RESUMEN
BACKGROUND: Early diagnosis of hepatitis C virus (HCV) infection is very important for the treatment of the disease. Development of sensitive and specific rapid detection assays is of great significance for the diagnosis. Here, we describe a promising method of using gold-labeled streptavidin fusion proteins as novel signal reporter in a rapid detection assay for HCV infection. METHODS: Recombinant genes encoding streptavidin fused with Escherichia coli maltose-binding protein (MBP) or with a portion of bacterial translational initiation factor 2 were cloned in expression vectors pMAL-5CX and pET28 and transformed in proper Escherichia coli host strains. The genes were induced and streptavidin fusion proteins, named M-STV and IF-STV, respectively, were purified by affinity chromatography to over 90% purity. The biotin-binding activity of M-STV and IF-STV was tested by enzyme-linked immunosorbent assay (ELISA). M-STV was labeled with colloidal gold nanoparticles and used as a signal reporter to develop a lateral flow-based rapid test for detecting anti-HCV antibodies in human blood samples. RESULTS: M-STV showed slightly higher biotin-binding activity and similar binding specificity as compared to commercial streptavidin. The gold-labeled M-STV bound specifically to biotin moieties immobilized on the rapid test strips in a dose-responsive manner and was successfully used in detecting HCV antibodies in serum samples of patients infected with HCV. The rapid test displayed higher detection sensitivity than gold-labeled commercial NeutrAvidin. CONCLUSION: Our results indicate that gold-labeled M-STV is a promising agent in rapid tests of HCV infection and possibly other viral infections.
Asunto(s)
Oro Coloide/química , Hepatitis C/diagnóstico , Proteínas Recombinantes de Fusión/metabolismo , Estreptavidina/metabolismo , Ensayo de Inmunoadsorción Enzimática/métodos , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Anticuerpos Antihepatitis/sangre , Humanos , Proteínas de Unión Periplasmáticas/química , Proteínas de Unión Periplasmáticas/genética , Proteínas de Unión Periplasmáticas/metabolismo , Proteínas Recombinantes de Fusión/química , Proteínas Recombinantes de Fusión/genética , Estreptavidina/química , Estreptavidina/genéticaRESUMEN
An energy-saving ethanol fermentation technology was developed using uncooked fresh sweet potato as raw material. A mutant strain of Aspergillus niger isolated from mildewed sweet potato was used to produce abundant raw starch saccharification enzymes for treating uncooked sweet potato storage roots. The viscosity of the fermentation paste of uncooked sweet potato roots was lower than that of the cooked roots. The ethanol fermentation was carried out by Zymomonas mobilis, and 14.4 g of ethanol (87.2% of the theoretical yield) was produced from 100g of fresh sweet potato storage roots. Based on this method, an energy-saving, high efficient and environment-friendly technology can be developed for large-scale production of fuel ethanol from sweet potato roots.