Current Issue

  • Whole genome and structural protein sequence analysis of PM1503-02 strain applied to human diploid cell rabies vaccine research

    ZUO Jing;ZHANG Weiye;TONG Wei;CUI Li;DONG Haohuan;DONG Jinqiu;R&D Center, Beijing Minhai Biotechnology Co., Ltd.;

    Objective To analyze the application of PM1503-02 strain in human diploid cell rabies vaccine(HDCV)production research at the molecular level.Methods The whole genome sequences and amino acid sequences of PM1503-02 were studied in comparison with 18 domestic and foreign vaccine strains using homology comparison and phylogenetic trees, and the antigen-binding sites and glycosylation sites of the glycoproteins(Gs) of PM1503-02 and 18 domestic and foreign vaccine strains were also studied in comparison. In addition, a phylogenetic tree was also constructed with 56 domestic street strains simultaneously to analyze the affinities between PM1503-02 and domestic street strains. The genetic stability of the whole genome sequence of PM1503-02 was confirmed across different passage generations.Results The whole genome sequence of PM1503-02 strain showed the highest homology with PM1503 and CVS-11 strains, reaching more than 99%, and showed the next highest homology with HEP-Flury, Flury LEP and Flury-LEP-C strains, reaching more than95%. There were few mutations in the antigenic sites in the extra-membrane domain of the G protein of PM1503-02, and only the 40 th amino acid of the antigenic siteⅡwas mutated to glutamic acid(E). The glycosylation sites were at amino acid positions 37, 247 and 319, respectively. The results of phylogenetic tree construction with 56 domestic strains showed that PM1503-02 was closely related to domestic street strains. PM1503-02 strain was passaged to the 63 rd, 64 th, 65 th, 66 th and70 th generations in HDCs, and single-point mutations appeared in the 66 th and 70 th generations of the whole genome sequences, but all of them were synonymous mutations with no effect on the protein structure.Conclusion This study provides favorable support for developing a more effective HDCV using PM1503-02 at the molecular level, which has a better prospect for application in China.

    2025 11 v.38 [Abstract][OnlineView][Download 1214K]

  • Analysis on serum metabolism changes in Chinese rhesus macaques after immunization with plague vaccine strain EV76

    ZHANG Qi;ZHANG Aiping;LI Qiang ;ZHANG Xiaolu;WANG Yongshun;WU Haisheng;ZHANG Qingwen;QI Zhizhen;ZHAO Zhijun;NHC Key Laboratory of Plague Prevention and Control, Qinghai Institute for Endemic Disease Prevention and Control;

    Objective To investigate the changes in serum metabolism of Chinese rhesus macaques after immunization with plague vaccine strain EV76, and to provide a scientific basis for evaluating the immunization effect of plague vaccine strain.Methods Six rhesus macaques were divided into EV76 immunization group and control group, with three in each group(two females and one male). The immunization group was injected with 1. 57 × 10~(10)CFU EV76 subcutaneously in the groin, and the control group was only injected with the same amount of PBS subcutaneously. Four weeks later, the blood was collected from femoral vein, and serum non-targeted metabolomics was detected by ultra-performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS). Differential metabolites were screened according to the criteria of variable importance in projection(VIP) > 1. 0, fold change(FC) > 1. 5 or < 0. 667 and P < 0. 05, and KEGG enrichment analysis of differential metabolites was performed.Results A total of 460 metabolites in positive ion mode and 440 in negative ion mode were identified. A total of 24 differential metabolites were found after database comparison. According to KEGG pathway enrichment analysis, the differential metabolites were mainly involved in the metabolism of glycine, serine and threonine, the biosynthesis of phenylalanine, tyrosine and tryptophan, the biosynthesis of valine, leucine and isoleucine, the metabolism of phenylalanine, the metabolism of cysteine and methionine, and the metabolism of tyrosine.Conclusion EV76 exerts its immune-enhancing effect by increasing the content of acetyl-L-carnitine, D-glutamine, hypoxanthine nucleoside and cortisol,and there are abnormal changes in a variety of amino acid metabolic pathways.

