2017年03期目次

  • Screening of cryoprotectant for freeze-dried rabies vaccine(Vero cells) for human use

    LI Chun-yan;ZHAO Shu-jie;SUN Hong-liang;YAN Feng;ZHENG Hua;ZHAO Bo;CHAI Bo-ya;YANG Yi;Changchun Institute of Biological Products Co.,Ltd.;

    Objective To screen the cryoprotectant suitable for freeze-dried rabies vaccine(Vero cells) for human use.Methods Six formula of cryoprotectants were developed by changing the contents of three main components, i. e. Dextran40, Trehalose and human serum albumin(HSA), with which the bulk of rabies vaccine(Vero cells)for human use of the same batch was prepared into six batches of final bulks, then lyophilized under the same condition, observed for appearance, and tested for potency and heat stability, based on which the formula of cryoprotectant was optimized. Six batches of final products were prepared to verify the suitability of the optimized cryoprotectant, of which three batches were verified for long-term stability. Results The optimal cryoprotectant consisted of 4% trehalose, 3% Dextran 40, 1%sucrose, 0. 5% mannitol and 2% HSA. All the indexes of six batches of final products of rabies vaccine met the requirements in Chinese Pharmacopoeia(Volume Ⅲ, 2010 edition), of which three batches showed stable titer after storage at 2 ~ 8 ℃ for 12 months. Conclusion The cryoprotectant for freeze-dried rabies vaccine for human use was optimized, which showed high stability.

    2017 03 v.30 [Abstract][OnlineView][Download 136K]

  • Immunogenicity of Haemophilus influenzae type b conjugates with different distributions of molecular sizes

    SUN Shu-xue;CHEN Zhong-wei;MU Shan-shan;WANG Zhen-ye;LIU Xiao-lei;WANG Zhen;LIU Feng;MF Hong-chu;CHU Zi-qing;Beijing Chengda Tianhe Biotechnology Co.,Ltd.;

    Objective To evaluate the immunogenicity of Haemophilus influenzae type b(Hib) conjugates with different distributions of molecular sizes and optimize the molecular size of the conjugate.Methods The Hib capsular polysaccharides(PRP) of various activation grades were covalently linked to the carrier protein CRM197 by amine reduction to generate a series of conjugates with different molecular sizes(K_D),of which the bulk was separated by gel filtration chromatography,and the fractions with K_Dvalues of 0.05 ~ 0.25,0.25 ~ 0.45 and more than 0.45 were collected.BALB / c mice were immunized with these conjugates,of which the specific Ig G titers in sera were measured by indirect ELISA,using a commercial vaccine as control.Results The antibody levels induced by two and three doses of conjugate with K_Dvalue of 0.35 was equivalent to that with K_Dvalue of 0.01(P = 0.166 and P = 0.882),while was higher than that with K_Dvalue of 0.53(P = 0.008 and P = 0.031),and also higher than that in control vaccine group.Similarly,the specific Ig G titers induced by two doses of conjugate with K_Dvalue of 0.25 was significantly higher than that with K_D value of 0.48(P = 0.037).However,for the conjugate components with different molecular sizes,the antibody levels induced by two and three doses of components with K_Dvalues of 0.05 ~ 0.25 were the highest,followed by those with K_D values of 0.25 ~ 0.45,and those by the components with K_Dvalues of more than 0.45 were the lowest.Conclusion The immunogenicity of conjugates with large molecular sizes was superior to that with small molecular sizes.However,the optimal K_Dvalue for preparation of Hib conjugate vaccine was 0.20 ~ 0.35.

    2017 03 v.30 [Abstract][OnlineView][Download 281K]

  • Expression of human Granzym B fusion protein in prokaryotic system and preparation of its immune serum

    WANG Wen-huan;FENG Fang-fang;CAI Yi-qi;ZHANG Chan-qiong;CEN Dan-wei;SONG Yi-ling;GUO Gang-qiang;ZHANG Li-fang;LI Wen-shu;Department of Microbiology and Immunology,Wenzhou Medical University;

