2021年11期目次

  • Progress in research on hand,foot and mouth disease-related enteroviruses and vaccines

    JIN Wei-ping;QIAN Sha-Sha;SHEN Shuo;Viral Vaccine Research Laboratory,Wuhan Institute of Biological Products Co.,Ltd.;

    Since the outbreak of hand,foot and mouth disease(HFMD)caused by enterovirus 71(EV-A71)in Fuyang City,Anhui Province in 2008,China and Southeast Asia have become epidemic areas of HFMD,which has also promoted the basic research and vaccine development of HFMD-related enteroviruses. In 2016,the monovalent inactivated whole virus EV-A71 vaccine was commercially launched,resulting in the decrease of the incidence,severe cases and mortality of HFMD caused by EV-A71 infection. However,the pathogen spectrum of HFMD-related enteroviruses has changed since 2012,with non-EV-A71 and non-CV-A16,including CV-A6 and CV-A10 and other serotypes,gradually become the main pathogens instead of EV-A71 or CV-A16. The monovalent inactivated EV-A71 vaccine is unable to protect HFMD caused by other serotypes. The development of a multivalent vaccine containing EV-A71 and several other major serotypes has become a consensus to prevent HFMD,control the spread of enteroviruses and delay the emergence of mutant and recombinant strains. The progress and challenge in research on the epidemiology,molecular biology of enteroviruses as well as the development of vaccines,especially multivalent vaccines,are reviewed in this paper.

    2021 11 v.34 [Abstract][OnlineView][Download 392K]

  • Stability of inactivated enterovirus type 71 vaccine at high temperature

    ZHOU Zhen-xin;YI Li;HU Yun-guang;YANG Jia-fang;HE Jia-ning;ZOU Jing;YUAN Yang-mei;CHEN Jing-xiu;LIANG Jing;DONG Shao-zhong;Institute of Medical Biology,Chinese Academy of Medical Sciences & Peking Union Mdeical College;

    Objective To evaluate the stability of inactivated enterovirus type 71(EV71)vaccine stored at(40 ± 2)℃and provide a reference for the efficacy and safety of the vaccine within the validity period when the cold chain is damaged with high temperature during storage and shipment. Methods Three batches(P1,P2 and P3) of inactivated EV71 vaccine qualified in releasing inspection after storage at 2 ～ 8 ℃ for 2 years(validity period)were placed at(40 ±2) ℃ for 0,24,48,72 and 96 h respectively,and determined for antigen content to understand the stability at high temperature. Another three batches(P4,P5 and P6)of inactivated EV71 vaccine under the same storage condition were placed at(40 ± 2)℃ for 24 h and subjected to control tests according the requirements in the Drug Registration Specification to understand the impact of high temperature on all quality indexes. Results After storage at(40 ± 2)℃ for 24 h,the antigen contents of vaccine of P1,P2 and P3 met the requirements in the Drug Registration Specification. The antigen contents showed decreasing tendencies with the increasing storage time at(40 ± 2)℃. Compared with that after storage for0 h,the antigen contents decreased insignificantly after storage for 24 h,while decreased significantly for 48 h,and were relatively stable after storage for 96 h. All the quality indexes of vaccine of P4,P5 and P6 met the requirements in the Drug Registration Specification after storage at(40 ± 2) ℃ for 24 h. Conclusion Inactivated EV71 vaccine showed a certain stability when exposed to(40 ± 2) ℃ for 24 h. It provides a reference for the study on quality of vaccine at extreme high temperature.

    2021 11 v.34 [Abstract][OnlineView][Download 230K]

  • Immune protective effect of Hp1188 protein of Helicobacter pylori

    HAN Fei;ZHANG Shuai-shuai;Department of Medical Laboratory,Chongqing University Three Gorges Hospital;

