Lipid nanoparticles (LNPs), with spherical vesicle structure composed of one or multiple phospholipid bilayers, are currently the most effective non-viral delivery vehicle in clinical research and application, mainly for the delivery of chemical drugs and nucleic acid molecules. Since the outbreak of COVID-19, mRNA vaccines standed out due to their simplicity of design and manufacture, low immunogenicity, and rapid large-scale production. As an important component, LNPs play an important role in the effectiveness and stability of mRNA vaccines against COVID-19. In this review, the structure, characteristics, application and quality control of the widely used LNPs are summarized, in order to provide a deeper and broader understanding to mRNA vaccine delivery systems and further promote the rapid development of mRNA vaccines and drugs.

According to difference of principle, virus detection methods can be divided into electron microscope method, cell culture method, immunoassay, nucleic acid testing, etc. Electron microscope method with a high resolution can show the structure of the viruses directly, which is suitable for the study of unknown virus. Cell culture methods can obtain the virus activity and detect virus titers. Immunoassay has strong specificity, low cost, and high speed, which is suitable for large-scale population screening. Nucleic acid detection methods allow simultaneous detection of multiple viruses with high sensitivity and strong specificity. Researchers should make a trade-off in the choice of method. This article discusses advantages and disadvantages of virus detection methods and applications, to provide reference for development of more accurate and efficient virus detection, prevention and control strategies.

HPV is a type of DNA virus that exists widely in nature, and the persistent infection of HPV can lead to the occurrence of a variety of cancers, especially cervical cancer. The development and application of HPV vaccine is of great significance for the prevention of cervical cancer and other related cancers. In recent years, with the advancement of science and technology, the research of HPV vaccine has made remarkable progress. There are currently three HPV vaccines on the global market including bivalent, quadrivalent and nine-valent vaccines. The bivalent and quadrivalent vaccines can prevent nearly 70% of HPV-related cervical cancers and precancerous lesions, in contrast, the nine-valent vaccine can provide nearly 90% protection. This article reviews the research progress of HPV vaccines.

Rabies caused by rabies virus is a lethal infectious zoonosis that infects the nervous system. The clinical conditions mainly include hydrophobia, salivation, mania or excitement. At present, there is no specific therapy for rabies and the disease is 100% lethal. The rabies vaccine remains the most effective strategy for rabies prevention and control. This article reviews the epidemiology of rabies, rabies vaccine strains and the research progress of human rabies vaccine in China, in order to provide a theoretical basis for optimizing rabies prevention and control strategies.

Objective　To establish and validate a colloidal gold immunochromatographic assay for the rapid and correct detection of monkeypox virus (MPXV).Methods　The labelling conditions of 7 anti-MPXV A35R monoclonal antibodies were optimized by NaCl precipitation assay.The antibody pairing assay was used to determine the gold standard antibody and the capture antibody, and to optimize the spotting concentration of the antibody detection line (T line) and the quality control line (C line), so as to establish the colloidal gold immunochromatographic detection method. The specificity, limit of detection, stability, repeatability, the hook effect, anti-interference performance of the method were verified.Results　The optimal antibody labelling amounts for 7 anti-MPXV A35R monoclonal antibodies 8D7, 10D2, 2F5, 10G4, 7G5, 10E4 and 2D7 were 0.96, 0.48, 0.96, 0.96, 0.96, 0.96, 0.48 μl, respectively, and the combination of the gold-labelled antibody 2D7 and the test antibody 8D7 was the best pair of antibodies. The optimal concentrations for the T and C line spot samples were 1.00 and 0.50 mg/ml, respectively. Colloidal gold test strips could detect MPXV but not other viruses; the minimum detection limits of MPXV A35R protein and inactivated MPXV solution were 2.34×10^-5 mg/ml and 1.00×10⁵ TCID₅₀/ml, respectively; the colour development of the negative samples was consistent with that of the positive samples; the colour development of 20 test strips was consistent; the concentrations of MPXV A35R protein and inactivated MPXV solution were proportional to the degree of colour development; the test strips were not interfered by interfering substances.Conclusion　A colloidal gold immunochromatographic assay is successfully developed for the rapid detection of MPXV, which has good specificity, detection range, stability, reproducibility and anti-interference performance without hook effect.

