2006年06期目次

  • Establishment of A Cell Strain for High Expression of Human-Mouse Chimeric Antibody against Human TNF-α and Analysis of Expressed Antibody

    NIE Yan-tao△,HE Tai-ping,XIE Rong-lin,et al ( △Rongsheng Pharmaceuticals,Chengdu 610023,China)

    Objective To establish a cell strain for high expression of human-mouse chimeric antibody against human TNF-α.Methods Transfect CHO cells with the constructed eukaryotic expression vector of gene encoding human-mouse chimeric antibody against human TNF-α and screen positive clones by DHFR/MTX gene amplification strategy.Analyze the characteristic of expressed antibody by ELISA,Western blot and neutralization test in vitro and compare with that of parental murine McAb.Results A transfected cell line 2F5 for high expression of chimeric antibody against human TNF-α was established.After static culture of the transfected cells for 4 d,the expression level of gene engineering antibody was about 80 mg/L.Compared with its parental McAb F7/2,the expressed antibody showed good specificity and neutralizing activity,and its relative affinity was 2.1×10 -10 mol/L.Conclusion A gene engineering cell strain for high expression of chimeric antibody against human TNF-α was successfully established.The expressed antibody showed potential prospect in clinical application.

    2006 06 [Abstract][OnlineView][Download 160K]

  • Construction and Expression of Trichosanthin Mutant Gene TCS KR173-174CG and Purification of Expressed Product

    AN Qun-xing△,MU Shi-jie,ZHANG Xian-qing,et al ( △Department of Blood Transfusion,Xijing Hospital,Fourth Military Medical University,Xi’an 710033,China)

    Objective To construct and express a trichosanthin(TCS) gene mutant and purify the expressed product.Methods Predict the potential antigenic determinant on TCS molecule by computer modeling and induce site-directed mutation.Amplify gene mutant TCS KR173-174CG by PCR using the genomic DNA of Trichosanthes kirilowii as a template and insert into expression vector pRSET-A,then transform to E.coli BL21(DE3) for expression under induction of IPTG.Purify the expressed product by Ni-NTA affinity column chromatography.Results The target protein in a soluble form was successfully expressed in E.coli.Homogenous TCS mutant protein was obtained after purification of expressed product.Conclusion TCS mutant gene TCS KR173-174CG was successfully constructed and expressed.

    2006 06 [Abstract][OnlineView][Download 258K]

  • Expression of Severe Acute Respiratory Syndrome(SARS) Virus S2 Protein in Pichia pastoris

    WANG Rui-lin△,JIN Ning-yi,ZHAO Bo,et al ( △College of Animal Science and Veteranary Medicine,Changchun 130062,China)

    Objective To express severe acute respiratory syndrome(SARS) virus S2 protein in Pichia pastoris.Methods A 1 590 bp gene fragment encoding the S2 protein of SARS virus was synthesized according to the SARS virus genome sequence reported in GenBank and recombined with CTL epitope gene(195 bp),then cloned into Pichia pastoris expression vector pPIC9K.The constructed recombinant plasmid pPIC9K-S2 was transformed to Pichia pastoris GS115,and the positive clones were screened by MD and MM plate cultures and identified by PCR.The obtained recombinant Pichia pastoris strain GS115/pPIC9K-S2 was induced with methanol,and the expressed product was identified by SDS-PAGE and Western blot.Results Recombinant plasmid pPIC9K-S2 was correctly constructed and highly expressed in Pichia pastoris.The expressed product showed specific reaction with SARS virus-positive serum.Conclusion SARS virus S2 protein was successfully expressed in Pichia pastoris and showed good antigenic specificity.