    Objective To investigate the immunoreactivity of Hp1188 protein of Helicobacter pylori and its protection in mice immunized by oral route and explore its significance in preparation of preventive vaccine. Methods Recombinant Hp1188 protein of H. pylori was purified and identified for immunoreactivity by Western blot. The corresponding antibodies in 100 clinical serum samples were determined by ELISA using the purified recombinant HP1188 protein as H. pylori antigen,of which the results were compared with those of urea breath test(UBT)and rapid urease test(RUT)to evaluate the feasibility of clinical application of the protein. Mice were immunized with purified Hp1188 protein,challenged with H. pylori Sydney strain 1(SS1),and determined for IgG,IgA and sIgA against Hp1188 in serum and intestinal mucus by ELISA. H. pylori culture,RUT and pathological examination were performed on gastric mucosa of mice to observe the quantity of H. pylori colonization and inflammation. Results The purified Hp1188 protein reached a purity of 90% and a concentration of 1. 0 mg/mL after purification,which could be recognized by sera of patients with H. pylolri infection as proved by Western blot. The results of serological test showed that the sensitivity and specificity of Hp1188 antibody kits were 93. 8% and 94. 3% respectively. The contents of IgG,IgA and sIgA in mice immunized with Hp1188 protein were significantly higher,while the count of clonized live H. pylori as well as the indexes in RUT and pathological examination were significantly lower,than those in PBS control group(each P < 0. 05),and the protective rate was 70%. Conclusion Hp1188 protein showed immune protective effect on mice,which provided an experimental basis for the development of H. pylori related vaccines.

    2021 11 v.34 [Abstract][OnlineView][Download 314K]

  • Prokaryotic expression and immunogenicity of truncated varicella-zoster virus glycoprotein E

    LI Chun-ming;WANG Li ;MA Cheng ;LIU Jia-yu;XU Yuan-hang;MU Zhong-mei;Research Department of Changchun Keygen Biological Products Co.,Ltd.;

    Objective To express varicella-zoster virus(VZV) glycoprotein E(gE) in prokaryotic cells and evaluate its immunogenicity. Methods The nucleic acid sequence of amino acids 25 ～ 537 of VZV surface glycoprotein E(gE25-537)was obtained from GenBank,based on which plasmid pUC-VZV-gE25-537-His was synthesized by codon optimization and used as a template for PCR. The amplified target fragment was cloned into vector pET-21 b,and the constructed recombinant plasmid pET-21 b-VZV-gE25-537-His was transformed to competent E. coli BL21(DE3)and induced with IPTG. The inclusion body was dissolved with urea,and the target protein was purified by nickel affinity chromatography. The purified protein was mixed with Freund adjuvant,with which mice were immunized s.c. in several sites for 3 times. Serum samples were collected 2 weeks after the last immunization and determined for neutralizing antibody titer. Results Restriction and sequencing proved that recombinant plasmid pET-21 b-VZV-gE25-537-His was constructed correctly. The target protein,with a relative molecular mass of about 62 000,mainly existed in a form of inclusion body,which reach a purity of 85% after purification and showed a specific binding to mouse monoclonal antibody against gE. The serum antibody titer of mice after immunization for 3 times reached 1 ∶ 54 954,while the neutralizing antibody titer was 1 ∶ 21. 75. Conclusion Truncated VZV-gE protein was expressed in prokaryotic cells, which showed good immunogenicity in mice after purification and might be used as a candidate antigen for preparation of herpes zoster vaccine.

    2021 11 v.34 [Abstract][OnlineView][Download 211K]

  • Effects of different cell culture media on critical quality attributes of human monoclonal antibody against tetanus toxin

    WANG Jian-feng;WANG Yan-yan;GUO Lan;YE Xing;NAN Jian-jun;AN Chen;SONG Lan-lan;CHEN Ji-jun;MAO Xiao-yan;Lanzhou Institute of Biological Products,Co.,Ltd,Engineering Research Center of Macromolecules Biological Drugs of Gansu Province,Vaccinal Engineering Technology Research Center of Gansu Province;

    Objective To investigate the effects of different media on growth of engineered cell strain and critical quality attributes of human monoclonal antibody(McAb)against tetanus toxin and screen the optimal medium. Methods Three commercial culture media,Growth A,Dynamis and T54,were used in fed-batch model,and the cell viability and viable cell density were determined. The antibody purity were analyzed by SEC-HPLC and CE-SDS,while the isoelectric point and charge heterogeneity by cIEF method. Results The viable cell density in Growth A medium was higher than,while the cell viability rate was between,those in Dynamis and T54 media. The purity of McAb in Growth A medium was higher than those in the other two media. However,there was no significant difference in the PI values of McAbs cultured in the three media. The number and proportion of acid peak of charge variation of antibody cultured in Growth A medium were less than,while the proportion of main peak was between,those in the other two media. Conclusion Growth A medium is more suitable for the cultivation of engineered cells for McAb against tetanus toxin.