    Objective To express human Granzym B(Gzm B) fusion protein in prokaryotic system and prepare its specific immune serum.Methods Recombinant prokaryotic expression vector p ET32a-Gzm B was constructed and transformed to E.coli BL21(DE3) for expression under induction of IPTG.The expressed recombinant Gzm B(r Gzm B)protein was purified by Ni-NTA agarose affinity chromatography,with which BALB / c mice were immunized s.c.in several sites for 3 times.The serum samples of mice were collected 2,4 and 6 weeks after the first immunization and determined for Gzm B specific Ig G by indirect ELISA,based on which the maximum dilution of antibody was determined.Immunodot test was used to further confirm the specific binding of serum polyclonal antibody to Gzm B.Results Restriction analysis and sequencing proved that recombinant plasmid p ET32a-Gzm B was constructed correctly.The relative molecular mass of expressed recombinant Gzm B was 48 000.The specific Ig G level against Gzm B in sera of mice immunized with purified r GZm B increased as time went on and reached the peak value at week 4 after the first immunization,and was maintained at a certain level afterwards.The maximum dilution of antibody was 1 ∶ 1 600.The prepared polyclonal antibody in immune serum showed specific binding to Gzm B.Conclusion Recombinant Gzm B protein and its specific polyclonal antibody were successfully prepared,which laid a foundation of further study on the application of Gzm B to target killing or target detection.

    2017 03 v.30 [Abstract][OnlineView][Download 518K]

  • Comparison of efficiencies of various urinary cells in induction of pluripotent stem cells

    SHI Liang;CUI Ya-zhou;ZHOU Xiao-yan;LUAN Jing;HAN Jin-xiang;School of Medicine and Life Sciences,University of Jinan-Shandong Academy of Medical Sciences;

    Objective To compare the efficiencies of urinary cells(UCs) of various origins and various batches in induction of pluripotent stem cells(PSCs).Methods Twenty-five urine samples produced at night and 81 urine samples produced at daytime(31 from male and 50 from female volunteers) were collected,and induced by reprogramming method without using virus,serum,feeder and oncogene c-MYC to obtain induced PSCs(i PSCs).The reprogramming efficiency was estimated by alkaline phosphatase(AP) staining.Results The isolation rates of UCs from the urine samples produced at night and at daytime were 8% and 44% respectively,which showed significant difference(P < 0.05).Significant differences were observed in the reprogramming efficiencies of UCs of various batches from the same origin and in those from various origins(P < 0.05).Conclusion The urine during the first micturition was unsuitable for collection of UCs,and various UCs showed influence on the reprogramming efficiency.It laid a foundation of study on increasing the induction efficiency.

    2017 03 v.30 [Abstract][OnlineView][Download 606K]

  • Expression of bone morphogenetic protein 10 in Chinese hamster ovary cells and purification and biological activity of expressed product

    ZHENG Xiong-fei;JIN Jian;GONG Xiao-mei;LI Wen-jun;XU Dong-sheng;WAN Ai-ni;Department of Drug Design and Molecular Pharmacology,Jiangnan University;

    Objective To express the full-length gene of bone morphogenetic protein 10(BMP10) in Chinese hamster ovary(CHO) cells,purify the expressed product and determine its biological activity in order to obtain bioactive BMP10 molecules.Methods A 6 × his-tag and DDDDK(enterokinase recognition site) were inserted into the part between propeptide and mature chain gene segment of recombinant human BMP10(rh BMP10),and the constructed recombinant plasmid p MH3-rh BMP10-histag-DDDDK was transformed to CHO cells by electroporation.The recombinant cells for stable expression of target protein were screened from single clones with 500 μg / ml G418 for suspension domestication expression for 8 d,of which the culture supernatant was collected and purified by anion exchange column and nickel column chromatography.The cell activity in vitro of the purified protein was determined by q-PCR.Results Western blot showed three bands with relative molecular masses of about 48 000,96 000 and 190 000 respectively,which were consistent with those of monomer and polymers of target protein.The target protein after digestion with restriction enzyme stimulated the up-regulation of Smad6 expression in P19 cells effectively.Conclusion The presence of propeptide plays an important role in the activity of BMP10.The modified BMP10 was expressed in CHO cells,which showed a certain biological activity.The study provided a novel route to preparation of active full-length BMP10 dimer.

