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C3d enhances the immune response against HBV-preS2/S following gene immunization in mice
Guan Q.-D.; Wang L.-X.; Guo Q.; Zhang J.-P.; Xu W.; Wang Y.; Xiong S.-D.
2005-01-12
Source PublicationNational Medical Journal of China
ISSN03762491
Volume85Issue:2Pages:101-105
AbstractObjective: To investigate whether C3d can enhance the immune response to HBV-preS2/S induced by direct injecting naked plasmid containing three copies of C3d and HBV-preS2/S in fusion form. Methods: HBV-preS2/S coding sequence was introduced into the eukaryotic expression vectors TR421 and TR421-C3d3 containing 3 copies of C3d respectively. PCR was used to identify the direction of insertion, DNA sequencing analysis was used on the positive clones. The recombinant plasmids TR421-preS2/S and TR421-preS2/S-C3d3 were transfected into the human breast cancer cells of the line 4T1 as a transient expression system. Semi-quantitative RT-PCR was used to detect the expression of HBV-preS2/S protein. TR421-preS2/S and TR421-preS2/S-C3d3 were injected into the anterior tibial muscles of female BALB/c mice, 6 in each group, 3 times at an interval of 3 weeks. Six mice were injected with blank plasmid TR421 as controls. The total blood was collected from the mice. ELISA was used to detect the level of specific anti-HBs antibody. The splenocytes of the immunized mice were collected and stimulated with HbsAg protein and then harvested to analyze the specific lympho-proliferative response by H-thymidine incorporation assay. CR2 positive Raji cells were added with plasmid transfected and mouse anti-HBs serum. Flow cytometry was used to detect the integration of preS2/S-C3d3 fusion protein. Results: The 4T1 cells transfected with TR421-preS2/S and TR421-preS2/S-C3d3 effectively expressed HBV-preS2/S, with the expression level of TR421-preS2/S-C3d3 lower by 15.25% than that of TR421-preS2/S. Twelve weeks after immunization, the specific anti-HBs-IgG level (A) was 0.81 ± 0.09 in the TR421 -preS2/S primed mice, significantly higher than that in the blank plasmid injected mice (0.49 ± 0.02, P <0.05); and the specific anti-HBs-IgG level (A) was 1.24 ± 0.29 in the TR421-preS2/S-C3d3 immunized mice, significantly higher than that of the TR421-preS2/S primed mice (P < 0.001). The lympho-proliferative response of the TR421-preS2/S primed mice was significantly stronger than that of the blank plasmid injected mice (P < 0.05) and the lympho-proliferative response of the TR421-preS2/S-C3d3 primed mice was significantly stronger than that of the TR421-preS2/S primed mice (P <0.05). Surface binding of protein could be detected only in the TR421-preS2 /S-C3d3 transfected supernatant. About 43.19% of the CR2 positive Raji cells integrated C3d fusion protein. TR421-preS2/S transfected supernatant and negative control did not show obvious integration. Conclusion: C3d enhances the humoral and cell-mediated immune responses against HBV induced by gene immunization.
KeywordComplement Gene Hepatitis B surface antigen Vaccines
URLView the original
Language英語
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Document TypeJournal article
专题University of Macau
AffiliationFudan University
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GB/T 7714
Guan Q.-D.,Wang L.-X.,Guo Q.,et al. C3d enhances the immune response against HBV-preS2/S following gene immunization in mice[J]. National Medical Journal of China,2005,85(2):101-105.
APA Guan Q.-D..,Wang L.-X..,Guo Q..,Zhang J.-P..,Xu W..,...&Xiong S.-D..(2005).C3d enhances the immune response against HBV-preS2/S following gene immunization in mice.National Medical Journal of China,85(2),101-105.
MLA Guan Q.-D.,et al."C3d enhances the immune response against HBV-preS2/S following gene immunization in mice".National Medical Journal of China 85.2(2005):101-105.
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