    2021 11 v.34 [Abstract][OnlineView][Download 393K]

  • Delaying effect of water-soluble coenzyme Q10 on rotenoneinduced Parkinson disease in rat model and relevant mechanism

    FENG Chi;SA Ri-gai-qi-qi-ge;LI Hai-ning ;LV Yue ;LI Hui-lei ;DONG Wu;YANG Jing-feng;Inner Mongolia Key Laboratory of Toxicant Monitoring and Toxicant and Toxicology,Collage of Animal Science and Technology,Inner Mongolia University for Nationalities;

    Objective To investigate the delaying effect of water-soluble coenzyme Q10(CoQ10) on rotenone-induced Parkinson disease(PD)in rat model as well as the relevant mechanism. Methods Wistar rats were randomly divided into CoQ10 control,sunflower oil(vehicle) control,rotenone-treatment(PD),CoQ10 pretreatment + PD(CoQ10 B) and PD + CoQ10 post-treatment(CoQ10 A) groups,ten for each. The recovery effect of CoQ10 on rotenone-induced PD in model rats was evaluated by open field experiment,neuromotor function test,histopathological examination as well as the expression levels of oxidation stress-related genes and amino acid decarboxylase gene. Results CoQ10 recovered the decreased rearing numbers caused by rotenone in open field test and latency before failing time in hanging test,which showed a tendency of increasing maintain time in rotarod test. Histopathological examination showed that rotenone caused nuclear condensation and cell degeneration in partial neurons,while CoQ10 slowed the degeneration down. Rotenone increased the expression of Nrf2/HO-1 related genes and decreased those of TPH and 5-HT1 A receptor genes,while CoQ10 decreased the expressions of Nrf2/HO-1 related genes and restored those of THP and 5-HT1 A receptor genes.Conclusion Water-soluble CoQ10 showed a certain neuroprotective effect on rotenone-induced neuropathy.

    2021 11 v.34 [Abstract][OnlineView][Download 456K]

  • Identification and expression pattern analysis of class Ⅲ peroxidase gene family in Camellia sinensis(L.)

    SHI Xiao-xia;ZHANG Bao-hui;YAO Xin-zhuan ;LV Li-tang;The Key Laboratory of Plant Resources Conservation and Germplasm Innovation in Mountainous Region (Ministry of Education),Institute of Agro-Bioengineering;

    Objective To identify the members of class Ⅲ peroxidase(POD)gene family in Camellia sinensis(L.)and analyze its expression pattern. Methods The members of POD gene family in genome of C. sinensis(L.)were identified by bioinformatics method,of which the chromosome localization,gene structure,conservative protein motif,evolutionary relationship and colinearity relationship were determined,and transcriptome data in various C. sinensis(L.)tissues as well as in the C. sinensis(L.)after drought,salt and cold stresses were analyzed. Results A total of 122 POD genes were identified in the genome of C. sinensis(L.),which were unevenly distributed in the chromosome,with several tandem gene repeats. The structures and conservative motifs of genes from the same subfamily were basically in agreement. The POD genes were divided into five clusters based on evolutionary relationship. There were colinearity relationship in 23 pairs of genes. POD gene was highly expressed in root tissues,and the expression levels of partial genes increased with the increasing leaf maturity. However,various POD genes were differentially expressed after drought,salt and cold stresses.Conclusion POD gene played an important role in the growth and development as well as response to abiotic stress of C. sinensis(L.),which laid a foundation of further study on the function of POD genes.

    2021 11 v.34 [Abstract][OnlineView][Download 654K]

  • Bioinformatics of bta-miR-126-5p target gene in exosomes derived from mammary epithelial cells of dairy cows and its regulatory function

    XU Hao-tian;LIAN Shuai;WU Rui;WANG Jian-fa;College of Animal Science and Veterinary Medicine,Heilongjiang Bayi Agricultural University;