    2017 03 v.30 [Abstract][OnlineView][Download 1054K]

  • Phylogenetic relationship of heamagglutinin protein gene of measles virus in Jilin Province during 2001～2014

    SHAN Yuan-chun;WANG Shuang;ZHOU Jian-hui;WU Dong-lin;YANG Yao;TIAN Xin;WEI Lei-lei;CHENG Tao;CAO Feng-rui;Jilin Provincial Center for Disease Control and Prevention;

    Objective To analyze the phylogenetic relationship of hemagglutinin(H)protein gene of measles virus in Jilin Province during 2001 ~ 2014.Methods A total of 35 measles virus strains in Jilin Province were selected as representative strains,from which H protein DNA was amplified fractionally by RT-PCR,sequenced,spliced by DNASTAR software,and analyzed for relationship and homology by Mega 4.0 and Bioedit software.Results All the 35 strains belonged to H1 a sub-genotype of H1 genotype and spread cross two clades distinctly.The homologies of nucleotides and amino acids of H protein of the 35 strains were 98.2% ~ 99.8% and 95.2% ~ 99.8% respectively.However,the homologies of nucleotides and amino acids were 98.8% ~ 99.6% and 96.7% ~ 98.8% to China93-4 / H1 a strain,and 94.5% ~ 95.2%and 84.7% ~ 86.7% to Shanghai-191 strain,respectively.Conclusion The phylogenetic relationship of H protein of measles virus in Jilin Province during 2001 ~ 2014 was farther and farther away from China93-4H1 a strain,while the homologies of nucleotide and amino acid to China93-4 / H1 a and Shanghai-191 strains decreased year by year,indicating that the protein in Jilin Province should be monitored and analyzed continuously.

    2017 03 v.30 [Abstract][OnlineView][Download 811K]

  • Effect of Salmonella on proliferation of mouse gastric carcinoma MFC cells and expression of mouse β-defensin-2

    CHEN Jing-xia;ZHANG Bo-ning;SUN Peng;NIU Yan;ZHAO Fang-xin;LU Jing-kun;ZHANG Xuan;ZHAO Peng-wei;Department of Anesthesiology,Gucheng Hospital of Hebei Provence;

    Objective To investigate the effect of Salmonella on proliferation of mosue gastric carcinoma MFC cells and expression of mouse β-defensin(m BD)-2.Methods The proliferation level of MFC cells treated with Salmonella at various concentrations(10,10~2,10~3 and 10~4 bacteria / ml) for various(12,24 and 48) hours were determined by MTT assay,based on which the concentration and time of Salmonella for inhibition of MFC cell proliferation were optimized.The apoptosis of MFC cells treated with Salmonella for various(12,24 and 48)hours was determined by TUNEL method.MFC cells were treated with Salmonella at concentrations of 10,10~2,10~3 and 10~4bacteria / ml for 4,8 and 12 h separately,in which the expression levels of m BD-2 m RNA and protein were determined by PCR and Western blot respectively.Results The proliferation of MFC cells was inhibited significantly after treatment with Salmonella at a concentration of 103 bacteria / ml for 24 h,while obvious apoptosis was observed.Both the expression levels of m BD-2 m RNA and protein reached the maximum in MFC cells after treatment with Salmonella for 8 h.Conlusion The expression level of m BD-2reached the maximum in MFC cells after treatment with Salmonella at a concentration of 103 bacteria / ml for 8 h,which laid a foundation of further study on mechanism of production of m BD-2.

    2017 03 v.30 [Abstract][OnlineView][Download 850K]

  • Cloning and functional identification of glycine betaine-cholinecarnitine transporter opuD gene from Halobacillus Y5

    WANG Yan-hong;CAO Ning;JIA Gui-yan;YU Li-yun;JIANG Ju-quan;College of Life Science,Northeast Agricutural University;

    Objective To clone the opu D gene of glycine betaine-choline-carnitine transporter(BCCT)from Halobacillus Y5,analyze the gene structure and protein function,and confirm the function by salt and alkaline tolerance test.Methods The total genomic DNA of Halobacillus Y5 partially digested with Sau3 A Ⅰ was ligated to linear p UC18 vector digested with Bam H Ⅰ and dephosphorylated with bacterial alkaline phosphatase,transformed to competent E.coli KNabc by electrotransformation and spread onto solid LBK medium containing 0.2 mol / L sodium chloride,agar and 50 μg / ml ampicillin.The positive colonies were screened,from which recombinant plasmid was extracted,analyzed for bioin-formatics and tested for salt and alkaline tolerance.Results A new salt and alkaline tolerant gene BCCT HY5_opu D was cloned from Halobacillus Y5,which enabled KNabc to grow in LBK medium containing 0.2 mol / L sodium chloride and5 mmol / L lithium chloride at p H 8.0.The protein encoded by the gene showed different sequence and structure from those of Opu D transporters reported previously.Conclusion A novel BCCT opu D gene was cloned successfully,which provided a theoretical basis for development and application of salt and alkaline tolerant gene of moderately halophilic bacteria.