Objective　To compare the effects of different signal peptide sequences on secretory expression of influenza virus hemagglutinin (HA) in Sf9 insect cells.Methods　The HA of H1N1 subtype influenza virus A/Victoria/2570/2019 (IVR-215) was used as a target.The full-length HA gene sequence was selected and naïve signal peptide, honeybee melittin signal peptide, glycoprotein 67 signal peptide（Gp67sp）, chitinase signal peptide, and human azurocidin gene signal peptide were connected to the 5´terminal of the gene， respectively. The genes were cloned into the genome of the pFastBac1 transfer plasmid， and five recombinant baculoviruses were constructed by the Bac-to-Bac system. The viral titers were detected by BacPAK^TM qPCR titration kit. The recombinant baculoviruses were used to infect Sf9 cells at different multiplicities of infection, and the cell supernatants were harvested at different time points after infection, and the protein expression levels were detected by immunoblotting to optimize the conditions for HA secretory expression.Results　The titers of the five recombinant baculoviruses harvested were all ＞10⁶ PFU/ml, and the Sf9 cells were able to correctly express and secrete HA, with a relative molecular mass of about 80 000. The amount of HA expressed was the highest when mediated by Gp67sp.The highest expression of HA was achieved when the recombinant baculovirus AcMNPV-Gp67sp-HA infected Sf9 cells for 96 h with multiplicity of infection 5.0 or 10.0, and the hemagglutination efficiency was 256 hemaqqlutinetion units/50 μl.Conclusion　All signal peptides tested effectively guide the secretory expression of influenza virus HA protein in Sf9 cells, among which Gp67sp is particularly advantageous for the efficient secretory expression of HA.

Objective　To evaluate the passage stability of pneumococcal subcultures of all serotypes for pneumococcal polysaccharide conjugate vaccine in a liquid medium.Methods　The culture duration of each generation of pneumococcus in a liquid environment was compared. The characterization of bacteria and capsular polysaccharide antigen gene sequences of the 15th generation (P15) and P4 (control group) strain were analyzed. Then, P15 was fermented and purified according to the production process, and the yield and quality of capsular polysaccharide antigen were analyzed by determination methods and compared with P4.Results　In P7-P15, the culture duration of each generation was similar, and there was no difference in strain identification and antigen gene sequence between P15 and P4, with the sequence similarity of 100%. After the fermentation of P15 strains, there was no apparent difference in the growth of cells, the yield of capsular polysaccharides and the related indexes of capsular polysaccharides compared with P4, and the verification indexes were in line with Cinese pharmacopoeia standards.Conclusion　Pneumococcal subcultures of all serotypes for pneumococcal polysaccharide conjugate vaccine have good passage stability in a liquid environment.

Recombinant virus vector vaccine is a kind of vaccine which uses virus as vector to deliver target antigen. The technical route of virus vector was mostly used for animal vaccines and the pathogen vaccines which were difficult to develop with traditional technical routes in the early stage. Among the five technical routes for the research and development of vaccines against COVID-19 in China, the recombinant virus vector vaccine was approved for marketing and played an important role in the prevention and control of the epidemic because of its advantages of simultaneously producing strong humoral and cellular immunity, being vaccinated through mucosal pathway, and completing the immunization program with one dose. However, due to the pre-existing immune problems of this vaccine, such as factors that may reduce the protective efficacy of the vaccine, and the increase of safety risks in order to enhance immunogenicity, there are few recombinant virus vector vaccine products on the market. With the development of genetic engineering and molecular biology technology, the variety of virus vectors is increasing day by day, and it has been applied in many vaccines. In this paper, the research and development status of recombinant virus vector vaccine is reviewed.

Cancer is one of the leading causes of death worldwide, and its etiology and pathogenesis are still not well understood. Human cytomegalovirus (HCMV) is a type of herpesvirus that encodes multiple oncogenes, activates various important signaling pathways related to cancer, and promotes cancer initiation and development after infection. Studies on breast cancer, glioblastoma, and muscle sarcoma etc have provided important evidence in this regard. This article reviews the molecular mechanisms and clinical researches of HCMV-induced carcinogenesis, aiming to provide new insights and targets for clinical prevention and treatment of HCMV infection.