    2006 06 [Abstract][OnlineView][Download 196K]

  • Construction of Recombinant Plasmid for Co-expression of Nucleoprotein and Glycoprotein of Rabies Virus aG Strain in Eukaryotic Cells

    YANG Hui-juan,CHEN Jun-ying,SUN Ming-bo,et al (Institute of Medical Biology,Chinese Academy of Medical Sciences,Peking Union Medical College,Kunming 650118,China)

    Objective To construct a recombinant plasmid for co-expression of nucleoprotein(NP) and glycoprotein(GP) of rabies virus aG strain in CHO-K1 cells.Methods The genes encoding the NP and GP genes of rabies virus aG strain were amplified by RT-PCR and cloned into vector pIRES to construct a dicistonic plasmid pING,then transfect to CHO-K1 cells in mediation of liposome.The transfectants were screened with G418,and the expressed product was analyzed by ELISA and IFA.Results The restriction map of recombinant plasmid pING showed NP and GP gene fragments at lengths of 1 353 bp and 1 575 bp respectively.ELISA and IFA proved the expression of both NP and GP in the transfected cells.Conclusion The recombinant plasmid pING for co-expression of NP and GP of rabies virus in eukaryotic cells was constructed.It laid a foundation of further development of recombinant rabies vaccine.

    2006 06 [Abstract][OnlineView][Download 226K]

  • Relationship between Soluble Fas Concentration and Expression of Vascular Endothelial Growth Factor(VEGF) and Tissue Inhibitor of Metalloproteinase-1(TIMP-1) in Metabasis of Breast Cancer in Lymph Node

    WANG Xin-ming△,CHEN Yun-bo,JIA Ming-ku,et al ( △Department of Oncology,The Second Hospital,Jilin University,Changchun 130021,China)

    Objective To analyze the relationship between soluble Fas(sFas) concentration and the expressions of vascular endothelial growth factor(VEGF) and tissue inhibitor of metalloproteinase-1(TIMP-1) in breast cancer and explore the influence of sFas concentration on the growth,infiltration and metabasis of breast cancer.Methods Determine the VEGF,TIMP-1 and serum sFas concentration of patients with breast cancer by S-P immunohistochemical method and ELISA respectively,and analyze the relationship among the unbalanced expression of VEGF and TIMP-1 and the serum sFas concentration in breast cancer with infiltration and metabasis in lymph node by the combination of determination results with the pathological parameters of breast cancer in clinic.Results The expression of VEGF was significantly related to the metabasis of breast cancer in lymph node.Of the VEGF-positive tumors,56% showed metabasis in lymph node and only 7% showed no metabasis in lymph node.The expression rate of VEGF was positively related to the progress of TNM phases and histological stages of breast cancer.Significant relationship was also observed between the expression of TIMP-1 and the metabasis of breast cancer in lymph node.The expression rate of TIMP-1 was negatively related to the progress of TNM phases and histological stages of breast cancer. The tumors positive for VEGF but negative for TIMP-1 showed the highest infiltration rate and metabasis rate in lymph node as well as the highest serum sFas concentration.However,the tumors negative for VEGF but positive for TIMP-1 showed the lowest metabasis rate in lymph node and the lowest serum sFas concentration.Conclusion Feedback mechanism existed among sFas,VEGF and TIMP-1,which could activate each other and showed close relationship.sFas concentration reflected the expressions of matrix metalloproteinase(MMPs) and VEGF indirectly,and its change reflected the growth,local infiltration and metabasis of breast cancer.

    2006 06 [Abstract][OnlineView][Download 110K]

  • Gene Cloning and Prokaryotic Expression of Human Cytomegalovirus Glycoprotein gB/AD-1

    LIU Lan-jun,HE Tai-ping,YANG Chun,et al (Chengdu Institute of Biological Products,Chengdu 610023,China)

    Objective To clone the gene fragment gB/AD-1 encoding the glycoprotein of human cytomegalovirus(HCMV) and construct its prokaryotic expression system.Methods Amplify gB/AD-1 gene fragment from the genomic DNA of HCMV AD169 strain and clone into vector pGEM-T,then,after identification by sequencing,subclone to expression vector pET-15b. Transform the constructed recombinant plasmid pET-15b/gB/AD-1 to E.coli for expression of gB/AD-1 under induction of IPTG.Purify the expressed product by one-step immobilized metal ion affinity chromatography(IMAC) and identify by Western blot.Results The expressed gB/AD-1 contained 35% of total somatic protein.The purity of expressed product after purification was more than 95%.Western blot showed specific reaction of the expressed product with CMV antibody-positive human serum.Conclusion The recombinant plasmid pET-15b/gB/AD-1 was successfully constructed,and gB/AD-1 was highly expressed in E.coli.