    2017 03 v.30 [Abstract][OnlineView][Download 3328K]

  • Effect of S3307 on anti-oxidant enzyme activity and DNA methylation level of coix seedlings under drought stress

    XIANG Jun-liang;HUANG Yu-lan;YIN Kui-de;LIU Zhao-bing;XU Ye-mei;FENG Jing-shu;REN Hai-juan;Faculty of Biological Life,Heilongjiang Bayi Agricultural University;

    Objective To investigate the effect of S3307 on anti-oxidant enzyme activity and DNA methylation level of Coix seedlings under drought stress.Methods The coix cultivar,"No 5 Yiliao",was cultured to the"two leaves and one core"stage,and divided into three groups.The coix seedlings in control group were treated with Hoagland medium,while those in PEG group with the nutrient medium containing 25% PEG,and those in PEG + S3307 group were pre-treated with S3307 then treated with 25% PEG.The morphological indexes of seedlings,such as height of seedling,diameter of stem,length and number of root in various groups were observed,while the activities of peroxidase(POD),superoxide dismutase(SOD)and catalase(CAT)were determined,and the DNA metylation levels were measured by HPLC.Results After pre-treatment with S3307,the height of seedlings and length of root decreased significantly(P < 0.05),while the diameter of stem and root cap ratio as well as the activities of SOD,CAT and POD increased significantly(P < 0.05).The percentage of 5-m C in leaves of seedlings increased by 34.29% as compared with that in control group,of which the increased extent decreased by 12.99% as compared with that in PEG group.Conclusion S3307 relieved the peroxide damage of membrane lipid and increase of DNA metylation level in coix seedlings under drought stress,which promoted the growth of seedlings.It laid a foundation of study on drought-relief mechanism of coix.

    2017 03 v.30 [Abstract][OnlineView][Download 804K]

  • Soluble prokaryotic expression,purification and antiviral activity of recombinant bovine interferon α

    LI Shu-qi;ZHAO Jun;WANG Li-li;YU Hai-yang;GAN Lin;WANG Ming-li;Anhui Jiuchuan Biotechnology Co.Ltd;

    Objective To highly express soluble recombinant bovine interferon-α(r Bo IFNα) of buffalo in E.coli,purify the expressed product and determine its antiviral activity.Methods IFNα gene was amplified from the total RNA of liver of buffalo by RT-PCR and inserted into prokaryotic expression vector p ET-32 a.The constructed recombiannt plasmid p ET-32a-IFNα was transformed to competent E.coli BL21(DE3)for expression under induction of IPTG.The expressed product was identified by SDS-PAGE and Western blot,purified by His Trap~(TM) HP nickel ion affinity chromatography,and determined for antiviral activity by cytopathic inhibition test in the vesicular stomatitis virus(VSV) / MDBK titration system.Results Sequencing result showed that the full-length of IFNα gene of buffalo was 498 bp,sharing a homology of100% with the bovine IFNα subtype C gene reported in Gen Bank.The expressed r Bo IFNα mainly existed in a soluble form and contained about 63.8% of total somatic protein.The purified r Bo IFNα reached a purity of 98.52% and showed specific binding to polyclonal antibody against bovine IFNα,of which the specific activity was(3.6 ± 0.25)× 10~6 U / mg.Conclusion Soluble r Bo IFNα was successfully expressed in E.coli,and the purified expressed protein showed high antiviral activity,which laid a foundation of clinical application of r Bo IFNα.