    2025 11 v.38 [Abstract][OnlineView][Download 898K]

  • Biological characteristics and genetic stability of 2BS celladapted strain of F-genotype mumps virus

    LIU Na;LIU Hongtao;SUN Yueqiu;ZANG Wanru;MENG Fandong;LI Shuang;SONG Wenjiao;LI Jiangdan;SHUANG Hui;CAI Yan;Research Department of Changchun Keygen Biological Products Co., Ltd.;

    Objective To investigate the biological characteristics and genetic stability of the 2 BS cell-adapted strain of F-genotype mumps virus(MuV), in order to lay the foundation for the subsequent development of live attenuated mumps vaccines.Methods The QBB-2 BS-3.2 and QBB-2 BS-9.3 strains were adapted through 40 passages in 2 BS cells, and the virus titers were measured at passages P15 to P40. Western blot was employed to evaluate the expression levels of key proteins hemagglutinin-neuraminidase(HN) and nucleocapsid protein(NP) in the MuV-QBB strains. After whole-genome sequencing, DNAstar 11.0 software was used to analyze the nucleotide and amino acid homology among different passages of viruses, and Mega11.0 and Generunner v 5.1.06 software was used for sequence alignment of the nucleotide and amino acid sequences, as well as for comparison of the variation of key antigenic sites.Results The titers of strains QBB-2 BS-3.2 and QBB-2 BS-9.3 graduallyincreasedfromP15 toP40,stabilizingatabout7.40 lgCCID_(50)/mLafterP20.TheexpressionlevelsofHN and NP proteins remained consistent across P15 to P40 for both QBB-2 BS-3.2 and QBB-2 BS-9.3 strains. The QBB-2 BS-3.2 strain maintained 100. 00% nucleotide and amino acid sequence homology across P15, P20, P25, P30, P35, and P40, while the QBB-2 BS-9.3 strain exhibited 99. 90% to 100. 00% homology across different passages. The QBB-2 BS-3.2 strain had three nonsynonymous mutations during P15 to P40, whereas the QBB-2 BS-9.3 strain had four. Compared to the S79 vaccine strain, the QBB strains had one additional N-glycosylation site in HN protein and four mutations in the immunogenic regions,with no mutations in the neutralizing epitopes.Conclusion The QBB-2 BS-3.2 and QBB-2 BS-9.3 strains maintain high virus titers and exhibit good genetic stability after continuous passaging in 2 BS cells, indicating the potential to be used as vaccine candidate strains, providing a solid foundation for the development of attenuated live mumps vaccines.

    2025 11 v.38 [Abstract][OnlineView][Download 884K]

  • Stability study on subculturing of working seed strain for recombinant human interferon α2b

    ZHANG Kaining;YU Jian;GU Xiaoyu;ZHU He;ZHANG Ying;ZHANG Ting;LI Tianwen;MO Fei;LIU Yunnan;XU Xiaoming;Changchun Institute of Biological Products Co., Ltd.;

    Objective To verify the passage stability of recombinant human interferon(rhIFN) α2b working seed strain at key generations through the construction of a 15 L scale-down model, in order to ensure the stability and effectiveness of rhIFNα2b during the production process.Methods The rhIFNα2b lyophilized strain was revived and subcultured, and glycerol stocks of the 0, 10 th, 20 th, 30 th, 40 th and 50 th generations of the culture were preserved respectively. The glycerol stocks of each generation preserved in the subculture stability study were selected and the fermentation-expression experiments were conducted in a 15 L reduced-scale model fermentation system under simulated production conditions. By real-time monitoring of the growth curve of the cells and combining with endpoint detection of product expression level,biological activity, plasmid stability, genetic integrity and microbial purity, a comprehensive comparison was made to determine whether the limited subculture times could cover the existing culture process.Results The rhIFNα2b working seed strains from passages 0, 10, 20, 30, 40, and 50 exhibited good stability and consistency after continuous subculturing.All strains showed typical E. coli colony morphology on LB agar plates with no contamination by foreign microorganisms.Antibiotic resistance profiles remained unchanged, Gram staining confirmed Gram-negative bacterial characteristics, and biochemical profiles were consistent with typical E.coli traits. The target gene expression levels consistently exceeded 20%without a declining trend, and the fermentation activity remained above 1. 0 × 108 IU/L. Plasmid restriction analysis produced bands consistent with theoretical expectations, with no observed sequence mutations, and the plasmid loss rates remained below 30%, indicating robust genetic stability.Conclusion The rhIFNα2b working seed strains demonstrated good fermen-tation stability under scaling-down simulated production conditions in the 15L bioreactor. The current passage limitations of the strain effectively meet the requirements of the updated production process, providing crucial data to support process improvement and optimization.

