Development of a Direct Competitive ELISA Kit for Detecting Deoxynivalenol Contamination in Wheat

Abstract

This study was conducted to develop a self-assembled direct competitive enzyme-linked immunosorbent assay (dcELISA) kit for the detection of deoxynivalenol (DON) in food and feed grains. Based on the preparation of anti-DON monoclonal antibodies, we established a standard curve with dcELISA and optimized the detection conditions. The performance of the kit was evaluated by comparison with high-performance liquid chromatography (HPLC). The minimum detection limit of DON with the kit was 0.62 ng/mL, the linear range was from 1.0 to 113.24 ng/mL and the half-maximal inhibition concentration (IC 50 ) was 6.61 ng/mL in the working buffer; there was a limit of detection (LOD) of 62 ng/g, and the detection range was from 100 to 11324 ng/g in authentic agricultural samples. We examined four samples of wheat bran, wheat flour, corn flour and corn for DON recovery. The average recovery was in the range of 77.1% to 107.0%, and the relative standard deviation (RSD) ranged from 4.2% to 11.9%. In addition, the kit has the advantages of high specificity, good stability, a long effective life and negligible sample matrix interference. Finally, wheat samples from farms in the six provinces of Henan, Anhui, Hebei, Shandong, Jiangsu and Gansu in China were analyzed by the kit. A total of 30 samples were randomly checked (five samples in each province), and the results were in good agreement with the standardized HPLC method. These tests showed that the dcELISA kit had good performance and met relevant technical requirements, and it had the characteristics of accuracy, reliability, convenience and high-throughput screening for DON detection. Therefore, the developed kit is suitable for rapid screening of DON in marketed products.

Rapid, Sensitive On-Site Detection of Deoxynivalenol in Cereals Using Portable and Reusable Evanescent Wave Optofluidic Immunosensor

“…The recoveries of the spiked samples were in the range from 81% to 127%, and the parallel test results show that the relative standard deviation (RSD) ranged from 3.92% to 10.46%. These are similar with the recoveries (77.1–107.0%) and RSD (4.2–11.9%) of the other immunological detection method in wheat [ 32 ]. These results indicate that the proposed method was capable of detecting DON in the real samples with suitable accuracy and precision, and the interference from the real samples was negligible.…”

Development of a Highly Sensitive and Specific Monoclonal Antibody Based on Indirect Competitive Enzyme-Linked Immunosorbent Assay for the Determination of Zearalenone in Food and Feed Samples

“…A seven-point standard curve of the icELISA was drawn under the above optimized conditions by plotting the gradient concentrations (Log C) against the B/B0% values (proportion of mean absorbance value of the standards (B) divided by that of the zero standards (B0) in triplicate experiments). A four-parameter logistic regression equation was deduced, and the IC50 was calculated [ 46 , 47 ].…”

Indirect Competitive ELISA for the Determination of Total Chromium Content in Food, Feed and Environmental Samples

“…The effect of the ionic strength was evaluated by preparing Cr(III) standard solutions and diluting the Cr(III)-EDTA mAb in HEPES-KCl buffer solution (10 mM HEPES and 3 mM KCl), which had been augmented with varying NaCl concentration (0.137, 0.2, 0.4, 0.8, and 1.6 M NaCl). The effect of the HBS buffer solution with different pH values (5.0, 6.0, 7.4, 8.0 and 9.0) was tested and optimized [ 49 , 50 ].…”