    2017 03 v.30 [Abstract][OnlineView][Download 1099K]

  • Preparation of immunomagnetic beads of enterohemorrhagic E.coli O157

    LIANG Bing;ZHANG Ya-jie;JI Xue;ZHU Ling-wei;GUO Xue-jun;LIU Yan-jing;LIU Jun;ZHOU Wei;LI Li-hong;FENG Shu-zhang;SUN Yang;Institute of Military Veterinary,Academy of Military Medical Science,Key Laboratory of Jilin Province for Zoonosis Prevention and Control;

    Objective To prepare the immunomagnetic beads(IMB)which enriched enterohemorrhagic E.coli(EHEC)O157 effectively and specifically.Methods Immunomagnetic beads were prepared with monoclonal antibody against E.coli O157 and five kinds of magnetic beads.The optimal magnetic beads were determined by comparing the antibody surplus in supernatant after coupling.The condition for preparation,such as time and temperature for activation,p H of coupling buffer as well as temperature and time for coupling,were optimized.The sensitivity and specificity of the immunomagnetic beads for detection and isolation of E.coli O157 were evaluated and compared with those of imported beads.Results The most appropriate magnetic beads with the highest coupling efficiency were carboxyl magnetic beads at diameters of 0.5 ~ 1.0 μm.The optimal time for activation after addition of NHS was 30 min,while the optimal temperature p H of 0.01 mol / L PBS as coupling buffer were 4 ℃ and 7.4,and the optimal temperature and time for coupling were 37 ℃ and 120 min,respectively.The sensitivity of E.coli O157 immunomagnetic beads reached 20 CFU / ml.When the concentration of E.coli O157 was 1.4 × 10~3 CFU / ml in the mixed suspension of different bacterial species,the specific capture ratio was about 90% by the prepared immunomagnetic beads.Conclusion The condition for coupling of carboxyl magnetic beads to E.coli O157 was optimized.The prepared immunomagnetic beads enriched E.coli O157 specifically,simply and rapidly,with higher capture efficiency,which might be used for the isolation and identification of EHEC O157 in clinical samples or food samples.

    2017 03 v.30 [Abstract][OnlineView][Download 241K]

  • Safety and immunogenicity of freeze-dried rabies vaccine(Vero cells) for human use in healthy population at ages of 10～60 years

    HUANG Li-rong;LV Teng-rong;PAN Gui-qiu;CAO Shou-chun;ZHANG Wei;HOU Yu-ting;LI Ming-li;LI Yan-ping;Center for Prevention and Control of Diseases;

    Objective To evaluate the safety and immunogenicity of freeze-dried rabies vaccine(Vero cells) for human use in healthy population at ages of 10 ~ 60 years.Methods A randomized(1 ∶ 1),blind,positive-controlled trial involving 1 200 healthy subjects at 10 ~ 60 years of age was conducted in Cenxi City and Cangwu County of Guangxi Zhuang Autonomous Region.The subjects were immunized with freeze-dried rabies vaccine(Vero cells)for human use on days 0,3,7,14 and 28,using a commercial product of the same kind as control.Local and systemic adverse reactions were observed within 42 d,while serum samples were collected 14 and 42 d after the first immunization and determined for neutralizing antibody against rabies virus by rapid fluorescence focus inhibition test(RFFIT),based on which the GMC of antibody was calculated.Results A total of 1 199 subjects were observed for safety,of whom the systemic adverse reaction rates in trial(600 cases)and control(599 cases)groups were 12.33% and 18.03% respectively,which showed significant difference(P < 0.05).The common symptom of systemic reactions was fever,of which the incidence rates in trial and control groups were 8.00% and 13.19% respectively.However,the local adverse reaction rates in trial and control groups were 5.33% and 10.52% respectively,which showed significant difference(P < 0.05).The symptoms of local adverse reactions were mainly pain,of which the incidence rates were 4.17% and 8.85% respectively.A total of1 147 subjects were observed for immunogenicity,of whom the antibody positive conversion rates in trial(571 cases)and control(576 cases) groups were 100.00% and 99.83% respectively on day 14 and both 100.00% on day 42 after the first immunization,which showed no significant difference(P > 0.05).The GMCs of antibodies were 8.94 and 7.96 IU / ml on day 14,and 7.26 and 15.04 IU / ml on day 42 after the first immunization,respectively,which showed significant difference(P < 0.05).Conclusion The freeze-dried rabies vaccine(Vero cells)for human use showed high safety and immunogenicity.