    2025 11 v.38 [Abstract][OnlineView][Download 1032K]

  • Expression of mouse hepatitis virus M protein in baculovirus expression system and evaluation of its antigenicity

    LUAN Mingzhu;MA Yueyu;SUN Li;LI Ming;FEI Dongliang;MA Mingxiao;QI Xinyao;Experimental Animal Center,Jinzhou Medical University,Liaoning Provincial Key Laboratory of Animal Product Quality and Safety;

    Objective To express mouse hepatitis virus(MHV) M protein by baculovirus expression system and evaluate its antigenicity, so as to lay a foundation for the development of MHV diagnostic kit.Methods The highly conserved MHV M protein gene sequence(GenBank: FJ647223) was screened by sequence alignment, and cloned into vector pFastBac I after codon optimization, then transformed into competent E.coli DH10 Bac~(TM). The recombinant bacmid Bacmid-M was obtained by blue-white spot screening, subsequently transfected into Sf9 insect cells, and the recombinant baculovirus rBV-M was obtainedbyrescue.Themorphologyofvirusparticleswasobservedundertransmissionelectronmicroscope,andtheexpression of recombinant M protein in Sf9 cells was identified by indirect immunofluorescence and Western blot. Using standard positive sera against reovirus type-3(Reo-3), minute virus of mice(MVM), Sendai virus(SV) and MHV as primary antibodies, the antigenic specificity of the recombinant M protein was analyzed by Western blot.Results The recombinant Bacmid-M was constructed correctly as identified by PCR, and the recombinant baculovirus rBV-M with typical rod-shaped morphology could be packaged in Sf9 cells. Sf9 cells infected with rBV-M showed red fluorescence under fluorescence microscope, and the expressed recombinant M protein exhibited specific binding to His-Tag monoclonal antibody at the relative molecular mass of26000.Therecombinantproteincouldonlyreactspecificallywithanti-MHVstandardpositiveserum,buthadnocross-reaction with anti-Reo-3, anti-MVM and anti-SV standard sera.Conclusion The M protein of MHV was expressed by baculovirus system, which has good specificity and antigenicity, and is expected to be used as coating antigen for the development of MHV diagnostic reagents.

    2025 11 v.38 [Abstract][OnlineView][Download 959K]

  • Construction and biological activity analysis of antimicrobial peptide RLT

    CHA Sina;FANG Yuxin;DONG Na;College of Animal Science and Technology,Northeast Agricultural University;

    Objective To construct antimicrobial peptide RLT by molecular modification of porcine beta-defensin 135(PBD135) and analyze its biological activity, so as to provide reference for the development of feed antibiotic substitutes.Methods Two molecules of histidine(His) in the amino acid sequence of PBD135 were replaced by arginine(Arg) with positive charge by amino acid mutation to obtain antimicrobial peptide RLT. Circular dichroism was used to analyze the secondary structure of antimicrobial peptide RLT, the minimal inhibitory concentration(MIC) method was used to evaluate its antibacterial activity against a variety of Gram-positive and Gram-negative bacteria, and the MTT method was used to detect its cytotoxicity against macrophage RAW264.7 of mice. Furthermore, the stability of antimicrobial peptide RLT in different salt ions(Na~+, Ca~(2+), NH_4~+, Zn~(2+), K~+, Mg~(2+), Fe~(3+)), pH(2 and 12) and serum concentration(50% and 25% fetal bovine serum)was investigated.Results The antimicrobial peptide RLT could form a regular α-helical structure in a simulated membrane environment, and its antibacterial activity against a variety of Gram-positive and Gram-negative bacteria was higher than that of PBD135. Meanwhile, the antimicrobial peptide RLT exhibited low toxicity to eukaryotic cells. RLT maintained high activity in acidic and alkaline environments, as well as environments with 25% and 50% serum, but its antibacterial activity was completely inhibited in Na+, Ca2+ and NH4+ environments.Conclusion A new antimicrobial peptide RLT was designed and synthesized, which has high broad-spectrum antibacterial activity, low cytotoxicity and good stability, and is expected to become an alternative to antibiotics for feeding purposes.

    2025 11 v.38 [Abstract][OnlineView][Download 845K]

  • Comparative analysis of centralized detection of leukoreduced suspended red blood cells in the Chongqing area from 2021 to 2023

    WU Siqi;DAI Huayou;CHEN Juan ;HUANG Runhua ;ZHANG Zhen ;MOU Juanjuan ;WANG Yunxia ;HUANG Xia;Chongqing Blood Center;

