The Construction Of Streptavidin-Fluorescence Phycobiliprotein(SA-CpcB-PEB) And Its Application To Fluorescence Immunoassay

Fluorescence immunoassay is widely used by environmental pollutants monitoring, medicinal testing, food inspecting and other fields. In this research, Biotin-Streptavidin system(BSA) is a high sensitivity, good specificity and stability of multi-level signal amplification system. Phycobiliprotein could be applied to fluorescence immunoassay, as a new fluorescence probe.The Streptavidin gene was amplified by PCR from the plasmid of pUC57-sa and cloned by the transform of enzyme sites. Core Streptavidin was successfully constructed, and molecular weight is 59.2KD, theoretical activity is 16.5u, measured activity is 16.3u, can be preserved by lyophilized powder for long time.The fusion protein SA-CpcB-PEB was prepared by reconstitution SA-CpcB with PEB in vivo in E.coli. Tested the fluorescence and the property of binding with biotin of SA-CpcB-PEB, the results show that the fusion protein retained the two proteins's bifunctional effects. By using SA-CpcB-PEB as a fluorescence probe to detect trace substances in BSA fluorescence immunoassay, we believed it can be used for fluorescence immunoassay.By studying the temperature, pH, different inhibitors on the activity and stability of the fusion protein SA-CpcB-PEB, got the better storage conditions is 4℃, 1mM EDTA, 0.05% NaN3, neutral pH, 20mM KPB, 150mM NaCl ,which is beneficial to the specific application.