ABSTRACT
Baculovirus expression vector system (BEVS) has been successfully applied to the over-expression of various proteins, thus providing sufficient materials for vaccine research. Compared to other systems, BEVS has many advantages: baculovirus solely being parasitic in invertebrates, the resultant products conferring high safety to mammalian, high expression level of recombinant proteins, preferable folding for eukaryotic protein, proper post-translational modification required for biological function, suitable for multiple genes co-expression and large-scale production with serum-free culture media. To better understand the advantages and prospective of BEVS for the vaccine research, this article will review the development of BEVS and its application on vaccine research.
Subject(s)
Animals , Baculoviridae , Genetic Vectors , Prospective Studies , Recombinant Proteins , VaccinesABSTRACT
Objective To establish an efficient baculovirus-insect cell expression system for the production of human immunodeficiency virus-1 ( HIV-1 ) envelope glycoprotein gp120 and to evaluate the physiochemical properties, antigenicity and immunogenicity of the recombinant protein. Methods The gene encoding HIV-1 NL4-3 gp120 was cloned into the downstream of pH promoter of the baculovirus transfer vec-tor pAcgp67B to construct the recombinant transfer vector pAc-gp120. Expression of the protein of interest was induced in baculovirus-infected High FiveTM insect cells. ELISA, analytical ultracentrifugation and size-exclusion chromatography were carried out to characterize physicochemical properties of the expressed gp120 protein. Immunogenicity of the recombinant gp120 protein was analyzed by HIV neutralization assay after im-munizing BALB/c mice with it. Results The recombinant HIV-1 gp120 protein was successfully obtained from the established insect cell expression system with a purity of more than 90％ and a mean yield of 13 mg/L in four batches. That recombinant HIV-1 gp120 protein was characterized by homogeneity in solution and possessed a good immunoreactivity to neutralizing antibodies and antisera against HIV. Immunogenicity analysis in BALB/c mice demonstrated that the recombinant gp120 protein could induce effective immune re-sponses against HIV-1 NL4-3. Conclusion A simple and scalable approach to obtain homogeneous and im-munogenic HIV-1 gp120 antigen is successfully established, which will promote further investigation of HIV vaccine candidates.
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Objective To establish an efficient baculovirus-insect cell expression system for the production of human immunodeficiency virus-1 ( HIV-1 ) envelope glycoprotein gp120 and to evaluate the physiochemical properties, antigenicity and immunogenicity of the recombinant protein. Methods The gene encoding HIV-1 NL4-3 gp120 was cloned into the downstream of pH promoter of the baculovirus transfer vec-tor pAcgp67B to construct the recombinant transfer vector pAc-gp120. Expression of the protein of interest was induced in baculovirus-infected High FiveTM insect cells. ELISA, analytical ultracentrifugation and size-exclusion chromatography were carried out to characterize physicochemical properties of the expressed gp120 protein. Immunogenicity of the recombinant gp120 protein was analyzed by HIV neutralization assay after im-munizing BALB/c mice with it. Results The recombinant HIV-1 gp120 protein was successfully obtained from the established insect cell expression system with a purity of more than 90％ and a mean yield of 13 mg/L in four batches. That recombinant HIV-1 gp120 protein was characterized by homogeneity in solution and possessed a good immunoreactivity to neutralizing antibodies and antisera against HIV. Immunogenicity analysis in BALB/c mice demonstrated that the recombinant gp120 protein could induce effective immune re-sponses against HIV-1 NL4-3. Conclusion A simple and scalable approach to obtain homogeneous and im-munogenic HIV-1 gp120 antigen is successfully established, which will promote further investigation of HIV vaccine candidates.
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Objective To discuss the feasibility and curative effect of intra-radiation stenting(125Iparticle stent)for the treatment of advanced esophageal cancers.Methods Fifteen patients with advanced esophageal cancer were enrolled in this study.Under X-ray guidance the esophageal stent,which was tied up with 125I radioactive particles,was orally inserted to the diseased region of the esophagus.The clinical manifestations and imaging findings were observed and the results were analyzed.Results After the operation all the clinical symptoms such as dysphasia showed an obvious improvement.No serious complications such as infection,hemorrhage,radiation pneumonia,etc.Occurred.The re-examination at 3-6 months after the treatment showed that the tumor size Was decreased in a certain degree in 14 patients,and in the remaining one patient the lesion became bigger and grew to the upper opening of the stent,resulting in esophageal restenosis.Conclusion The intra-esophageal implantation of radioactive stent is a feasible and safe treatment for the advanced esophageal cancers with excellent curative results.