    Objective To retrospectively analyze the centralized detection data of 1 unit of leukoreduced suspended red blood cells from blood collection and supply institutions in the Chongqing area from 2021 to 2023, investigate the key processes of blood collection and preparation, and evaluate the quality of regional blood products, so as to provide a basis for establishing unified regional blood quality homogenization.Methods Centralized detection was conducted on leukoreduced suspended red blood cells from six blood collection and supply institutions in Chongqing from 2021 to 2023. The detection data trends were statistically analyzed, and the Kruskal-Wallis H test was used to analyze the quality control items of the six institutions over the three years. The centralized detection results were fed back on-site, and paper survey forms were distributed to understand the quality control situation of each institution.Results The compliance rate of each quality control item of leukoreduced suspended red blood cells from the six institutions was > 85%. The Kruskal-Wallis H test revealed statistically significant differences among the institutions in each item, with H of 44. 86 to 246. 96 and P < 0. 001. Over the three years,the volume and haemoglobin(Hb) results of institutions B and C were more dispersed, while those of institution F were more concentrated. The red blood cell hematocrit(HCT) data of institution B were more dispersed, while those of institutions A, D,and F were more concentrated. The residual leukocyte count data of institution C were more dispersed, while those of institutions A and F were more concentrated. The hemolysis rate at the end of storage fluctuated significantly for institution A.Conclusion Since the implementation of centralized detection, improvements have been made in all links of the blood chain for each institution. However, data analysis still reveals that blood product homogenization needs to be further strengthened.

    2025 11 v.38 [Abstract][OnlineView][Download 1012K]

  • Efficacy and safety evaluation of recombinant human interferon alpha2b suppository in treatment of cervical high-risk human papilloma virus infection

    WEI Meng;GUO Xinxin;HAN Xiao;WANG Ning;YUAN Chunli;The First Hospital of Jilin University;

    Objective To evaluate the efficacy and safety of recombinant human interferon alpha2b(rHuIFNα2b) suppository in the treatment of cervical high-risk human papilloma virus(HPV) infection, so as to provide experimental basis for its clinical application.Methods From September 2021 to July 2023, 118 patients with high-risk HPV infection who met the inclusion and exclusion criteria of this study were collected from the gynecological outpatient department of the First Hospital of Jilin University. The patients were randomly assigned to the parallel experimental group and placebo-control group with blind method. The experimental group was given rHuIFNα2b suppository, and the control group was given placebo with uniform appearance. All patients started taking the medicine about 3 days after the end of menstruation. The usage method was to place the rHuIFNα2b suppository in the posterior fornix of vagina before going to bed, 1 suppository each time, once every other day, and stop using it during menstruation. Nine times were a course of treatment, and three courses of treatment were used continuously. The efficacy was assessed by comparing the post-treatment HPV clearance rate and total effective rate, and adverse events throughout the entire study period were documented.Results After the treatment, the HPV negative conversion rate(76. 67%) and total effective rate(85. 00%) of the experimental group were significantly higher than those of the control group(negative conversion rate: 43. 10%, total effective rate: 51. 72%), and the difference was statistically significant(χ~2= 13. 86 and 15. 17, respectively, each P < 0. 001). In the experimental group, there were no significant differences in the total effective rates of rHuIFNα2b suppository against HPV16, HPV18, HPV52 and HPV58(P = 0. 140-1. 000). Among the118 patients, a total of 2 cases had adverse events, including 1 case in the experimental group and 1 case in the control group, with the both symptoms of transient vulvovaginal discomfort.Conclusion Topical administration of rHuIFNα2b suppository is effective in the treatment of cervical high-risk HPV infection, and it is effective against all major high-risk subtypes of HPV with good safety, which is worthy of clinical application.

    2025 11 v.38 [Abstract][OnlineView][Download 840K]

  • Development of national standards for insect cell Sf9 and HighFiveTM DNA residue detection

    QIN Haiyang;LI Jiayi;XIU Pengcheng;NIE Jianhui;HUANG Weijin;ZHANG Li;Division of HIV/AIDS and Sex-transmitted Virus Vaccines, Institute for Biological Products Control,National Institutes for Food and Drug Control;