    Objective To predict the candidate target gene of bta-miR-126-5 p in exosomes derived from mammary epithelial cells of dairy cows and analyze its function by the relevant bioinformatics techniques so as to lay a foundation of functional verification of bta-miR-126-5 p. Methods The candidate target genes of bta-miR-126-5 p were predicted by using Targetscan and miRWalk online analysis system,on which GO enrichment analysis,protein interaction analysis and KEGG enrichment analysis were performed by using DAVID,STRING,KEGG and other biological information analysis systems. Results The sequence of mature region of bta-miR-126-5 p was highly conserved in various species. A total of203 bta-miR-126-5 p candidate target genes were screened,which mainly involved in the process of intracellular molecular function such as combinations of metal ions(especially zinc ion)and ATP. The enrichment of signaling pathways focused on the biosynthesis and degradation of fatty acids as well as metabolisms of purine,propionic acid salt and inositol phosphate. Conclusion The bta-miR-126-5 p may play an important role in the process of inflammation and physiological metabolism in cows by regulating candidate target genes and participating in relevant biological processes.

    2021 11 v.34 [Abstract][OnlineView][Download 410K]

  • Influence of super-activated platelet lysate on proliferation of osteoblasts and repairing effect on apoptosis

    CUI Tong;WU Qi-jian ;LIU Chun-xiang ;LIAN Hong-yu;ZHAO-Xue;LI Zi-tao;ZHANG Yi;Hongqi Hospital Affiliated to Mudanjiang Medical University;

    Objective To investigate the influence of super-activated platelet lysate(sPL)on proliferation of osteoblasts and repairing effect on apoptosis so as to determine whether it can be effectively used for bone healing. Methods Osteoblasts hFOB.1.19 were added with sPL at various concentrations(0%,5%,10% and 20%) and counted for 7 consecutive days,based on which a growth curve was plotted to analyze the effect of different concentrations of sPL on the proliferation of osteoblasts. An osteoblast apoptosis model was established by induction with transforming growth factor-β（TGF-β）. The model osteoblasts were treated with sPL at various concentrations(0%,5%,10% and 20%)for 36 h respectively,and determined for apoptosis rate by flow cytometry. Results The sPL at concentrations of 5% and 10%showed strong promoting effect on the proliferation of osteoblasts,while the effect of 5% sPL was superior to that of10% sPL. However,20% sPL showed no significantly promoting effect on the cell proliferation. The 5% and 10% sPL showed significantly repairing effect on the apoptotic osteoblasts as compared with 0% sPL(each P < 0. 01),while 20%sPL showed no significantly repairing effect(P > 0. 05). Conclusion The sPL can promote the proliferation of osteoblasts and repair apoptosis so as to promote the healing of bone tissue,of which the effect is satisfactory at a concentration of 5%.

    2021 11 v.34 [Abstract][OnlineView][Download 236K]

  • Collaborative calibration of national reference for human leukocyte antigen tissue match typing

    HU Ze-bin;SUN Bin-yu;LIU Xiang ;LAN Geng-xin ;SUN Jing;YU Ting;HUANG Jie;National Institutes for Food and Drug Control;

    Objective To collaboratively calibrate the national reference for human leukocyte antigen(HLA)tissue match typing. Methods Four laboratories with good qualification were selected to carry out the experiment according to the unified collaborative calibration scheme. PCR-SSO,PCR-SBT and NGS sequencing were used to calibrate 38 samples of the national reference. Each sample was determined by PCR-SBT sequencing and NGS sequencing for one time each,Meanwhile,the international HLA typing control panel(51 samples)was determined by PCR-SBT sequencing to control the quality of the experiment. Results All the HLA-A,B,C,DRB1 and DQB1 sites of 38 samples of the national reference were detected by four laboratories,and the typing results of the five sites at 2-bit and 4-bit resolutions were in agreement. Finally,4-bit high-resolution typing results were selected for the value of the national reference. All the typing results of HLA-A,B,C,DRB1 and DQB1 sites of 51 samples of international HLA typing control panel were consistent with those reported in IHWG. Conclusion The value of national reference for HLA tissue match typing is accurate,which may be used for the accuracy evaluation of HLA-A,HLA-B,HLA-C,HLA-DRB1,HLA-DQB1 gene typing detection reagents.