Objective　To investigate the IgG antibody levels against Neisseria meningitidis serogroups among children after group A and C meningococcal polysaccharide conjugate vaccine (MPCV-AC) was included in the immunization program in Anhui province for 11 years, to provide reference for further optimizing of the immunization strategy against epidemic meningococcal meningitis.Methods　A multi-stage random sampling method was employed to select healthy children under 11 years old from six cities in Anhui province. Questionnaires were conducted and blood specimens were collected.ELISA was used to detect the levels of antibody against groups A, C, Y and W135, and the data were statistically analyzed.Results　A total of 1 002 children were investigated. The proportions of children with protective antibody concentrations against serogroups A, C, W135 and Y were 89.82%, 92.12%, 31.04% and 66.87%, respectively. The median antibody concentrations were 9.93, 10.55, 4.63 and 7.75 μg/ml, respectively. The positive rates of serum anti-Nm IgG antibodies in four different age groups were statistically significantly different, with χ² values of 27.75, 17.70, 132.00 and 122.55, respectively, and P＜0.05. Children who received booster immunization with group A, C, Y, W135 meningococcal polysaccharide vaccine had significantly higher positive rates of anti-Nm IgG antibodies in W135 and Y groups compared to those who received MPCV-AC（χ² values were 10.30 and 38.25, P＜0.05）.Conclusions　The IgG antibody levels against serogroups A and C of Nm among children aged 0 to 11 years in Anhui province are relatively high after the inclusion of MPCV-AC in the immunization schedule, showing significant effectiveness in meningococcal prevention and control. However, the IgG antibody levels against serogroups W135 and Y remain relatively low in this population, indicating a need for strengthened educational efforts to improve the vaccination rate of meningococcal polyvalent vaccine.

Liposomes are artificially prepared nanometer-scale spherical vesicular structures composed of phospholipids, cholesterol, surfactants, etc. Liposomes exhibit a variety of potential applications in drug delivery and vaccine development due to advantages such as flexible charge, particle size regulation and surface modification properties. In cancer therapy, liposomes have become an important tool for improving therapeutic efficacy and reducing side effects by enhancing drug targeting, improving drug biocompatibility and stability and controlling drug release. Liposomes are the key technology for improving drug delivery efficiency and enhancing the efficacy of anti-tumor treatments. This article reviews the current state of the application of liposomes in pharmaceuticals to provide reference for developing novel liposomes.

Immunoglobulin A nephropathy (IgAN) tends to occur in young people, and some patients may develop end-stage kidney disease, causing serious family and social burden, but for current clinical treatment, IgAN-specific drugs are scarce. With the development of disease cognition, drugs discovery and drug development technology in recent years, it is hopeful that IgAN could be overcome in the near future. At present, the recognized pathogenesis of IgAN is the "four-fold strike" theory, in which the complement system plays an important biological role in the occurrence and development of IgAN. In this paper, the basic concept, components, activation pathway, function and related drug discovery and development progress of the complement system are briefly introduced. Then, the pathogenesis of IgAN and the role of the complement system in IgAN are analyzed in detail, as well as the strategies for treating IgAN based on the complement system, and the discovery and development of new drugs are discussed. Finally, the prospects and challenges of the complement system drugs in the treatment of IgAN are prospected.