    Objective To establish national standards of genomic DNA for insect cell Sf9 and HighFive~(TM)(Hi5), in order to standardize the quantifying of residual DNA in related biological products.Methods Genomic-tip 500/G kit was used to extract genomic DNA of Sf9 and Hi5 cells, and the concentration and purity were analyzed by UV spectrophotometry and agarose gel electrophoresis. After stability and homogeneity analysis, these standard substances were calibrated by four laboratories for quantifying and assignment. Then the standard substances were evaluated in real-time PCR and fluorescence staining for applicability.Results Genomic DNA of Sf9 and Hi5 cells with high-purity were obtained, which were qualified with A_(260)/A_(280)between 1. 7 and 2. 0, and showed a single specific band in agarose gel electrophoresis. The homogeneity and stability all met the requirements of national standards. After collaborative study in four independent laboratories, the geometric mean of the concentration of insect cell Sf9 DNA residue standard was assigned as 97. 95μg/mL, the 95%confidence interval of the average concentration was 97. 27-98. 40μg/mL, and the 95% confidence interval in single test was 92. 02-103. 76μg/mL; The geometric mean of the concentration of insect cell Hi5 DNA residue standard was assigned as 97. 05μg/mL, the 95% confidence interval of the average concentration was 96. 61-97. 72μg/mL, and the 95%confidence interval in single test was 91. 46-102. 90μg/mL.Conclusion Insect cell Sf9 and Hi5 DNA residue national standards were successfully established, which met the requirements of real-time PCR and fluorescence staining and can be used to detect the residual DNA in related biological products.

    2025 11 v.38 [Abstract][OnlineView][Download 1038K]

  • Development, verification and application of CCK-8 assay for antiviral drug susceptibility determination of herpes simplex virus-1

    TIAN Na;DENG Li;MA Tiancong;LI Jinghua;GAO Wenrui;YU Shouzhi;PAN Shuyuan;Research and Development Office, Beijing Institute of Biological Products Co., Ltd.;

    Objective To develop a CCK-8(cell counting kit-8) assay for the determination of antiviral drug susceptibility of herpes simplex virus-1(HSV-1) in Vero cells, and to use the method for susceptibility determination of wild and modified strains after verification.Methods After optimizing and screening CCK-8 reagent culture time(3, 4, 5 h), acyclovir(ACV)dilution ratio(2, 4 times) and sample culture time(72, 96, 120 h), the CCK-8 assay for the determination of antiviral drug susceptibility of HSV-1 was developed. The method was verified for specificity, accuracy, repeatability and system applicability, and then was used to detect the susceptibility of wild-type HSV-1 and its modified strains oHSV-GFP and oHSVGM-CSF to ACV, which was compared with plaque method.Results The developed CCK-8 method was as follows: HSV-1 was mixed with different concentrations(5, 1. 25, 0. 313, 0. 078 1, 0. 019 5, 0. 004 88 μg/mL) of ACV and then inoculated into Vero cells. After culturing for 72 h, CCK-8 reagent was added. After another 4 h of continuing culture, the absorbance value at 450 nm wavelength was detected by a microplate reader, and the IC_(50)of ACV on HSV-1 was calculated. The IC_(50)value of ACV against oHSV-1 standard strain was 0. 13 μg/mL. The method exhibited good specificity and system suitability.The sample recovery rates of accuracy and repeatability verification were both within the range of 70% to 130%, with the RSD < 20% for repeatability verification. The IC_(50)values of ACV against wild-type HSV-1 and its modified strains oHSVGFP and oHSV-GM-CSF were 0. 10, 0. 09 and 0. 11 μg/mL, respectively. The IC_(50)value of ACV against wild-type HSV-1 strain measured by plaque assay was 0. 15 μg/mL, which was close to that detected by CCK-8.Conclusion The developed CCK-8 assay for the determination of antiviral drug susceptibility of HSV-1 is a simple, practical and effective method, which can be used to study the susceptibility of various wild-type HSV-1 and its modified strains to various antiviral drugs.

    2025 11 v.38 [Abstract][OnlineView][Download 905K]

  • Establishment and validation of a high-performance liquid chromatography with charged aerosol detection method for determination of sucrose content in mRNA-based SARS-CoV-2 vaccines

    WU Fan;LIU Qin;YANG Huan;SHEN Xue;WANG Sijie;HE Wenzhi;FEI Chengrui;Yunnan Institute of Supervision and Inspection for Food and Drug, Public Service Platform for Industrial Technology Foundation of the Ministry of Industry and Information Technology;

