Technology Study On Colloidal Gold-immunochroma-togrphy Assay Strip For Detection Of Clenbuterol

With the development of immunization technique and the emerging food security issues, gold immunochromatography assay for its particular competitive advantages is playing an increasingly important role in the clinical, field testing, family self-test, and so on. However, there exists a considerable technical gap between domestic and foreign. This article aims to establish R & D technology platform of it from antigen design to sensitivity adjusting through the technological study of clenbuterol strip.The first step is antigen design according to the molecular characteristics of clenbuterol. Complete antigen used in the current commercial products was mostly synthesized by diazotization-coupling method(OVA-GA-CL, coupling ratio of 25.2:1). But it is synthesized by cross-linking with glutaraldehyde in this article(OVA-GA-CL, coupling ratio of 25.2:1). Immune response was more easily flourished, because the five-carbon bridg reduces the coupling protein steric effect. Meanwhile, polyclonal antibody was purified by affinity chromatography and ammonium sulfate precipitation in this article and the antibody titer was detected by indirect ELISA. The purified antibody titers were respectively 1:16000 and 1:32000. Compared to the antiserum, the purified antibody titer was decreased, the analysis result of SDS-PAGE showed that the purity of purified antibody raised greatly, and the strips were prepared on this basis.Secondly, colloidal gold solution was prepared by improved Frens method. Reaction system was heated by water bath with rotary evaporator. The sodium citrate was fistly added to the systerm and after that the chlorine acid was mixed in the odium citrate solution. In the end, single scattered small particles were prepared and were identified by transmission electron microscopy and full wavelength scanner.At last, through the optimization of the pretreatment solution of the sample pad, and gold conjugate pad, the antigen and antibody coating buffer and the technology of preparation for the antibody-colloidal gold conjugates, the sensitivity of clenbuterol strip is 16ng/mL. And the results of stability test and reapeat tests show that the strip can be preserved at room temperature for 3 months and inter-and intra-batch repeatability is up to 95%. The strip prepared in this study indicated a good prospect.