    2017 03 v.30 [Abstract][OnlineView][Download 213K]

  • Epidemiological investigation on an outbreak of 3 cases of measles infected in hospital

    REN Ya-ping;PENG Yi;FEI Yi;XIAO Shao-tan;Pudong New Area Centre for Disease Control and Prevention;

    Objective To investigate the causes of an outbreak of measles in a pediatric liver transplantation ward of a hospital in Shanghai,China.Methods Medical data of 3 cases of measles in a pediatric liver transplantation ward from May 4 ~ 15,2014 were analyzed retrospectively.Results All the measles virus antibody Ig M were positive in 3 children.The time durations of hospital stays of 3 children were longer than the longest latent period of measles(21 d),while the onset time intervals were shorter than the shortest latent period of measles(7 d).Conclusion This event was a hospital infection outbreak.However,no obvious source of common infection was found.The early diagnosis of measles in the children with basic diseases should be paid more attention.The pre-inspection on fever and ventilation disinfection in the ancillary departments should be strengthened to avoid the cross-infection of measles cases.

    2017 03 v.30 [Abstract][OnlineView][Download 108K]

  • Development of a method for determination of antibody-dependent cell-mediated cytotoxicity of recombinant monoclonal antibody against TNF-α

    LV Li-juan;CHEN Bao-han;ZHOU Dong-mei;XU Jun;SUEN Wen-zheng;YANG Bin;Sunshine Lake Pharma Co.,Ltd.;

    Objective To develop a method for determination of antibody-dependent cell-mediated cytotoxicity(ADCC)of recombinant monoclonal antibody(m Ab) against tumor necrosis factor alpha(TNF-α).Methods The ADCC of recombinant m Ab against TNF-α was determined using serially diluted innovator original drug(m Ab-O) drug or its biosimilar candidate(m Ab-S)combined with normal human peripheral blood mononuclear cells(PBMCs)as effector cells and a CHO cell line expressing membrane bound TNF-α as target cells.The median inhibitory concentration(IC_(50))of m Ab and the relative biological activity of m Ab-S were calculated by four-parameter equation.Under the optimized experimental condition,the method was verified for specificity and precision.Results The cytotoxicity of m Ab against TNF-α was in a dose-dependent mode,which was consistent with the result acquired from the four-parameter equation.The dose-response curve was in a typical S shape on semilogarithmic coordinate paper,with a R2 value of more than 0.99.The optimal condition of each critical parameter of this method when using PBMCs from the same individual source as effector cells was as follows: the number of target cells was 4 × 10~4 cells / well,while the ratio of effector cells to target cells was 25 ∶ 1,the incubation time of antibody with target cells was 1 h,and the induction time was 4 h.The method showed good specificity.The ADCC of m Ab-S was similar to that of m Ab-O(50% ~ 200%) using PBMCs from the same individual source as effector cells,while the RSD in reproducibility test on relative activity was 4.64%.However,the RSD of test results by one person in 2 working days at an interval of 3 d was 12.50%,while than by two persons in the same working day was 8.21%,both of which met the acceptance criteria for method validation.Conclusion A method for determination of ADCC of recombinant m Ab against TNF-α was successfully developed,which showed high specificity and precision,and might be used for evaluation of ADCC of the m Ab and commercial innovator original drugs.

    2017 03 v.30 [Abstract][OnlineView][Download 419K]

  • Effect of methods for sample treatment on subsets and function analysis of T lymphocytes

    REN Jing;ZHANG Yao-fang;WANG Gui-qin;FAN Wei-ping;Department of Immunology and Microbiology,Shanxi Medical University;

    Objective To investigate the effect of three methods for sample treatment,i.e.Ficoll-Hypaque discontinuous density gradients as well as erythrocyte lysis before and after antibody labeling,on the subsets and function analysis of T lymphocytes.Methods The samples were prepared by three methods and determined for surface molecules and subsets by flow cytometry,for cell survival rate by trypan blue staining,and for cell activity by Con A stimulation.Results The percentages of subsets of T lymphocytes in samples treated by erythrocyte lysis showed no significant difference with that by Ficoll-Hypaque discontinuous density gradient centrifugation(P > 0.05),while cell survival rate and activity were higher(P < 0.05).However,the test results of samples treated by erythrocyte lysis before and after antibody labeling showed no significant difference.Conclusion Discontinuous density gradient centrifugation was more suitable for scientific research,while erythrocyte lysis after antibody labeling for clinical laboratory.

