*Article* **Validation of New ELISA Technique for Detection of Aflatoxin B1 Contamination in Food Products versus HPLC and VICAM**

**Elsayed Hafez 1, Nourhan M. Abd El-Aziz 2, Amira M. G. Darwish 2, Mohamed G. Shehata 2,\*, Amira A. Ibrahim 1, Asmaa M. Elframawy <sup>3</sup> and Ahmed N. Badr <sup>4</sup>**


**Abstract:** AbstractToxin-contaminated foods and beverages are a major source of illness, may cause death, and have a significant negative economic impact worldwide. Aflatoxin B1 (AFB1) is a potent toxin that may induce cancer after chronic low-level exposure. This study developed a quantitative recombinant *AflR* gene antiserum ELISA technique for aflatoxin B1 detection in contaminated food products. Aflatoxin B1 residuals from 36 food samples were analyzed with HPLC and VICAM. DNA was extracted from aflatoxin-contaminated samples and the *AflR* gene amplified using PCR. PCR products were purified and ligated into the pGEM-T vector. Recombinant plasmids were sequenced and transformed into competent *E. coli* (BL21). Molecular size and B-cell epitope prediction for the recombinant protein were assessed. The purified protein was used to induce the production of IgG antibodies in rabbits. Serum IgG was purified and labeled with alkaline phosphatase. Finally, indirect-ELISA was used to test the effectiveness of polyclonal antibodies for detection of aflatoxin B1 in food samples.

**Keywords:** aflatoxin B1; recombinant *AflR* gene; VICAM; HPLC; I-ELISA; peanut; wheat flour; milk powder

**Key Contribution:** The indirect-ELISA technique was compared with HPLC and the VICAM system in food samples. The new technique showed reasonable accuracy; cost- and time-effective detection of aflatoxin B1.