    2006 06 [Abstract][OnlineView][Download 181K]

  • Preliminary Study on Immunological Activity of BCG-CpG-DNA

    ZHAO Ai-hua,KOU Li-jie,QIAO Lai-yan,et al (National Institute for the Control of Pharmaceutical and Biological Products,Beijing 100050,China)

    Objective To study the immunological activity of BCG-CpG-DNA.Methods Determine the immunological activity of BCG-CpG-DNA by lymphocyte proliferation test,T cell subgrouping,the production of cytokine and the effect on killing activity of NK cells.Results BCG-CpG-DNA promoted the proliferation of T and B lymphocytes in mice,activated macrophages to secrete IL-12,enhanced the ability of ConA in inducing IFN-γ,increased the contents of CD3+,CD4+ and CD8+T cell subgroups in murine splenic cells and enhanced the killing activity of murine NK cells.All the results of test and control groups showed significant difference.Conclusion BCG-CpG-DNA activated antigen-presenting cells and showed good immunological activity.

    2006 06 [Abstract][OnlineView][Download 121K]

  • Expression of Recombinant Thermomonospora fusca Cellulase Cel6A in Pichia pastoris and Purification of Expressed Product

    MU Jing-yu△,WANG Xi,WANG Qiao,et al ( △College of Pharmacy,Jilin University,Changchun 130021,China)

    Objective To express the cellulase Cel6A of Themomonospora fusca in Pichia pastoris.Methods Amplify cellulase Cel6A gene by PCR using the extracted genomic DNA of Themomonospora fusca as a template.Clone the amplified gene into vector pGAPZαA and transform to Pichia pastoris X-33 strain by electroporation.Screen the positive transformants by colony PCR method for subculture.Select stable cell strains and culture for 3 d.Collect the culture supernatant and purify the expressed recombinant cellulase Cel6A of Themomonospora fusca by His-Bind Quick Cartridge 300 chromatography.Results Electrophoretically pure recombinant cellulase Cel6A of Themomonospora fusca was obtained by one-step His-Bind Quick Cartridge 300 chromatography.SDS-PAGE showed that the relative molecular weight of expressed product was 60 000.The recovery rate of expressed protein was 200 mg/L.The activity of expressed protein was 500 U/mg by carboxymethyl cellulose method and 1 U/mg by filter paper method.Conclusion Recombinant cellulase Cel6A of Themomonospora fusca was successfully expressed in Pichia pastoris.

    2006 06 [Abstract][OnlineView][Download 127K]

  • Influence of Vitamin A on Immunoregulatory Function of Mice

    XING Jie△,LI Yu-fen,LUO Hai-ying,et al ( △Department of Pediatrics,The Second Hospital,Jilin University,Changchun 130041,China)

    Objective To explore the influence of vitamin A on the immunoregulatory function of mice.Methods Twenty-four mice were divided into three groups and fed by the prepared forages with low,normal and high vitamin A contents for 5 weeks respectively,then determined for serum vitamin A concentration by microfluorescence spectrophotometry,and for the IL-4 and IFN-γ levels in CD4+T cells by trichromatic flow cytometry.Results Significant difference was observed in the serum vitamin A concentrations of mice in the three groups.However,the IFN-γ level of mice fed by the forage with low vitamin A content(28.60%±10.82%) was significantly higher than that with normal vitamin A content(20.13%±6.39%).The IL-4 level of mice fed by the forage with high vitamin A content(5.31%±2.04%) was significantly higher than that with normal vitamin A content(2.80%±1.37%). Along with the increasing content of vitamin A in forage,the IFN-γ expression and Th1 immune response level decreased,and the IL-4 expression and Th2 immune response level increased.Conclusion Vitamin A showed significant influence on the Th1/Th2 immune response of mice.