    2017 03 v.30 [Abstract][OnlineView][Download 615K]

  • Scale-up culture of Marc-145 cells by using microcarriers in various bioreactors and propagation of PRRSV

    REN Fei;FENG Er-kai;YIN Mo-li;LUAN Yang;CHENG Yue-ning;ZHANG Miao;YU Jing-jie;SHENG Cheng-cheng;CHENG Shi-peng;CHEN Li-zhi;Jilin Te Yan Biological Technology Co.,LTD;

    Objective To investigate the condition for scale-up culture of Marc-145 cells by using microcarrires in various bioreactors and propagate porcine reproductive and respiratory syndrome virus(PRRSV).Methods Marc145 cells were cultured in primary bioreactor(BC-7L) containing 4 g / L cytodex-1 for 72 h,then sterilized,rinsed,digested with trypsin and inoculated to secondary bioreactor(BC-14L) for propagation of the cells to 5 times of the original volume.PRRSV(TJM-F92 strain) was inoculated at a MOI of 0.05,and the supernatant and suspension samples were collected24,36,48,60 and 72 h after inoculation and determined for virus titer(TCID50).Results Marc-145 cells reached a density of 28.7 × 10~5 cells / ml 72 h after culture in primary bioreactor,and 24.9 × 105 cells / ml 72 h after culture in secondary bioreactor after digestion with trypsin.The virus titer reached a peak value of 108.19TCID5036 h after inoculation,which showed no significant difference in supernatant and suspension.Conclusion It was feasible to perform a scale-up culture of Marc-145 cells from BC-7L to BC-14 L bioreactors,which laid a foundation of optimization of novel production procedure and large-scale production of live PRRSV vaccine.

    2017 03 v.30 [Abstract][OnlineView][Download 2907K]

  • Identification,isolation and purification of transformed product of gypenoside XLIX by thermophilic glycosidase Fpendo5A

    YU Bo-hao;LI Nan;ZHAO Huan-xi;LI Jing;ZHENG Fei;DAI Yu-lin;LI Zhuo;LIU Shu-ying;YU Shan-shan;Changchun University of Chinese Medicine;

    Objective To identify,isolate and purify the transformed product of gypenoside XLIX by thermophilic glycosidase Fpendo5 A.Methods Gypenoside XLIX was transformed with thermophilic glycosidase Fpendo5 A,and the transformed product was identified by HPLC,rapid resolution liquid chromatography-quadrupole-time of flight-mass spectrometry / mass spectrometry(RRLC-Q-TOF MS / MS),then isolated and purified by fully preparative high performance liquid chromatography(HPLC).Results Single product was obtained 48 h after transformation of gypenoside XLIX by thermophilic glycosidase Fpendo5 A at 65 ℃ and p H 6,which was gylongiposide Ⅰ generated by hydrolysis of glycosidic bond at site C20 of gypenoside XLIX with Fpendo5 A.The purity and yield of the transformed product were95.42% and 21.6% respectively after purification.Conclusion Gypenoside XLIX was successfully transformed by thermophilic glycosidase Fpendo5 A.The purified transformed product reached a high purity and a high yield,which laid a foundation of further study on pharmacological activity as well as the relevant basic research.

    2017 03 v.30 [Abstract][OnlineView][Download 525K]

  • Feasibility of determination of immunogenicity of Hib-CRM197 conjugate vaccine by immature rabbit method

    CAO Fang;LIU Hai-chang;LI Qiang;WANG Liang-chao;PAN Ling-ling;YANG Ling;Zhejiang Weixin Biological Pharmaceutical Co.,Ltd.;

    Objective To evaluate the feasibility of determination of immunogenicity of Hib-CRM197 conjugate vaccine by immature rabbit method.Methods Rabbits were injected s.c.with Hib-CRM197 conjugate vaccine at a dosage of12.5 μg polysaccharide on days 0,7 and 14 respectively,using physiological saline as negative control and the comme-rcial products of the same kind from three manufacturers as positive control.Serum samples were collected before immunization and 1 ~ 2 weeks after the last immunization,and determined for antibody level by ELISA.Results The serum antibody levels(A_(490) values) of rabbits immunized with Hib-CRM197 vaccine and those in positive control group were more than 5 times higher than those before immunization,and more than 2 times higher than the cutoff value.Conclusion It is feasible to determine the immunogenicity of Hib-CRM197 conjugate vaccine by immature rabbit method.

