RESUMO
OBJECTIVE: Obtain the hepatitis C virus high purified subunit fusion protein and detect its immunogenicity. METHODS: With the vector of pET-11d, fusion protein was expressed in Escherichia coli BL21 (DE3) after induced by IPTG. The protein was then purified by DEAE negative ion exchange chromatography and Ni2+ affinity chromatography. Western Blot analysis was used to detect the antigenicity of the fusion protein. At the same time, the sera were collected and prepared from the immunized experimental animals in order to investigate the immunogenicity of the protein by EIA. RESULTS: High purified hepatitis C virus subunit fusion protein was obtained successfully. The EIA indicated that the fusion protein could elicit specific antibodies in the animals with very high titers. CONCLUSION: The hepatitis C virus subunit fusion protein expressed in prokaryotic system was proved to have strong immunogenicity. It could provide some helpful and useful information to the hepatitis C virus prophylactic and therapeutic vaccine development.
Assuntos
Hepacivirus/metabolismo , Proteínas Recombinantes de Fusão/imunologia , Proteínas Recombinantes de Fusão/metabolismo , Proteínas Virais/imunologia , Proteínas Virais/metabolismo , Animais , Ensaio de Imunoadsorção Enzimática , Escherichia coli/genética , Escherichia coli/metabolismo , Hepacivirus/genética , Imunoensaio , Coelhos , Proteínas Recombinantes de Fusão/genética , Proteínas Virais/genéticaRESUMO
OBJECTIVE: To construct virus-like particles of hepatitis B core antigen, with HBsAg "a" epitope exposed on the surface. METHODS: Hepatitis B surface antigen "a" epitope were inserted into the Hepatitis B core antigen, between the 78th (Asp) and the 79th (Pro) amino acids. The gene was synthesized after the codon optimized, then it was ligated to the express vector after been enzyme digest. The virus-like particles were observed by electron microscope and detected by ELISA after been expressed and purified. Immune the rabbits by the VLPs, then detect the antibody. RESULT: The virus-like particles were confirmed by electron microscope. Its antigenicity and immunogenicity were identified by ELISA. CONCLUSION: The prokaryotic express plasmid with the fusion gene has been constructed successfully. The virus-like particles have been expressed, purified and identified, which lays the foundation for its application in the further.
Assuntos
Epitopos/genética , Antígenos de Superfície da Hepatite B/genética , Vírus da Hepatite B/genética , Engenharia de Proteínas , Vírion/genética , Animais , Ensaio de Imunoadsorção Enzimática , Epitopos/análise , Epitopos/imunologia , Antígenos do Núcleo do Vírus da Hepatite B/análise , Antígenos do Núcleo do Vírus da Hepatite B/genética , Antígenos do Núcleo do Vírus da Hepatite B/imunologia , Antígenos de Superfície da Hepatite B/análise , Antígenos de Superfície da Hepatite B/imunologia , Vírus da Hepatite B/química , Vírus da Hepatite B/imunologia , Vírus da Hepatite B/ultraestrutura , Coelhos , Vírion/química , Vírion/imunologia , Vírion/ultraestruturaRESUMO
OBJECTIVE: To determine the main genotype of hepatitis B virus (HBV) in Xinjiang. METHODS: 200 HBsAg positive serum specimens were detected from more than 2000 serum of Xinjiang inhabitants, and HBV S gene was detected by using nPCR amplifying, and compared with the standard S region HBV nucleotide sequences of genotypes A-H retrieved from GenBank, then analyzed and drawn the polygenetic tree by MEGA3 software. RESULT: Gene in 127 (63.5%) serum specimens was detected from 200 samples. Among 127 serum specimens, 10 (7.8%) was genotype B, 58 (45.7%) was genotype C, and 59 (46.5%) was genotype D. CONCLUSION: Genotype B, C and D have been found in Xinjiang.