    2006 06 [Abstract][OnlineView][Download 104K]

  • Construction and Identification of Recombinant Fowlpox Virus Expressing GP5/GP3 of Pig Reproduction and Respiration Syndrome Virus(PRRSV) and Swine IL-18

    SHEN Guo-shunΔ,JIN Ning-yi,QIN Xiao-guang,et al ( ΔCollege of Animal Science and Veterinary Medicine,Jilin University,Changchun 130062,China)

    Objective To develop a safe and effective recombinant fowlpox(FPV) as a vaccine against pig reproduction and respiration syndrome virus(PRRSV).Methods Insert the GP5/GP3 gene of Changchun strain of PRRSV downstream to the composite promoter of fowlpox virus transferring vector pUTAL-IL-18.Co-transfect chick embryo fibroblast(CEF) with the constructed recombinant plasmid pUTAL-IL-18-GP5-GP3 and FPV 282-E4 strain for homologous recombination.After 3 cycles of pressure screening in presence of 5′-bromo-deoxyuridine(BrdU),the recombinant virus was identified by PCR,RT-PCR and IFA.Results The target gene at a length of 340 bp was amplified from the genome and total RNA of recombinant fowlpox virus.The CEF transfected with recombinant fowlpox virus reacted with PRRSV specific fluorescent antibody.It indicated that the target gene was successfully expressed in recombinant fowlpox virus.Conclusion A recombinant fowlpox virus for expression of GP5/GP3 gene of PRRSV was successfully constructed.

    2006 06 [Abstract][OnlineView][Download 148K]

  • Stability of Recombinant E.coli Strain for Expression of MAGE-1/HPS70/MAGE-3

    HU Pei-zhen△,SUI Yan-fang,LI Xia,et al ( △Department of Pathology,Xijing Hospital,The Fourth Military Medical University,Xi’an 710032,China)

    Objective To study the stability of biological property of recombinant E.coli strain for the expression of MAGE-1/HSP70/MAGE-3.Methods Subculture the constructed recombinant E.coli strain pET28a-MAGE-1/HSP70/MAGE-3/BL21(DE3) for 50 passages,and select monoclones every 10 passages for induction and test for losing rate of plasmid. Extract the plasmids of strains of various passages for scanning electron microscopy. Determine the expression level of fusion protein and test for the biochemical property of the recombinant strain.Results Recombinant strain pET28a-MAGE-1/HSP70/MAGE-3/BL21(DE3) showed typical morphology of E.coli.The test results of the strain of various passages showed no significant difference from those of primary strain.Conclusion Recombinant E.coli strain pET28a-MAGE-1/HSP70/MAGE-3/BL21(DE3) showed stable biological property and might be used as the bacterial seed for production.

    2006 06 [Abstract][OnlineView][Download 167K]

  • Preparation and Property of Rabies Virus Liposome Vaccine for Human Use

    GUO Xiu-xia△,FU Quan,CHEN Yan,et al ( △Changchun Institute of Biological Products,Changchun 130062,China)

    Objective To prepare the rabies virus liposome vaccine for human use and explore its pharmaceutical and toxicological properties.Methods Prepare liposome with phospholipid,cholesterol and stearylamine by reverse evaporation,then add purified rabies virus antigen for preparation of rabies virus liposome vaccine by lyophilization.Observe the prepared vaccine for morphology under electron microscope,and determine for envelope rate and release in vitro.Analyze the particale distribution and size of the vaccine by laser diffraction particle size analyzer.Perform local stimulation,hypersensitivity and abnormal toxicity tests on the prepared vaccine.Results The prepared rabies virus liposome vaccine under electron microscope were in forms of round or elliptic particles distributed evenly,at a mean size of 12.25 μm.The envelope rate of the vaccine was more than 60%.The vaccine showed no stimulation or abnormal toxicity and was qualified in hypersensitivity test.Conclusion The prepared rabies virus liposome vaccine showed stable property and was value of further study.

