Document Type: Research Paper
Department of Pathobiology, Faculty of Veterinary Medicine, Shahid Chamran University of Ahvaz, Iran.
Graduate Student of Veterinary Medicine, Shahid Chamran University of Ahvaz, Iran.
Department of Basic sciences, Faculty of Veterinary Medicine, Shahid Chamran University of Ahvaz, Iran.
Antibody based methods have numerous advantages compared to other detection methods. However, heavy metals cannot stimulate the immune antibody response; it is the main obstacle for preparation of the antibodies that used in detection methods of the metals. The production of the immunogenic Pb complexes by using cost benefit linkers of EDTA and DTPA were investigated in this study.BSA molecules were functionalized using EDTA and DTPA linkers in various BSA/linker ratio, pH, incubation times and buffers. The complexes were formed after addition of the Pb to the [BSA-linker]. Different concentrations of the glutaraldehyde were mixed with the formed complexes and shaken at room temperature for 12 hours. The prepared conjugates were
dialyzed in phosphate buffer saline for 72 hours. Stimulation of the mice antibody responses against the prepared [Pb-linker-BSA] complex were evaluated using ELISA. In optimized condition, the DTPA linker coupled Pb to each BSA molecules 2 times higher than EDTA. The optimum pH was 9.6 in EDTA binding to BSA; also, EDTA molarity was 25 times higher than Pb molarity in [PbEDTA-BSA] complexes.These conditions for DTPA conjugation to Pb and BSA are pH 7.4-9.6 and 9.6 respectively. Also, optimum molarity of DTPA was 4 times higher than Pb in [Pb-DTPABSA] complexes. Also, the higher antibody responses against Pb were stimulated by immunization with an antigenic complex that has more solubility and Pb coupled to each albumin molecules. In conclusion, in optimized conditions, DTPA more efficient than EDTA for synthesizes of the immunogenic Pb complex.