    2021 11 v.34 [Abstract][OnlineView][Download 139K]

  • A survey on cognition of poliomyelitis and its vaccination among parents of children at ages of not more than 6 years in Yunnan Province,China

    SHI Hao-yu;ZHONG Zhi-lei;WANG Jing-dong;LI Ying;LI Jia-xuan;CHEN Qiu-yu;YIN Xing-xiao;YANG Jing-si;Institute of Medical Biology,Chinese Academy of Medical Sciences & Peking Union Medical College;

    Objective To understand the status quo of parents′ awareness of poliomyelitis and its vaccination in Yunnan Province,China,and to explore effective methods of population-education for poliomyelitis eradication. Methods A questionnaire survey was conducted among parents of children at ages of not more than 6 years in 24 community health service centers in Yunnan Province by using the convenient sampling method. Results A total of 600 questionnaires were issued and 553 valid questionnaires were recovered. Vaccinators were main source of vaccine knowledge for the parents(70. 5%). The recommendation of the inoculating physician was the major influencing factor of choice of vaccine(68. 2%). However,the influencing factors of choice of vaccine were different in the parents with different characteristics,such as parent-child relationship,education ground,family income and whether the occupations of major family members were medically related. The correction rates of four aspects of knowledge about polio and its vaccine were different in the parents with different characteristics. There were significant differences in the choice of parents with different characteristics as to whether they will choose to receive IPV at their own expense. However,when the IPV at their own expense was chosen,the priority factors were different in the parents with different characteristics of parentchild relationship and family monthly income. Conclusion There are differences in children′ s parents′ cognition of poliomyelitis and its vaccination. According to parents′ situation and needs,targeted popularization education methods should be carried out to improve parents′ knowledge of poliomyelitis and its vaccine,so as to help maintain the poliomyelitis-free status in China.

    2021 11 v.34 [Abstract][OnlineView][Download 217K]

  • Surveillance of adverse events following immunization in Inner Mongolia Autonomous Region,China from 2018 to 2019

    LI Dong-ying;YAN Shao-hong ;TIAN Xiao-ling ;LEI Xia;Baotou Medical College;

    Objective To analyze the epidemiological characteristics of adverse events following immunization(AEFI)in Inner Mongolia Autonomous Region,China from 2018 to 2019 and provide a basis for improving the safety of vaccines and the quality of vaccination services. Methods The data reported in Inner Mongolia Autonomous Region from 2018 to2019 were collected through the China Immunization Program Information Management System and analyzed by using descriptively epidemiological methods. Results A total of 6 696 cases of AEFIs were reported from 2018 to 2019 in Inner Mongolia Autonomous Region,with a mean reported incidence of 48. 10/100 000 doses. The ratio of male to female in the reported cases was 1. 26 ∶ 1. Most(6 536,97. 61%)of the cases appeared in the children at ages of not more than 6 years,which were mainly reported from March to August. Of the cases,6 221(92. 90%) were general adverse reactions which were mainly fever(4 426 cases,31. 79/100 000 doses),while 418(6. 24%)were rare adverse reactions which were mainly allergic reactions(222 cases,2. 77/100 000 doses). The top three reported incidence rates of AEFIs were caused by group ACYW135 meningococcal polysaccharide vaccine,acellular DTP and live attenuated Japanese encephalitis vaccine. Conclusion The sensitivity of AEFI monitoring in Inner Mongolia Autonomous Region has declined,and the reported AEFIs are mainly general reactions. It is necessary to strengthen the training and guidance of AEFI monitoring.

    2021 11 v.34 [Abstract][OnlineView][Download 189K]

  • Determination of kit (ELISA) for screening material plasma of human COVID-19 immunoglobulin (pH4) for intravenous injection and establishment,validation and application of quantitative detection method

    ZHOU Zhi-jun;XING Yan-tao;LIANG Xiao-long;XIAO Long;ZENG Shuang-ying;LI Pu;DENG Kun;WANG Yue;YU Jian-hong;LI Tao-jing ;HU Yong;HAN Ren;LI Ce-sheng;Sinopharm Wuhan Plasma-derived Biotherapies Co.,Ltd.;