    2006 06 [Abstract][OnlineView][Download 181K]

  • Preparation,Safety and Immunogenicity of S.paratyphoid A-Tetanus Toxoid Conjugate Vaccine

    QIN Min,YANG Guo-you,TU Xiao-ping,et al (Chengdu Institute of Biological Products,Chengdu 610023,China)

    Objective To prepare S.paratyphoid A-tetanus toxoid(TT) conjugate vaccine and study its safety and immunogenicity.Methods The polysaccharide O-SP of S.paratyphoid A was activated with CDAP,derived with ADH,then coupled to TT using EDAC as condensing agent and purified by column chromatography to prepare S.paratyphoid A-tetanus toxoid(TT) conjugate.After control tests,the conjugate was further tested for safety and immunogenicity in animals.Results The ratio of polysaccharide to protein of the prepared conjugate was as stable as 0.6-0.8.The recovery rate of conjugate,determined by Sepharose Cl-4B chromatography,was more than 75% when Kd value was not more than 0.2.The content of high molecular conjugate was not less than 90%.Double immunodiffusion test showed visual precipitation lines of the conjugate with hyperimmune rabbit serum against S.paratyphoid A and TT antiserum.The conjugate showed good safety in animal test.The antibody titer against S.paratyphoid A of mice immunized with the conjugate increased significantly,and the antibody positive conversion rate reached 100%.Conclusion The S.paratyphoid A-tetanus toxoid(TT) conjugate vaccine prepared by activating polysaccharide O-SP with CDAP showed good stability,safety and immunogenicity.

    2006 06 [Abstract][OnlineView][Download 97K]

  • Construction of Hepatitis B Virus DNA Vaccine Carrying Interleukin-2 Gene

    WU Xiao-juan△,ZHAO Da-peng,LENG Mei,et al ( △Changchun Institute of Biological Products,Changchun 130062,China)

    Objective To construct the hepatitis B virus(HBV) DNA vaccine carrying interleukin-2(IL-2) gene and explore its immune effect.Methods Insert internal ribosomal entry site(IRES) gene into the polyclonal site of plasmid pVAX1,then insert HBsAg and IL-2 genes into the same plasmid,at the two sides of IRES gene respectively,to construct HBV DNA vaccine pHII.Transfect COS-7 cells with pHII for transient expression.Immunize BALB/c mice with the extracted plasmid pHII for 3 times at weeks 0,2 and 4 respectively and determine the HBsAg and HBsAb contents in sera starting from week 1,using plasmid pVAX1 as negative control and plasmid pVAX/HBsAg as positive control.Kill the mice at week 13 and determine for the CD4+ and CD8+ on surface of T lymphocyte in murine spleens by flow cytometry.Results HBsAg and IL-2 were detected in the supernatant of COS-7 cells transfected with plasmid pHII.HBsAbs were detected in the sera of mice in test and positive control groups at weeks 2 and 4 respectively,but not detected in those of mice in negative control group.However,no HBsAg was detected in the three groups.Compared with those in positive control group,the antibody in test group appeared 2 weeks earlier,the antibody positive rate was 2.5 times higher,and the quantity of antibody increased about 3 folds.Both the number of CD4+ molecule and the ratio of CD4+ to CD8+ of mice in test group were significantly higher than those in positive control group.Conclusion The constructed HBV DNA vaccine pHII induced immune response in mice successfully.Compared with that of HBV DNA vaccine without molecular adjuvant,the immune effect of the constructed HBV DNA vaccine was good.

    2006 06 [Abstract][OnlineView][Download 192K]

  • Preparation of Antibody against Rabies Virus and Development of ELISA Method for Determination of Rabies Virus Antigen

    ZHAO Zhi-jing,ZHUANG Hui,ZHANG Jia-ming,et al (Department of Microbiology,Peking University Health Science Center,Beijing 100083,China)

    Objective To prepare the antibody against rabies virus(RV),with neutralizing activity,and develop an ELISA method for the determination of RV antigen.Methods The polyclonal antibody(PcAb) against RV was prepared by immunizing New Zealand rabbits with RV strain.The hybridoma cell strain stably secreting the monoclonal antibody(McAb) against RV was established by fusing murine myeloma SP2/0 cells with the splenocytes of BALB/c mice immunized with RV strain.The neutralizing activity of antibody was determined by murine neutralization test.The RV antigen was determined by double antibody sandwich ELISA and competitive ELISA.Results The ELISA titers of PcAbs of two rabbits were 1∶6.0×103 and 1∶1.2×104,and the neutralizing activities of purified rabbit IgG against RV were 46.3 and 29.2 IU/ml respectively.Nine hybridoma cell strains stably secreting McAb to RV,of IgG1 or IgG2b subtype,were established.The ELISA titers of McAbs secreted by the strains in ascites were 1∶1×105-1∶1×107.McAbs 3E5,4C2 and 4F8 showed neutralizing activities.Western blot proved that McAb 4C2 recognized the linear epitope of RV glycoprotein.A double antibody sandwich ELISA for RV antigen was developed using McAb 4C2 as capture antibody and rabbit PcAb as detection antibody.Meanwhile,a competitive ELISA was developed by incubating McAb 4C2 at a constant concentration with RV then determining RV antigen by indirect ELISA.Conclusion Both the prepared PcAb and McAb showed neutralizing activities and could be used for the determination of RV antibody by ELISA.