Objective　To establish a rapid method for the detection of mycoplasma using real-time quantitative PCR (qPCR) technology.Methods　Eight mycoplasma species available for validation under European Pharmacopoeia（10.0 edition） 2.6.7 were selected. Specific primers were designed based on mycoplasma 16S RNA sequence. A qPCR method for the detection of mycoplasma was established and validated for specificity, sensitivity and reproducibility.Results　Suitable primers for the detection of mycoplasma by qPCR were successfully screened. Eight mycoplasma species showed a cycle threshold (CT) between 20-30 at a concentration of 10⁵-10⁶ CFU/mL, and there was no CT for the bacterial control at same concentration. Except for Mycoplasma sperminum, which could be detected at a concentration of 10-100 CFU/mL, the other 7 mycoplasma species could be detected at a concentration of 10 CFU/mL. CTs were between 35-40 at 10 CFU/mL. In specificity experiment that repeated 3 times, the control bacteria were all undetected, CTs of 8 species within 10⁵-10⁶ CFU/mL were between 20-30, and for each species, the coefficient of variation (CV) for 3 times was less than 10%. In sensitivity experiment that repeated 3 times, the CTs of each species with different dilution concentrations were between 20-40 in each experiment, and the CVs were less than 10%.Conclusion　The established method has good specificity, sensitivity and reproducibility, and can support timely control of possible mycoplasma contamination risks in biologics.

Objective　To evaluate the immunogenicity of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and influenza virus combined vaccine formulated with 4 adjuvants Al(OH)₃, MF59, AS03 and QS21 in mice.Methods　Adjuvants Al(OH)₃, MF59, AS03 and QS21 were used respectively to prepare SARS-CoV-2 (inactivated) and tetravalent influenza virus (split) combined vaccine. BALB/c mice were immunized by intraperitoneal injection on day 0 (D0) and D14, respectively. Blood samples were collected on D14 and D28 to detect antibody titers against SARS-CoV-2 and hemagglutination inhibition titers of influenza. Spleen lymphocytes were isolated on D28 to detect cellular immune responses to both SARS-CoV-2 and influenza virus.Results　SARS-CoV-2 and influenza virus combined vaccine with different adjuvants induced both antigen-specific antibody responses and cellular immune responses in mice. At D28 post-initial immunization, MF59 adjuvant group induced high levels of SARS-CoV-2 binding antibodies and neutralizing antibodies, with geometric mean titers (GMTs) of 89 144 and 5 418, respectively, and MF59 group also induced high levels of hemagglutination-inhibiting antibodies against the quadrivalent influenza virus strains (H1N1, H3N2, BV, BY), with GMTs of 4 457, 5 120, 1 470 and 5 881, respectively. Both the MF59 and AS03 groups induced robust Th1-type (IFN-γ, IL-2) cellular immune responses against SARS-CoV-2, with spot forming units (SFUs) statistically significantly higher than those of Mock group （IFN-γ： H=16.69，P＜0.01； IL-2： H=15.21， P＜0.05）. The AS03 group induced a strong Th1-type (IFN-γ, IL-2) cellular immune response against the quadrivalent influenza virus strains (H1N1, H3N2, BV, BY), with SFUs statistically significantly higher than those of Mock group （IFN-γ： H=12.93, 12.17, 11.82, 13.61, P＜0.05； IL-2： H=12.24, 12.42, 11.72, 12.43， P＜0.05）.Conclusion　The immunogenicity as well as specific antibody and cellular immune responses of SARS-CoV-2 and influenza virus combined vaccine with different adjuvant formulations are different, indicating the importance of adjuvants in the development of combined vaccine formulations.

Objective　To evaluate the stability of the self-made aluminum phosphate adjuvant under different storage conditions so as to provide supportive data for the storage and expiration date of the adjuvant.Methods　Three consecutive batches of industrial-scale self-made aluminum phosphate adjuvant were selected and stored at (5±3) ℃ and (25±2) ℃, respectively. The main evaluation indices of aluminum phosphate adjuvant (appearance, pH value, aluminum content, adsorption rate, sodium chloride content, analysis of sedimentation supernatant, sterility, bacterial endotoxin, arsenic salt, iron salt, sulfate salt, ammonium salt, heavy metals, phosphorus to aluminum molar ratio, zero charge point, particle size and distribution) were investigated by sampling at regular intervals.Results　The appearance and sterility of 3 batches of aluminum phosphate adjuvant were qualified after stored at two temperatures for 15 months. The pH value was 5.4-6.2, the adsorption rate was 93%-100%, the aluminum content was 2.77-3.36 mg/mL, the sodium chloride content was 7.90-8.60 g/L, and the sedimentation supernatant analysis ranged from 12 to 15 mL .The bacterial endotoxin ranged from 0.5 to 3.0 endotoxin unit/mg aluminum, and the molar ratio of phosphorus to aluminum ranged from 0.82 to 1.13. The particle size corresponding to the cumulative distribution reaching 10%, 50% and 90% were 1.75-2.53, 2.87-3.98 and 4.94-6.90 μm, respectively. The zero charge point was 5.11-5.47, with the residual impurity of arsenic salt ≤0.001‰, iron salt ≤0.015‰, sulfate salt ≤0.5%, ammonium salt ≤0.005% and heavy metal ≤0.002%. All main evaluation indices of the adjuvant were stable and in line with the quality standard.Conclusion　Three batches of self-made aluminum phosphate adjuvant show good stability after being stored at (5±3) ℃ and (25±2) ℃ for 15 months, which provides a basis for the formulation of storage conditions and efficacy of aluminum phosphate adjuvant.