    Objective To screen five currently commercially available 2019-nCoV IgG antibody detection kits(A,B,C,D and E) by comparing the specificity,sensitivity and linear range,and establish and validate a quantitative detection method for material plasma of human COVID-19 immunoglobulin(pH 4)for intravenous injection. Methods The COVID-19 convalescent plasma with neutralizing antibody activity was used as the internal control standard and 2-fold serially diluted to seven dilutions(S1 ～ S7). According to the instructions of the kits from five manufacturers,the absorbances of S1 at the highest concentration were compared,while the linear ranges were compared by using a four-parameter fitting curve. A quantitative detection method was developed with three candidate kits and used for determination of partial representative negative and positive samples,and the correlation of results was evaluated. A kit was selected based on comprehensive consideration,with which the dilution of the internal control standard was assigned after serial dilution to fit the standard curve and establish a quantitative detection method. The established method was validated for linear range,accuracy and precision,and used for establishmetnt of a plasma screening mode. after verifying its linear range,accuracy and precision. Results The linear range of the internal control standard showed that six dilutions were detected by kit A,while the absorbance of S1 was 2. 8;five dilutions were detected by kit B,while the absorbance of S1 was 2. 4;five dilutions were detected by kit C,while the absorbance of S1 was 1. 4;three dilutions were detected by kit D,while the absorbance of S1 was 3. 3;however,five 5 dilutions were detected bu kit E,while the absorbance of S1 was 3. 3. By using kits A,B,and E with similar linear ranges,a quantitative detection method was established with the internal control standard,of which the R2 of the curve and the recovery rates of various dilutions met the relevant requirements. The representative negative samples tested by kit A showed high background and poor specificity. The correlations between the determination results by three kits and the neutralization test of the representative positive samples were more than 0. 9.Kit B was selected finally,by which the dilution of the internal control standard determiend with three batches for 4 times was assigned as 32,and the four-parameter fitting curve was linearly established,and the linear range of dilution was 1 ～16,with a R2 value of more than 0. 99,indicationg that the accuracy and precision were qualified. The plasma screening mode was establilshed as single test each sample,and the dilution of retest sample was determiend. Conclusion The kit from manufacturer B is optimized as a quantitative detection kit,which may be used for the material plasma screening of human COVID-19 immunoglobulin for intravenous injection after validation.

    2021 11 v.34 [Abstract][OnlineView][Download 373K]

  • Optimization of procedure for purification of group C meningococcal polysaccharide by chromatography

    SU Xiao-ye;GAO Lei;CHENG Ya-jun;LI Qian;TIAN Yu;LIU Gang;Bei Jingzhifei Lvzhu Biopharmaceutical Co.,Ltd.;

    Objective To optimize the technological condition for purification of group C meningococcal polysaccharide by chromatography so as to increase the column capacity of the chromatographic medium and decrease the application of sodium deoxycholate(SDC)on the premise of ensuring the separation effect and polysaccharide recovery rate. Methods Crude group C polysaccharide was pretreated in Tris-HCl buffer containing 1% SDC,then separated and purified by a series of chromatography columns packed with Capto Adhere and Capto DEAE and desalinated by ultrafiltration to obtain the bulk of polysaccharide. The effects of SDC content in buffer as well as crude polysaccharide concentration and volume for loading on the separation effect were investigated to optimize the technological condition. Under the optimal condition,the service life of chromatography column was investigated to facilitate the cleaning and regeneration of chromatography medium. Results When the the chromatographic flow rate was controlled at 90 cm/h,the loading concentration of crude polysaccharide increased to 5 g/L,the SDC content of mobile phase decreased to 0. 5%,and the loading volume increased to 25%. The residual SDC content after desalination by ultrafiltration was less than 10 mg/g polysaccharide.Chromatographic purification could be carried out for 5 times in succession,and the purification and separation effects of chromatographic column were basically unchanged. Conclusion Group C meningococcal polysaccharide can flow through the chromatography column. Under the optimal condition,chromatography may be used for the separation and purification of group C meningococcal polysaccharide during production.

    2021 11 v.34 [Abstract][OnlineView][Download 379K]

  • Development and verification of whole column imaging detectioncapillary isoelectric focusing electrophoresis for determination of isoelectric point of virus-like particles of recombinant enterovirus 71

    WANG Wen-wei;HE Cheng ;WANG Bei;LOU Jue-ren;Shanghai Wellvax Co.,Ltd.;