    2006 06 [Abstract][OnlineView][Download 174K]

  • Purification and Biological Activity of Recombinant Melittin- 88ArgIL-2

    LIU Ming-jun,WANG Bin,QIAN Dong-meng,et al (Department of Microbiology,Key Laboratory of Medicine and Biotechnology,Medical College,Qingdao University,Qingdao 266021,China)

    Objective To prepare purified recombinant melittin- 88ArgIL-2 chimeric protein and analyze its biological activity.Methods Transform the constructed recombinant plasmid pGEX-4T-2/melittin- 88ArgIL-2 to E.coli DH5α for expression under induction of IPTG.Purify the expressed product by ion exchange and affinity chromatography and identify by SDS-PAGE.Determine the inhibitory effect of expressed chimeric protein on the proliferation of Hela cells by MTT method.Results The purity and protein concentration of expressed soluble protein after purification reached 95% and 0.5 g/L respectively.The purified chimeric protein,at a concentration of 20 μg/L,showed significant inhibitory effect in vitro on the proliferation of Hela cells.Conclusion The purified recombinant melittin- 88ArgIL-2 chimeric protein was successfully prepared and showed biological activity.It laid a foundation of scale-up of pilot procedure and development of purification procedure of the chimeric protein.

    2006 06 [Abstract][OnlineView][Download 171K]

  • Test for Contamination with Adventitious Agents in Cells Used for Production of Biologics in China

    MENG Shu-fang△,LI Xiu-lan,FENG Jian-ping,et al ( △National Institute for the Control of Pharmaceutical and Biological Products,Beijing 100050,China)

    Objective To test the contamination with adventitious agents in the cells used for production of biologics in China.Methods Test the bacteria,fungi,mycoplasma and common adventitious viruses in the cells used for production of biologics according to the requirements in Chinese Pharmacopoeia(Volume Ⅲ,2005).Results A total of 79 cell lines from the cell bank for production of biologics were tested from 2003 to March 2006,of which 10 lines(12.7%) were contaminated with mycoplasma and 5 lines(6.3%) with adventitious viruses.Of the 5 cell lines contaminated with adventitious viruses,two 293 cell lines caused the abnormal morphology of human diploid cells inoculated,two Vero cell strains caused the deaths of all the embryonated eggs inoculated,and the other one Vero cell strain caused the deaths of all the suckling or adult mice on the 7th day after inoculation.Histopathological examination proved the pathological changes at various degrees in the brain tissues of mice inoculated.Conclusion The cells for production of biologics in China are contaminated with adventitious agents at a certain degree.

    2006 06 [Abstract][OnlineView][Download 292K]

  • Serological Diagnosis Using Recombinant 38 kD Protein of Mycobacterium tuberculosis as Antigen

    YUE Qiao-hong△,SU Ming-quan,YANG Jun-lan,et al ( △Department of Clinical Laboratory,Xijing Hospital,The Fourth Military Medical University,Xi’an 710032,China)

    Objective To clone and express the gene encoding 38 kD protein of Mycobacterium tuberculosis and perform serological diagnosis using the protein as an antigen.Methods Amplify 38 kD antigen gene from the genomic DNA of Mycobacterium tuberculosis by PCR,identify by sequencing,then clone into expression vector pGEX-4T-2 and transform to E.coli BL21 for expression.Purify the expressed product by GSTrapFF protein column chromatography and test for sensitivity and specificity by ELISA.Results The expressed product contained 28% of total somatic protein,and its sensitivity and specificity were 79.8% and 99.0% respectively.Conclusion The recombinant 38 kD protein might be used as an antigen for diagnosis of infection with Mycobacterium tuberculosis.