    Objective To establish a high-performance liquid chromatography(HPLC) with charged aerosol detection(CAD)method for the determination of sucrose content in mRNA-based SARS-CoV-2 vaccines, and to validate and preliminarily apply the method, so as to provide a new detection method for the determination of sucrose content in more mRNA vaccines.Methods Chromatographic column: Zorbax NH_2(Agilent, 4. 6 mm × 250 mm, 5. 0 μm); Chromatographic conditions: acetonitrile dissolved in water(50% acetonitrile) as the mobile phase, the flow rate of 0. 8 mL/min, the nebulization temperature of 70 ℃, the detector acquisition frequency of 10 Hz, the filtration of 3. 6 s, and the column temperature of 20 ℃. The linearity, limit of quantification(LOQ) and limit of detection(LOD), specificity, repeatability and accuracy of the method were validated, and the sucrose content in three batches of mRNA-based SARS-CoV-2 vaccines was detected by the established method.Results The concentration of the sucrose standard exhibited a good linear correlation with peak area in R therangeof0.25 to6 mg/mL,withacorrelationcoefficient()of0.9986.TheLODandLOQwere0.03125 and0.125 mg/mL respectively. Lactose and sucrose could be completely separated under the experimental conditions, and the resolution was1. 65. The relative standard deviation(RSD) of peak area of the six test samples was 2. 4%. The recovery rates for spiked samples at three different concentrations ranged from 101. 3% to 102. 2%. The sucrose content of three batches of SARS-CoV-2 mRNA vaccine samples detected by the established method was 81. 377 4, 80. 339 0 and 74. 157 3 mg/mL, respectively.Conclusion The established HPLC-CAD method has a wide measurement range, good linearity and repeatability, and high accuracy, allowing for accurate quantification of sucrose content in the mRNA-based SARS-CoV-2 vaccines.

    2025 11 v.38 [Abstract][OnlineView][Download 916K]

  • Validation of analytical method for molecular size variants in monoclonal antibodies based on size-exclusion high-performance liquid chromatography

    YANG Yalan;LIU Bing ;CUI Yongfei;LI Meng;WU Gang;YU Chuanfei;Division of Monoclonal Antibodies,National Institutes for Food and Drug Control,NHC Key Laboratory of Research on Quality and Standardization of Biotech Products,NMPA Key Laboratory for Quality Research and Evaluation of Biological Products;

    Objective To verify the size-exclusion high-performance liquid chromatography(SEC-HPLC) method for molecular size variant analysis in monoclonal antibodies which is to be included in the Chinese Pharmacopoeia(VolumeⅢ, 2025 edition), in accordance with the requirements of the ICH Q2(R2) Guideline for Analytical Method Validation.Methods Firstly, the validation plan, acceptable standards, and samples for analytical and instrument linearity were established. Then,the method was validated for specificity, range, accuracy and precision. Finally, the statistical analysis of test results was conducted to obtain the validation report of the SEC-HPLC method.Results The SEC-HPLC method exhibited a good specificity, with a reportable range of 0. 21% to 8. 33% for aggregates, 0. 06% to 2. 58% for fragments, and ≥ 89. 09% for monomers. The working range of the method was 5-300 μg, with limit of quantitation(LOQ) of 0. 21% for aggregates and0. 06% for fragments. The linear correlation coefficient(R2) exceeded 0. 99. The mean recovery rates ranged between 80%and 110%. The RSDs for both repeatability and intermediate precision analyses were less than 8%.Conclusion The specificity, range, accuracy, and precision of the analytical method for molecular size variants in monoclonal antibodies based on SEC-HPLC all meet the acceptance criteria.

    2025 11 v.38 [Abstract][OnlineView][Download 959K]

  • Establishment and verification of a method for in vivo efficacy detection of inactivated SARS-CoV-2 vaccine(Vero cells)

    SHEN Ailin;TANG Wei;YAN Jiachi;HE Hongwu;LUO Min;YU Juan;CHENG Yao;HU Fenfen;ZHU Lin;TIAN Yao;CHEN Cai;PAN Ting;SHI Jinrong;Wuhan Institute of Biological Products Co., Ltd., National Engineering Technology Research Center for Combined Vaccines;