    Objective To develop and verify a method for determination of isoelectric point(pI) of virus-like particles(VLPs)of recombinant enterovirus(EV)71(EV71 VLPs)by whole column imaging detection-capillary isoelectric focusing(WCID-CIEF)electrophoresis. Methods The pI of recombinant human EV71 VLPs was determined by WCID-CIEF electrophoresis. The developed method was verified for specificity,accuracy,reproducibility and intermediate precision.Results In WCID-CIEF electrophoresis,the focusing was performed at 3 000 V for 5 min,and no urea was added when the samples were treated. Blank buffer showed no interference to the determination of pI of EV71 VLPs. The relative standard deviations(RSDs)of determination results of pI marker 5. 85 and pI marker 7. 05 in six repeat tests by WCIUDCIEF electrophoresis were 0. 18% and 0. 06% respectively,while that of EV71 VLPs was 0. 08%. However,the RSD of determination results of pI of recombinant human EV71 VLPs in 12 repeat tests by different personnel on different dates was 0. 14%. Conclusion The WCID-CIEF electrophoresis for determination of pI of recombinant human EV71 VLPs showed high specificity,accuracy,reproducibility and intermediate precision,which might be used for the quality evaluation of recombinant human EV71 VLPs.

    2021 11 v.34 [Abstract][OnlineView][Download 222K]

  • Establishment of MDCK suspension cell line in serum-free medium at high cell density in bioreactors

    ZHAO Cai-hong;WANG Mei-hao;LI Zi-liang;JIN Dong-wu ;MA Hua ;MA Zhong-ren;QIAO Zi-lin;CHEN Hong ;ZHANG Jia-you ;WANG Jia-min;Gansu Tech Innovation Center of Animal Cell,Biomedical Research Center,Northwest Minzu University;

    Objective To establish a MDCK cell line in serum-free suspension culture and analyze the growth characteristics,virus sensitivity,metabolic kinetics characteristics and the feasibility of linear amplification in bioreactor.Methods A MDCK suspension cell line was obtained by stepwise serum-low and serum-free suspension domestication,and determined for virus sensitivity. The cell growth characteristics,metabolic kinetics characteristics and optimal time for subculture of the suspension cells were investigated in 1. 2 L quadrupled parallel bioreactor,and the cells were subjected to linear scale-up culture in 5 and 75 L bioreactors. Results Adherent MDCK cells were stably cultured at a serum concentration of 3% for five passages and directly adapted to serum-free medium to obtain a MDCK suspension cell strain(named MDCK-XF03)with a resuscitation activity of 96. 8%,in which the influenza virus of various types grew well.The maximum proliferation concentrations of MDCK-XF03 cells cultured at different initial inoculating densities were more than 1 000 × 10~4 cells/mL,which showed no significant difference(P > 0. 05). High specific growth rates were observed within 48 h after culture,which showed significant difference(P < 0. 05),while the doubling time was also short and significantly different(P < 0. 05). MDCK-XF03 cells made full use of glucose within 84 h after culture. Conclusion A MDCK suspension cell line MDCK-XF03 was successfully obtained and cultured in bioreactor at an initial inoculating density of 200 × 10~4 cells/mL,which achieved the goal of scale-up culture from 5 L to 75 L bioreactors. It provided a cell matrix for the industrial production of influenza vaccine.

    2021 11 v.34 [Abstract][OnlineView][Download 725K]

  • Development and verification of a method for determination of biological activity of recombinant human interleukin-1 receptor antagonist

    YU Lu;WANG Ying;ZHANG Yu;LIU Ying;LIU Jing-hui;LIU Yu-lin;Department of Cytokines,Changchun Institute of Biological Products Co.,Ltd.;

    Objective To develop and verify a method for determination of biological activity of recombinant human interleukin-1 receptor antagonist(IL-1 Ra). Methods The biological activity of IL-1 Ra was evaluated according to its ability in blocking the killing effect of interleukin-1β （IL-1β） on human melanoma A375.S2 cells,and the result was corrected with the standard. The initial sample concentration(8,10,12 and 14 μg/mL)and final IL-1β concentration(2,4 and 6 ng/mL)were optimized,and the method was verified for specificity,accuracy,linear range,intermediate precision and durability. Three batches of bulks and three batches of final products of IL-1 Ra prepared by Changchun Institute of Biological Products Co., Ltd. were determined by the developed method. Results The initial sample concentration and final IL-1β concentration of the developed method were optimized as 8 μg/mL and 4 ng/mL respectively. The components of buffer in bulk and subsidiary materials in final product showed no impact on the determination result. The linear range of the developed method was 6 ～ 13 μg/mL,with a correlation coefficient(R2)of 0. 992 1. The slope of linear regression equation in verification for accuracy was 0. 975 8. The geometric coefficients of variation(GCVs)of a batch of bulk and a batch of final product by different persons at different time points were 13. 895% and 8. 670%respectively,both of which were less than 20%. However,the GCV of determination result of relative potencies with cells of different passages and densities wsa 5. 595%,which was less than 20%. The determination results of three batches of bulks and three batches of final products by the developed method were relatively stable. Conclusion A method for determination of biological activity of recombinant human IL-1 Ra was successfully developed, which showed high accuracy,precision and durability,and might be used for the biological activity evaluation and quality control of the product.