    2006 06 [Abstract][OnlineView][Download 157K]

  • Identification of Live Attenuated Varicella Vaccine by Amplification of VZV gE Glycoprotein Gene

    JIN Yu-lan,CHEN Zhe-wen,QU Ai-dong,et al (Shanghai Institute of Biological Products,Shanghai 200052,China)

    Objective To identify live attenuated varicella vaccine by the amplification of varicella-zoster virus(VZV) gE glycoprotein gene.Methods Extract viral DNA from the samples of live attenuated varicella vaccine,amplify by PCR and identify by agarose gel electrophoresis.Results The amplified gE genes from live attenuated varicella vaccine prepared with VZV Oka strain at a titer of not less than 2.0 lgPFU/ml showed specific bands on agarose gel electrophoretic profile.However,no VZV gE gene was amplified from the viral DNA of control vaccines,such as live attenuated measles vaccine,measle-mumps combined vaccine and rubella vaccine as well as herpes simplex virus types Ⅰ and Ⅱ,each at a titer of not less than 4.2 lgCCID 50/ml.Conclusion gE glycoprotein gene might be used as a specific indicator for the identification of VZV.

    2006 06 [Abstract][OnlineView][Download 174K]

  • Quality Control of Interferon Liposome

    LI Yun-fu△,ZHANG Shui-hua,LI Jiu-li,et al ( △College of Life Science and Biotechnology,Shanghai Jiao Tong University,Shanghai 200030,China)

    Objective To develop a stable quality control system of interferon liposome.Methods Analyze the morphology,particle size,envelope rate,residual organism content and stability by using electron microscope,size distribution analyzer,gas phase chromatography and EIA reader.Results Interferon liposome was in a form of round particles in diameters of 50-350 nm under transmission electron microscope.Analysis by size distribution analyzer showed normal distribution of size of interferon liposome,in a mean diameter of 200-210 nm.The span of particles was 0.80-1.10,and their envelope rate reached more than 90%.The residual organism content was less than the standard described in Chinese Pharmacopoeia.After storage at 2-8℃ for 6 months,the leakage rate of liposome was not more than 20%,and the total activity of interferon showed no significant change.Conclusion The developed quality control system of interferon liposome was stable and practical.

    2006 06 [Abstract][OnlineView][Download 163K]

  • Curative Effect of Recombinant Human Interferon α2a Gel on Herpes Simplex Virus Infection in Animals

    ZHANG Shu-zi△,LI Fan,YI Shi-hong ( △Jilin Yatai Biopharmaceutical Industry Co. Ltd,Changchun 130033,China)

    Objective To study the curative effect of recombinant human interferon α2a(rhIFNα2a) gel on in vivo infection with herpes simplex virus(HSV) in animals.Methods The guinea pig model of skin infection with HSV-1 and the mouse model of vaginitis caused by HSV-2 infection were established.Divide the model guinea pigs and mice into three test,one parallel control and one blank control groups separately.Treat the animals in the three test groups with rhIFNα2a gel at dosages of 3×105,2×105 and 1× 105 IU/g respectively,while those in parallel control groups with acyclovir.However,the animals in blank control groups were untreated. Evaluate the curative effect of each group.Results The rhIFNα2a gel at dosages of 2×105 and 3×105 IU/g decreased the HSV-1 content in skin tissue of guinea pigs as well as the mortality rate of mice,and prolonged the survival period of mice significantly.The curative effect of rhIFNα2a gel showed no significant difference with that of acyclovir.Conclusion rhIFNα2a gel inhibited the pathological change on skin caused by HSV-1 and shortened the course of disease.It showed good curative effect on vaginitis caused by HSV-2 in mice.