Objective　To structurally optimize and preliminarily identify the biological activity of anti-tumor necrosis factor receptor superfamily member 4 (OX40) monoclonal antibody Ab18.Methods　The species cross-binding activity of Ab18 to OX40 of different species was measured by ELISA. The binding activity of Ab18 to HEK293-hOX40 cells was measured by flow cytometry (FCM). The blocking activity and agonistic activity of Ab18 were measured by reporter gene methods. The Fc fragment of Ab18 was engineered by point mutagenesis technology to change the affinity of the Fc fragment with Fc receptors. ELISA and FCM were used to evaluate the binding activity to human Fcγ receptor ⅡB and human neonatal Fc receptor, and the reporter gene method was used to measure the agonistic activity and antibody-dependent cell-mediated cytotoxicity (ADCC) activity of Ab18 and its mutants.Results　The Ab18 antibody had species cross-binding activity with mouse, rat and rhesus monkey OX40s, and stronger binding activity and stronger blocking activity with HEK293-hOX40 than the control antibodies GBR830 and KHK4083. The median effect concentration (EC₅₀) value of binding activity of Ab18 with HEK293-hOX40 was 366.9 ng/ml, and the half maximal inhibitory concentration value of blocking activity was 657.3 ng/ml. In addition, Ab18, GBR830 and KHK4083 showed agonistic activity. The ADCC activity EC₅₀ value of Ab18 was 25.7 ng/ml, and the agonistic activity EC₅₀ value was 14.9 ng/ml. The mutated Ab18 had lower agonistic activity, better binding activity with human neonatal Fc receptor, consistent ADCC activity, and good biological activity.Conclusion　The Ab18 antibody is successfully optimized, showing better biological activity.

Objective To investigate the mechanism of pyroptosis production in colorectal cancer cells by Newcastle disease virus (NDV) via natural killer (NK) cell activation through granzyme A (GZMA) secretion, with specific focus on the role of gasdermin B (GSDMB) expression level in this process. Methods NK cells were co-cultured with NDV at different concentrations [NDV1, 2, 3: 0.001, 0.010, 0.100 hemagglutinin/mL] for 16 h, and blank control (culture medium only) as well as non-stimulated NK cell control were set. All groups were subsequently incubated with high (SW837) or low (SW480) GSDMB expression colorectal cancer cells. Quantitative PCR and ELISA were used to detect the gene expression and protein secretion levels, respectively, of effector molecules in NK cells. Cell proliferation, cytotoxicity and lactate dehydrogenase (LDH) release were measured to assess the cell viability and pyroptosis in 2 tumor cell lines. Cell morphology observation and Western blot of GSDMB cleavage were applied to verify NK cell-mediated pyroptosis effect. Results Quantitative PCR and ELISA results indicated that NDV stimulation upregulated the gene expression and protein secretion levels of GZMA, IFN-γ, and perforin in NK cells. The highest protein secretion levels of GZMA (t = 6.70, P = 0.003) and IFN-γ (t = 13.66, P < 0.001) were reached in NDV3 stimulation, and were statistically significantly higher than non-stimulated NK control. Comparing with non-stimulated NK cell control treated, cell viability inhibition [NDV3-NK at effector-to-target ratio (E/T) =1, 2: t = -10.17, -25.94, both P < 0.001], LDH release (NDV3-NK at E/T=1, 2: t = 13.70, 9.75, both P < 0.001), and N-terminal GSDMB (NDV2-NK at E/T=1, 2: t = 6.80, 3.07, both P < 0.05) of with stimulated NK cell treated SW837 all increased statistically significantly. Pyroptosis-related features such as plasma membrane blebbing and swelling were observed in SW837, but not in SW480 cells, even though viability inhibition and cell damage were seen in SW480. Conclusion NDV-activated NK cells induce pyroptosis in colorectal cancer cells via GZMA secretion, particularly in cells with high GSDMB expression.