    2021 11 v.34 [Abstract][OnlineView][Download 224K]

  • Application and research progress of rabies virus CTN-1 strain in rabies vaccine for human use

    SHI Lei-tai;LI Yu-hua;National Institutes for Food and Drug Control;

    The rabies virus CTN-1 strain was isolated,domesticated,attenuated,established and applied in China. It has an independent intellectual property right and has been fully adapted to Vero cells,which has the advantages such as high potency,good immunogenicity,low concentration multiple and low production cost. CTN-1 strain has a good protective effect on the population. Comprehensive verification for biology,immunology,immunochemistry,molecular biology,gene sequencing and animal epidemiology prove that CTN-1 strain meets the national and WHO requirements and quality standards for production of human rabies vaccine. This paper reviews the application and research progress of rabies virus CTN-1 strain in rabies vaccine for human use.

    2021 11 v.34 [Abstract][OnlineView][Download 227K]

  • Progress in research on role of small molecular compounds in field of induced pluripotent stem cells

    LIU Xin-yang;QIN Jie-chen;ZHAO Qing;School of Pharmaceutical Science & Yunnan Key Laboratory of Pharmacology for Natural Products,Kunming Medical University;

    The use of transcription factors can reprogram differentiated cells into induced pluripotent stem cells(i PSCs).However,transcription factors are prone to gene mutations which will bring safety problems. The ultimate goal of regenerative medicine is to directly obtain somatic cells from patients and induce them to differentiate into pluripotent stem cells through small molecule compounds. The small molecule compound induced pluripotent stem cells is an important tool to effectively produce high-quality i PSCs. There are many types of small molecule compounds with stable structures and relatively controllable effects. This article reviews the classification and effects of small molecule com-pounds that have been reported so far,as well as the methods to induce i PSCs from all compounds.

    2021 11 v.34 [Abstract][OnlineView][Download 169K]

  • Effect of CD44 expression on breast cancer stem cells

    ZHANG Rui-juan;YANG Ying;College of Life Science,Tianjin University;

    The stemness of breast cells is mainly maintained by CD44 standard isoform(CD44 s),while the cell proliferation by CD44 variant isoform(CD44 v). As an important signal molecule,CD44 receives signal transmission of cytokines which come from microenvironment,integrates the signal and releases CD44 ICD through the action of lyase,and transmits the signal to the nucleus. CD44 v subtype cells also play a leading role in the metastasis process of breast cancer stem cells(BCSCs)which express CD44. During epithelial-mesenchymal transition(EMT),the transformation and structural changes of CD44 subtype have a significant effect on cell invasion,migration and distal positioning. This paper reviews the maintenance of stemness of breast cancer cells by CD44 and the preference of different subtypes of breast cancer cells during metastasis to lung,bone,liver and brain during metastasis.

    2021 11 v.34 [Abstract][OnlineView][Download 196K]

  • Progress in research on correlation of human leukocyte antigen typing to leukemia

    PEI Yong-feng;YU Mei;Institute of Transfusion Medicine,Nanning Blood Center;

    Human leukocyte antigen(HLA) is the major histocompatibility complex(MHC) in humans,of which the typing methods are mainly based on low-resolution polymerase chain reaction-sequence specific primers(PCR-SSP) and high-resolution polymerase chain reaction-direct sequencing typing(PCR-SBT). At present,the pathogenesis of leukemia is not completely known. It is found that HLA is an important genetic factor for leukemia. Many HLA genes are genetically susceptible or antagonistic to the occurrence of leukemia,and some HLA molecules are closely related to the diagnosis,treatment and prognosis of leukemia. This article reviews the progress in research on the correlation of HLA typing to leukemia in recent years,which will provide a reference for the research on the pathogenesis,diagnosis and prevention of leukemia.

    2021 11 v.34 [Abstract][OnlineView][Download 210K]


@2012 Editorial Department of "Chinese Journal of Biologicals"