    2017 03 v.30 [Abstract][OnlineView][Download 264K]

  • Analysis of fatty acid composition of Rosa davurica Pall. by gas chromatography-mass spectrometry

    JIA Peng-yu;LI Liang-yu;LI Chao-yang;SONG Da-wei;National Coarse Cereals Engineering Research Center,Heilongjiang Bayi Agricultural University;

    Objective To analyze the fatty acid composition of Rosa davurica Pall.from Greater Khingan in the northern region of Heilongjiang Province,China by gas chromatography-mass spectrometry(GC-MS).Methods Rosa davurica Pall.were collected on the same day and stored under the same condition,of which fatty acids were extracted from different parts of flowers(petal,stamens,torus and calyx) by soxhlet extraction with petroleum ether,and analyzed quantitatively and qualitatively by means of GC / MS after boron trifluoride-methyl esterification.Results The fatty acid profiles varied significantly in petal,stamens,torus and calyx of Rosa davurica Pall.A portion of 14,15,13 and 20 kinds of fatty acids were detected in petal,stamens,torus and calyx of Rosa davurica Pall.,of which the relative contents of unsaturated fatty acids were 63.78%,59.1%,47.02% and 40.3%,respectively.Conclusion The oil fatty acid compositions in various parts of Rosa davurica Pall.were different,while petal was rich in unsaturated fatty acids.The study provided a reference for improving the quality of rose essential oil and utilization of Rosa davurica Pall.resources.

    2017 03 v.30 [Abstract][OnlineView][Download 764K]

  • Comparison of requirements for quality control of intravenous immunoglobulin

    LEI Min;MA Li;LI Chang-qing;Institute of Blood Transfusion,Chinese Academy of Medical Sciences and Peking Union Medical College;

    Intravenous immunoglobulin(IVIG) p H 4 have been widely used for the treatment of a variety of immunodificiency and autoimmune diseases.Strict quality control is important to ensuring the clinical efficacy of IVIG.Although some rigorous quality control regulations on IVIG have already been described in the European Pharmacopoeia,United States Pharmacopoeia and Chinese Pharmacopoeia,they should be replenished,refined and improved due to the complicate composition as well as the increasing understanding of action mechanism and increasing indications in clinic of IVIG.In this paper,the requirements for quality control of IVIG in the international mainstream pharmacopoeia are compared,while some new methods for quality control of IVIG reported in recent years are reviewed,so as to provide a reference for the relevant researchers and manufacturers of blood products.

    2017 03 v.30 [Abstract][OnlineView][Download 177K]

  • Progress in research of biological effect of interleukin 12

    ZHOU Peng;BAI Qin-qin;CHEN Li-li;School of Public Health,University of South China;

    As a heterodimeric cytokine,interleukin-12(IL-12)is an initial factor in cellular immune response.IL-12 can activate T lymphocytes and natural killer cells,and stimulate their proliferation and differentiation.IL-12 can also regulate cellular immunity and induce T lymphocytes or NK cell to secrete interferon(IFN)γ.This paper reviews the role of IL-12 in anti-infection,anti-tumor and vaccine development.

    2017 03 v.30 [Abstract][OnlineView][Download 189K]

  • Progress in research on anticancer drugs derived from microorganisms

    WEI Yan-hong;YAO Chen-guang;XI Cai-li;HU Kang-hong;Sino-Germany Biomedical Center,Hubei University of Technology,Hubei Collaborative Innovation Center for Industrial Fermentation;

    The extremely rich microorganisms in the world contain numerous natural products with different chemical structures,such as proteins,polysaccharides,anthracyclines,organic acid esters,terpenes,alkaloids,macrolides and enediynes,many of which show anti-tumor effects and have been developed as anti-cancer drugs already used in clinical trial.In this paper,the taxonomy of anti-cancer agents derived from terrestrial,marine and symbiotic microorganisms,challenge and relevant strategies in development as well as advantages and prospects of anti-cancer drugs derived from microorganisms.

    2017 03 v.30 [Abstract][OnlineView][Download 223K]


@2012 Editorial Department of "Chinese Journal of Biologicals"