Structural Insights Into the Mechanisms of Recognition of Recombinant Full-Length Antibodies for the Detection of Staphylococcus aureus Enterotoxins C1, C2, and C3

作者：Chunhao Wei, Aihong Wu, Xinxin Xu, Aihua Qu, Chuanlai Xu, Hua Kuang · 发表于：Analytical Chemistry · 年份：2025 · DOI：10.1021/acs.analchem.4c06364 · 被引用次数：9 · 研究领域：Monoclonal and Polyclonal Antibodies Research、Biosensors and Analytical Detection、Antimicrobial Resistance in Staphylococcus

) and its enterotoxins has become a global food safety issue. Herein, three types of Staphylococcal enterotoxins (SE), including SEC1, SEC2, and SEC3, were successfully expressed and immunized in mice to prepare monoclonal antibodies (mAbs). We screened a pair of mAbs 16E12-9B7 from 12 strains that could simultaneously recognize SEC1, SEC2, and SEC3. Furthermore, the genes from hybridoma cells 9B7 and 16E12 were extracted, amplified, and inserted into expression vectors to obtain recombinant antibodies (rAbs), whose affinities were consistent with those of ascites antibodies. The paired rAbs 16E12-9B7 were applied to a gold immunochromatographic strip (GICS) system to enable the rapid detection of SEC1, SEC2, and SEC3 with visual limits of detection (vLOD) of 2, 0.5, and 2 ng/mL in milk samples. Noticeably, we found that hydrogen bonds and salt bridges played a significant role in these antigen-antibody interactions. The key sites for 9B7 were ASN28 and SER31 in complementarity determining region (CDR) and the key sites for 16E12 were SER32 in the 16E12 variable light chain (VL) and ARG100, SER101, and TYR102 in the 16E12 variable heavy chain (VH). The analysis of key rAbs sites has potential in the screening of mutant antibodies.