Preparation of Nitrofurazone Metabolite Hapten and Its Applications in Immunoassay
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Abstract:
In the present work,a novel method for preparation of nitro-terminal group-containing SEM hapten was established by a reaction of SEM with 4-(3-aldehyde-4-nitro-phenoxy)-butyric acid s, which introduced a reactive terminal nitro groups into SEM structure. The prepared SEM hapten was then coupled with bovine serum albumin (BSA) and ovalbumin (OVA) respectively to prepare SEM-BSA immunogen and SEM-OVA coating antigen. The BALB/C mice were immunized with immunogen SEM-BSA, and the monoclonal antibodies were prepared by cell cell fusion technology. The titer and specificity of the monoclonal antibodies were evaluated by using indirect competitive enzyme-linked immunoassay (ciELISA). The results showed that the prepared monoclonal antibody could specifically recognize the 2- nitrobenzaldehyde derivates (2-NPSEM) of SEM with a titer of 1:2560000. The established ciELISA method exhibited a good linear relationship in the range of 0.05~4.05 μg/L with a detection limit of 0.04 μg/L and a half maximal inhibitory concentration value (IC50) of 0.23 μg/L, and the cross reaction rates of NPAMOZ, NPAOZ and NPAHD were 0.03%, 0.02% and 0.26%, respectively. This methods for SEM hapten synthesis provided new ideas and ways for the high-efficient detection of nitrofurazone metabolite.