Objective　To establish a method for the detection of biological activity of human growth hormone (hGH) injection based on hGH promoting proliferation of the rat lymphoma cell line (Nb2-11 cell) and to validate the method.Methods　The proliferation of hGH stimulated cells was measured by luminescent cell viability assay, and the relative potency of hGH was calculated by the 4-parameter logistic regression model. The detection method was established by screening the analysis medium formulae (1%, 5% and 10% horse serum), cell treatment methods (cells starved for 24 h or without starvation treatment) and cell densities (5 000 or 8 000 cells/well). The Nb2-11 cell proliferation method adopted was verified for specificity, accuracy, precision, linearity, durability and range according to the general requirements 9401 of Chinese pharmacopoeia 2020 edition volume Ⅳ.Results　The optimal curve fit was obtained using assay medium containing 5% horse serum, cells starved for 24 h, and a density of 8 000 cells/well. The hGH preparation buffer had no effect on the detection results. The linear regression equation was: y=0.942 8x + 0.072 6, with a coefficient of determination of 0.980 9, and the linear range was 60%-140%. The single experiment recovery rates of different active samples were 70%-130%. The coefficient of variation of the validation experiment was less than 20%.Conclusion　A method for determination of biological activity of hGH injection is successfully developed, which shows good specificity, accuracy, precision, linearity and durability, and can be used for the biological activity detection of hGH injection.

Objective　To establish and verify a pharmacokinetic detection method for rabbit anti-human thymus/lymphocyte immunoglobulin (r-ATG) , for detection of the total concentration of r-ATG in the sera of patients after clinical medication.Methods　Mouse anti-rabbit IgG was used to coat the ELISA plate, and horseradish peroxidase-labeled goat anti-rabbit IgG was used as the secondary antibody to establish a double-antibody sandwich ELISA method for the detection of r-ATG content in human serum. The linear range, parallelism, repeatability, intermediate precision, accuracy, specificity, and stability of recovery rates for standard curve concentrations of reference materials at different time points of the method were verified. Clinical blood samples of 5 patients were detected and pharmacokinetic analysis was carried out by the established method.Results　When the initial concentration was 0.25 μg/ml and the dilution times were between 2¹-2⁹, the reference concentration and the absorbance value showed an S curve, the coefficient of determination (R²) was > 0.99, and the recovery rate was 90%-110%. The coefficient of variation (CV) of slope of linear equation between reference sample and three blood samples was < 10%, with R² > 0.99, CV of R² < 5%, showing good parallelism . The repeatability CV and intermediate precision CV were <15%, and the recovery rates were all 90%-110%. The average recovery rates of accuracy verification results were 80%-120%. There was statistically significant difference between the sera of the injected patients, healthy people, and non-injected patients, the t-values were 32.607 and 21.949, respectively, with P <0.01. The average recovery of each concentration in the standard curve was 90%-110% when the reference was stored at 2-8 ℃ for 16 months, and the relative standard deviation of the recovery at different time points of each concentration was less than 15%. The serum concentration of r-ATG reached the highest value on the 5th day after administration, with an average half-life time of (6.5±5.3) d.Conclusion　The established method has good dilution linearity and parallelism, repeatability, intermediate precision, accuracy, specificity between samples, and the recovery rate of each concentration in the reference standard curve at different time points is stable, thus can be used for the detection of serum concentration after clinical administration of r-ATG.