    Objective To establish a method for in vivo efficacy test of inactivated SARS-CoV-2 vaccine(Vero cells) by immunizing BALB/c mice combined with ELISA, explore its influencing factors and verify the method.Methods At 14 d after the mice were immunized with inactivated SARS-CoV-2 vaccine(Vero cells), the blood samples were collected from the eyeballs and the inactivated serum was isolated, and the titer of specific SARS-CoV-2 antibody in mouse serum was determined by ELISA. The results of two animal sources and three age groups of mice, as well as three batches of key reagents(coating plate, enzyme dilution and enzyme-labeled antibody) in the detection kit, were compared and analyzed, and the precision, specificity and relative accuracy of the established method were verified according to the guidelines for the validation of bioactivity/potency assays for biological products in the Chinese Pharmacopoeia 9401(Volume IV, 2020 edition). The in vivo efficacy of 10 batches of semi-finished products of inactivated SARS-CoV-2 vaccine(Vero cells) was detected by the established method.Results The geometric coefficients of variation(GCVs) of the results of mice from different sources immunized with three batches of samples ranged from 17. 1% to 30. 3%, and there was no statistically significant difference between the results of two sources of mice in the 95% confidence interval(t = 0. 25, P > 0. 05). The GCV of the results of mice at different weeks of age immunized with the same batch of samples was 10. 9%. The GCV of A_(450)_-_(630)value detected by the same batch of kits, the same batch of antibodies and different batches of enzyme dilutions was 4. 8%, the GCV of A_(450)_-_(630)value detected by the same batch of kits, the same batch of enzyme dilutions and different batches of antibodies was9. 9%, and the GCV of A_(450-630)value detected by the same batch of antibodies, the same batch of enzyme dilutions and different batches of coating plates was 6. 3%. The GCVs of six tests of the same batch of semi-finished products was 9. 1%-23. 8% by the same experi-menter and 24. 9% by different experimenters at different time and places. The GCV of geometric mean titer(GMT) of antibody titer in aluminum hydroxide adjuvant group and negative control group was 8. 4%. The relative bias(RB) of the same sample at four different dilutions ranged from-25. 0% to 21. 7%. The titers of 10 batches of vaccine reference products ranged from 1 131 to 2 263, and the GCV was 13. 0%.Conclusion The established mouse immunization combined with ELISA method has high accuracy, precision and good specificity, which can be used to detect the in vivo efficacy of inactiva-ted SARS-CoV-2 vaccine(Vero cells), and can be used as an evaluation index of product efficacy.

    2025 11 v.38 [Abstract][OnlineView][Download 877K]

  • Development and verification of a 1H nuclear magnetic resonance detection method for O-acetyl and pyruvate content in pneumococcal polysaccharide conjugate vaccine

    LV Xinyang;GUO Deqian;QIU Peng;LEI Yonghong;LIN Jisheng;LIAN Xiaojuan;WANG Jiandong;HU Yaling;Sinovac Life Sciences Co.,Ltd.;

    Objective To develop and verify a~1H nuclear magnetic resonance(~1H NMR) method for determining the relative content of O-acetyl and pyruvate in all intermediates of pneumococcal polysaccharide conjugate vaccine, providing a reliable quality control tool for the development and manufacture of pneumococcal polysaccharide conjugate vaccines.Methods A quantitative 1H NMR protocol was developed in which the methyl resonance of the capsular polysaccharide served as the characteristic peak for calculating the relative content of O-acetyl and pyruvate in both the bulk polysaccharide and downstream intermediates. The method was validated for linearity and range, specificity, and accuracy. The validated assay was then applied to determine the relative levels of O-acetyl and pyruvate in the degraded polysaccharides, activated polysaccharides and conjugate bulks of Streptococcus pneumoniae serotypes 1, 4, 7F, 17F, 18C and 22F prepared by either reductive amination or cyanation chemistry.Results Within the range of 2. 5-20. 0 mg/mL of the test sample, the polysaccharide peak-area ratio remained constant, establishing the valid quantitation range. Neither internal standard nor heavy water interfered, quantitation peaks of all serotypes were baseline-resolved, and post-degradation chemical-shift changes did not impair identification. The content of O-acetyl and pyruvate detected by 1H NMR agreed with the spectrophotometric data. The RSDs for O-acetyl content in degraded, activated and conjugate bulks of serotypes 1, 7F, 17F, 18C and22F were ≤ 6. 03 %, and for pyruvate content in serotype 4 intermediates ≤ 4. 51 %.Conclusion The 1H NMR method demonstrated excellent repeatability and broad applicability for quantifying O-acetyl and pyruvate content, making it suitable for quantitative determination of these modifications in all intermediates of pneumococcal polysaccharide conjugate vaccines.

    2025 11 v.38 [Abstract][OnlineView][Download 933K]

  • Screening and validation of novel electrophoretic buffers for immunoelectrophoresis

    ZHAO Yuhao;ZHAO Qin;ZHOU Qin;CAI Min;ZHANG Youjie;DUAN Xuhua;SHAO Hong;NMPA Key Laboratory for Quality Control of Therapeutic Monoclonal Antibodies, Shanghai Institute for Food and Drug Control;

