Development and Preliminary Application of Monoclonal Antibody against Heavy Metal Cadmium Ion

Artificial hapten Cd-ITCBE was synthesized using isotrhiocyanobenzyl-EDTA(ITCBE) to chelate Cd²⁺. The isothiocyanate method was used to conjugate Cd-ITCBE to bovine serum albumin (BSA) and artificial immunogen Cd-ITCBE-BSA was obtained, the detection antigen Cd-ITCBEOVA was obtained in the same way. Ultraviolet (UV), sodium dodecyl sulfate polyacrylamide gel electropheresis (SDS-PAGE) and inductively coupled plasma atomic emission spectroscopy (ICP-AES) were used to identify Cd-ITCBE-BSA and Cd-ITCBE-OVA. Balb/C mice were immunized with Cd-ITCBEBSA and hybridoma lines that secreted Cd²⁺ mAb were generated by cell fusion and then screened by indirect ELISA, blocking ELISA. The Cd²⁺-Kit was established with Cd²⁺ mAb and its traits were tested. The Results showed that Cd-ITCBEBSA was synthesized successfully and its concentration of BSA and Cd²⁺ was 7.1 mg/mL and 191.7 μg/mL respectively. Four hybridoma lines of 1A3, 2B7, 2E10, 4F3 were screened out and the best one was 2E10 and its affinity constant(Ka) was 7.58×10⁸ L/moL. The calibration curve of Cd²⁺-Kit was typical sigmoid curve fitted to the four parameter logistic equation with the IC₅₀ of 16.3 μg/L, the detection limit of 3 μg/L. The recoveries of Cd²⁺ spiked in water were 89.28%, in soil were 94.28% and in feed were 92.88% and the coefficient variation were below 15%. The specificity of Cd²⁺-Kit had 96.5% cross reactivity with Hg²⁺ and little or no cross-reactivity with other compounds. The validity of Cd²⁺-Kit in 4°C was above 180 d.