    2006 06 [Abstract][OnlineView][Download 93K]

  • Specificity and Sensitivity of VIDAS HIV Duo Assay Kit

    XU Si-hong△,SONG Ai-jing,LI Xiu-hua,et al ( △National Institute for the Control of Pharmaceutical and Biological Products,Beijing 100050,China)

    Objective To evaluate the specificities and sensitivities of VIDAS HIV Duo Quick kit and VIDAS HIV Duo Ultra kit for simultaneous detection of HIV antibody and antigen.Methods A total of 287 serum samples from the patients or doubtful patients with HIV infection and 1 240 serum samples from the patients without HIV infection were tested by VIDAS HIV Duo Quick kit,VIDAS HIV Duo Ultra kit and HIV antibody reference kit,separately.The samples with inconsistent test results with that by HIV antibody reference kit were further confirmed.The specificities and sensitivities of VIDAS HIV Duo Quick kit and VIDAS HIV Duo Ultra kit were compared with that of HIV antibody reference kit.Test the serially diluted culture supernatant of three HIV strains by VIDAS HIV Duo Quick kit,VIDAS HIV Duo Ultra kit and HIV-1 p24 reference kit separately and compare the sensitivities of the three kits in test for p24 antigen.Results A total of 1 277 samples were identified as HIV antibody negative,and 250 as positive.Compared with those of HIV antibody reference kit,the specificity and sensitivity of VIDAS HIV Duo Quick were 99.45% and 100%,and those of VIDAS HIV Duo Ultra kit were 99.14% and 99.20% respectively.However,the sensitivities of VIDAS HIV Duo Quick kit and VIDAS HIV Duo Ultra kit in test for culture supernatant of HIV strain were not less than that of HIV-1 p24 antigen reference kit.The samples from patients in window period could also be confirmed by VIDAS HIV Duo Quick kit and VIDAS HIV Duo Ultra kit.Conclusion On the basis of test for HIV p24 antigen,the specificities and sensitivities of VIDAS HIV Duo Quick kit and VIDAS HIV Duo Ultra kit in test for HIV antibody showed no significant decrease.Both the kits were suitable for the samples from patients in window period.

    2006 06 [Abstract][OnlineView][Download 146K]

  • Application of HpaA Protein in Diagnosis of Helicobacter pylori Infection

    WU Li-xian,WANG Guo-fu,BAI Li,et al (Department of Microbiology and Immunology,Dali College,Dali 671000,China)

    Objective To develop an indirect ELISA method for determination of serum HpaA antibody of Helicobacter pylori and explore the application of recombinant HpaA protein as an antigen in the diagnosis of H.pylori infection.Methods Express the HpaA gene amplified from clinical isolate of H.pylori in E.coli.Purify the expressed HpaA by Ni 2+-NTA column chromatography.Develop an indirect ELISA method for determination of serum HpaA antibody using the purified HpaA protein as an antigen.Evaluate the feasibility of application of HpaA protein in diagnosis of H.pylori infection using laboratory diagnostic standard.Results SDS-PAGE proved that the recombinant HpaA protein mainly existed in ultrasonic supernatant of Helicobacter pylori.The sensitivity and specificity of the purified HpaA protein in the determination of HpaA antibody in clinical specimens were 100.0% and 90.1% respectively.Conclusion The developed indirect ELISA method for determination of HpaA antibody showed good sensitivity and specificity.It laid a foundation of preparation of commercial kit.

    2006 06 [Abstract][OnlineView][Download 128K]

  • Preparation of Purified Protein Derivative of Histoplasmin

    CHEN Bao-wen,SHEN Xiao-bing,SU Cheng,et al (National Institute for the Control of Pharmaceutical and Biological Products,Beijing 100050,China)

    Objective To prepare purified protein derivative of histoplasmin.Methods Culture Histoplasma capsulatum on the surface of liquid,collect the culture liquid,purify by salting out,precipitation by trichloroacetic acid,then dialyze and sterilize to prepare the bulk of purified protein derivative of histoplasmin.Perform skin and safety tests on the bulk in guinea pigs.Results The purified protein derivative of histoplasmin showed good safety and reached a purity of more than 80%.The skin reaction in guinea pigs induced by 1 μg/ml of purified protein derivative of histoplasmin was equivalent to that induced by the imported product of the same kind.Conclusion The purified protein derivative of histoplasmin might be used as a in vivo diagnostic reagent for infection of Histoplasma capsulatum and for histoplasmosis in clinic.

    2006 06 [Abstract][OnlineView][Download 90K]


@2012 Editorial Department of "Chinese Journal of Biologicals"