of foods by DON is a potential hazard for livestock and human beings. It is therefore of utmost importance to assay DON contamination.This main purpose of this thesis is to develop a DON-TRFIA and a DON/ZEN-TRFIA for simultaneous detection of DON and ZEN. DON-TRFIA was based on a indirect TRFIA format, deoxynivalenol–bovineserum albumin conjugate was coated onto a 96–well microplate and incubated with standard DON and anti–DON antibody, and then a goat anti–rabbit IgG-Eu3+ conjugate was used to enable detection. The practical working concentration range was from 0.01 to 100ng/mL and the sensitivity for detection was 0.01ng/mL. The shelf life of reagents on the assay was over 6 months at 4℃. The recoveries of detection for DON spiked in corn, wheat, bread and beer were 70-130%. The results obtained by the TRFIA and ELISA correlated well and the coefficient of correlation is 0.9669. These results suggest that the DON-TRFIA is a good method with high sensitivity for quantitative analyzing DON in cereal. Then a TRFIA for simultaneously detection of DON and ZEN contamination was developed, with Eu3+ and Sm3+-chelates labeling goat anti–rabbit IgG and goat anti–mouse IgG, respectively. The measurement ranges of DON were 0.194~100 ng/mL and that of ZEN were 0.73~50ng/mL. The results obtained by the dual-label assay agreed well with those by single label TRFIA-DON and TRFIA-ZEN. The correlation ratios of DON and ZEN with dual and single label TRFIA were 0.9719 and 0.9645, respectively.Besides those we also initially established a homogenously indirect competitive LICLIA for the detection of DON and concisely examined constancy and recovery of the assay. This immunoassay only took 30min to complete the detection. Results showed that the detection limit of the assay was 0.007ng/mL for homogenously indirect competitive LICLIA at a practical working concentration range from 0.007 to 100ng/mL.