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Objective To investigate the genetic features of pan-drag resistant pseudomonas aeruginosa. Methods The susceptibilities to 14 antibacterial agents were detected in pseudomonas aerations by K-B paper diffusing method. A strain of pan-drug resistant pseudomonas aeruginosa was selected randomly, and was used to amplify genes for β-Lactam antibiotic resistance (TEM, SHV, CTX-M-1 group,CTX-M-2 group, CTX-M-9 group, OXA-1 group, OXA-2 group, OXA-10 group, PER, GES, VEB,CARB, LCR, BEL, DHA, IMP, VIM, SPM, GIM, SIM and oprD2), aminoglycosidase drug resistance genes (including aminoglycosides modification enzymes gent,16S rRNA methylase gene) and genetic markers of plasmid (traA and traF), integrons (tnpA, tnpU and merA), transposons Ⅰ (int Ⅰ 1 ) by PCR,and the sequences of above genes were analyzed. Results The pseudomonas aeruginosa was resistant to all 14 antibacterial agents, and the following genes were positive in PCR: TEM-1 and CARB-3 types of genes for β-Lactam antibiotic resistance (deletion of oprD2), aac (6')-Ⅱ and ant (2")-Ⅰ of aminoglycosidase resistance genes, transposons Ⅰ (int Ⅰ 1 ), and the merit of integrons. Conclusions Pseudomonas aeruginosa shows high resistance to most antibacterial agents, which should draw attention in clinic.
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OBJECTIVE To investigate the 16S rRNA methylases gene and AMEs of 30 strains multi-resistant Pseudomonas aeruginosa isolated from clinic work in our hospital.METHODS Collected 50 P.aeruginosa strains from two hospitals(30 strains of multi-resistant P.aeruginosa from Yantai,Shandong,20 strains of pan-drug resistant P.aeruginosa from Jiangsu),and analyzed 6 kinds of 16S rRNA methylases(armA,rmtA,rmtB,rmtC,rmtD and npmA) and 6 kinds of AMEs gene aac(3)-Ⅰ,aac(3)-Ⅱ,aac(6′)-Ⅰb,aac(6′)-Ⅱ,aac(3″)-Ⅰ,and aac(2″)-Ⅰ by PCR and sequence analysis.RESULTS Among 30 strains of multi-resistant P.aeruginosa from Yantai,the positive rate of 4 kinds of genes aac(3)-Ⅱ,aac(6′)-Ⅱ,aac(3″)-Ⅰ,and aac(2″)-Ⅰ was 20%(6/30),46.7%(14/30),36.7%(11/30) and 3.3%(1/30).The other 8 kinds genes were all negative.The total positive rate of AMEs gene was 46.7%(14/30).In 20 strains of pan-drug resistant P.aeruginosa,the positive rate of 6 kinds of genes rmtB,aac(3)-Ⅱ,aac(6′)-Ⅰb,aac(6′)-Ⅱ,aac(3″)-Ⅰ,and aac(2″)-Ⅰ was 55%(11/20),60%(12/20),35%(7/20),60%(12/20),50%(10/20) and 45%(9/20).The other 6 kinds genes were all negative.The total positive rate of AMEs gene was 95%(19/20).CONCLUSIONS It is the first report that 16S rRNA methylases gene is existed in P.aeruginosa;there is very high positive rate of AMEs genotypes in P.aeruginosa;there are differences in gene existing among two hospitals.
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OBJECTIVE To provide laboratory evidence for the prevention and control of coagulase negative staphylococcus(CNS),and study the distribution and drug resistance of CNS in our hospital.METHODS CNS of inpatients from Oct 2005 to Dec 2007 was isolated and identified with ATB Expression microbe identification and drug sensitivity system.RESULTS A total of 354 strains of CNS were isolated,from the main samples of secretion,sputum,blood and cerebrospinal fluld.The isolation rate from departments of pediatrics,ICU,orthopedics and neurology were 9.90%,9.30%,9.00% and 5.60%,respectively.CONCLUSIONS CNS is playing a significant role in nosocomial infection.The drug resistance of CNS is very serious.To pevent nosocomial infection,it is critically important to monitor the antimicriobial resistance of CNS and use autibiotics more rationally.
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OBJECTIVE:To correct the standard of bismuth content in Weishusan.METHODS:The contents of effective bismuth in Weishusan and other three reference substances were detected by compleximetry.After adjusting the dosage of Weishusan,clinical rank sum test was carried out.RESULTS:The content of bismuth in original formula of Weishusan was rather high,therefore the inventory of bismuth subcarbonate in preparation technic of Weishusan should be reduced to7.0kg.CONCLUSION:The quality standard should be reformulated as bismuth subcarbonate0.1g(?10%)in per gram of Weishusan.