    Objective To screen and verify a buffer to replace barbiturate solution(BBTS) in immunoelectrophoresis, in order to obtain a safer, environmentally friendly buffer whose electrophoresis effects are equivalent to BBTS.Methods Five types of buffers to be examined, trihydroxy aminomethane-acetic acid-disodium ethylenediamine tetraacetate(TAE), glycine-sodium hydroxide(Gly-NaOH), trihydroxy aminomethane-hydrochloric acid(Tris-HCl), borate buffer solution(BBS) and phosphate buffer solution(PBS), were prepared. The buffers were preliminarily screened and confirmed via immunoelectrophoresis using human albumin and human immunoglobulin as test samples, and the precipitation arc clarity and migration distance as indicators. In addition, the specificity, durability, stability and universality of the method were verified.Results Among the five buffers, TAE and PBS exhibited the best electrophoresis effects, with the moderate migration distance of human albumin and immunoglobulin, as well as the forming of clear precipitation arcs, which were equivalent to those of BBTS. As buffers,TAE and PBS showed good specificity, durability, stability and universality.Conclusion The reagents required to prepare PBS are more readily available, less costly, and safer to use, and PBS buffer was finally selected as an alternative buffer to BBTS in immunoelectrophoresis.

    2025 11 v.38 [Abstract][OnlineView][Download 1015K]

  • Research status of lactoferrin and its anti-human coronavirus effect

    ZHAO Yu;SHU Xingfu;ZHANG Haixia;MA Zhongren;Key Laboratory of Biotechnology and Bioengineering of State Ethnic Affairs Commission, Biomedical Research Center, Northwest Minzu University;

    Coronaviruses(CoVs) are plus-stranded RNA viruses that can infect a variety of hosts. Currently, several CoVs have been found to cause human disease, such as human coronavirus(HCoV), severe acute respiratory syndrome coronavirus(SARS-CoV), Middle East respiratory syndrome coronavirus(MERS-CoV), and SARS-CoV-2. Among them, SARS-CoV-2 mainly causes acute infectious coronavirus disease 2019(COVID-19), which seriously threatens human health and has become one of the major public health problems in the world. At present, some SARS-CoV-2 vaccines or drugs have been marketed, but there are shortcomings such as requiring multiple injections and being not applicable to some people. There is an urgent need to develop new drugs with good safety and low side effects. Lactoferrin(LF), a natural iron-binding glycoprotein, has broad-spectrum antiviral, anti-inflammatory and immunomodulatory functions. In this paper, the main biological functions of LF and the research progress on its anti-HCoV effects are reviewed, in order to provide reference for the application of LF in the clinical prevention and treatment of HCoV infection.

    2025 11 v.38 [Abstract][OnlineView][Download 867K]

  • Research status and progress on respiratory syncytial virus vaccines

    LI Chenglong;ZHAO Di;PAN Ruowen;AN Wenjue;Hualan Biological Bacterin Inc.;

    Respiratory syncytial virus(RSV), an RNA virus, is one of the leading pathogens causing respiratory tract infections in infants, the elderly and immunocompromised people, resulting in a huge medical burden of disease worldwide every year. The fusion protein(F protein) on the surface of RSV is the main target of neutralizing antibodies, especially the discovery of its pre-fusion conformation(pre-F), which has laid a key theoretical foundation for the design of a new generation of vaccines. In view of the current limited effective prevention and treatment methods for RSV-related diseases, it is necessary to develop effective vaccines, and vaccines of various technical routes, including subunit vaccines, mRNA vaccines,live attenuated vaccines and viral vector vaccines, are under the development stage. Among them, some RSV vaccine prouducts for the elderly and pregnant women have been approved for marketing. This paper mainly outlines the problems encountered in the early development of RSV vaccines, the current status and progress of RSV vaccine research, in order to provide reference for the follow-up development, clinical evaluation and immunization strategy formulation of RSV vaccines.

    2025 11 v.38 [Abstract][OnlineView][Download 919K]

  • Application and evaluation of CNBG product traceability system in the field of vaccine circulation

    ZHANG Wei;LIU Danrui;XU Yuan;MENG Taiqi ;LI Yasong ;ZHOU Jie;Storage & Transportation Department, Wuhan Institute of Biological Products Co., Ltd.;

    Under the background of the rapid development of informatization, intelligent warehousing and logistics, the whole process of product traceability, as an effective supervision method, has promoted the improvement and development of vaccine traceability system. This paper comprehensively expounds the architecture of CNBG product traceability system(including system composition, system association and system operation), and analyzes the application implementation and effectiveness of the system in the field of vaccine circulation based on system application examples to prove that the system can meet various traceability management requirements in the field of vaccine circulation, providing a strong guarantee for the construction of a vaccine traceability system and ensuring the quality safety of the whole process of vaccine circulation.

    2025 11 v.38 [Abstract][OnlineView][Download 976K]


@2012 Editorial Department of "Chinese Journal of Biologicals"