Objective　To establish and validate a method for the detection of dimethyl sulfoxide (DMSO) in pneumococcal polysaccharide conjugate bulk.Methods　The mobile phase, column temperature and flow rate in the reversed phase high performance liquid chromatography(RP-HPLC) were optimized to establish the optimal detection method. The method was validated in terms of specificity, linearity, accuracy, precision and stability.Results　The RP-HPLC method was optimized with 10% acetonitrile as the mobile phase at a flow rate of 0.6 ml/min and a column temperature of 35 ℃. The relative errors of this method for the detection of DMSO content in the test and control groups of the interfering components ranged from -2.6% to 2.1%, and the resolutions were all ≥1.5; the concentrations showed good linearity with the peak areas, and the regression coefficient of determination was ≥0.990 0; the biases of different concentration levels ranged from -5.31% to -0.89%, and the upper limit of bias in 95% was≤8.0%; the precision of different concentrations ranged from 0.70% to 4.45%, and the upper limit of confidence of the precision in 95% was ＜8.0%. The evaluation of the comprehensive ability of the method showed that the total method variation was ≤7.0% in the range of 4-50 μg/ml. Both the 95% prediction interval and the 95%/90% tolerance interval were in the range of 80%-120%. The relative standard deviation of the mean peak area of the test solution within 20 h were ≤2.0%.Conclusion　The method for the determination of DMSO in pneumococcal polysaccharide conjugate bulk is successfully established, which shows good specificity, linearity, accuracy, precision, stability and high comprehensive evaluation ability.

Adenovirus vectors, renowned for their efficacy as gene delivery tools, have garnered significant attention in vaccine research over the past decades. Currently, adenovirus vectors have been successfully applied to the vaccine development of Ebola virus and severe acute respiratory syndrome coronavirus 2, and have shown good protection and safety. However, the application of adenovirus vectors in vaccines still faces challenges, such as interference of pre-existing antibodies and poor targeting. This article provides a review of the application, existing problems, and improvement directions of adenovirus vectors in vaccine research.

Pertussis is a highly contagious respiratory disease. With the increase in pertussis cases in recent years, it has become more and more concerned. Currently, vaccination is the most effective prevention method for pertussis, and developing new vaccines is a major priority in pertussis prevention and treatment. This paper summarizes the current pertussis epidemiological trends, analyzes the causes of pertussis resurgence, and tracks the latest progress in pertussis vaccine development, in order to provide a reference for developing novel pertussis vaccines.

Objective To establish and compare 3 tetanus antibody detection methods in vitro. Methods The indirect method, double antigen assay and toxin binding inhibition (ToBI) test were established. The developed methods were validated for linearity and range, intermediate precision, accuracy and detection limit. Results The linear range of indirect method was 0.007 8-1.000 0 mili-international unit (mIU)/mL [coefficient of determination (R²) = 0.998 6], of double antigen assay was 0.078-10.000 mIU/mL (R²=0.998 7) and of ToBI test was 1.95-31.25 mIU/mL (correlation coefficient > 0.98). The validation results indicated that three methods meet the guiding principles of analytical method validation in terms of precision, accuracy, and specificity. Comparing the positive detection rates of these methods, there was no statistically significant difference (χ²=1.34，P=0.513). Conclusion Three detection methods for tetanus antibody in vitro, namely indirect method, double antigen assay and ToBI test, are successfully established, laying research foundation for in vitro method to replace in vivo neutralizing assay.

Vaccine is a special drug containing bioactive substances, and the storage and transportation of vaccine have certain cold chain requirements on temperature. After the Shandong vaccine case, a series of laws and regulations have been issued to further strengthen the management of vaccine production, circulation and vaccination. In this paper, the laws and regulations related to vaccine circulation management are systematically combed and analyzed in chronological order, aiming at clarifying the context of laws and regulations for vaccine production, distribution, storage enterprises and vaccination agencies, and providing legal and regulatory basis for them to better carry out vaccine circulation management.

Objective　To investigate the applicability of cytopathic inhibition assay for the detection of human cytomegalovirus neutralizing antibody (CMV-NA) potency in plasma and human cytomegalovirus immunoglobulin for intravenous injection (CMV-IVIG).Methods　Cytopathic inhibition assay for detection of CMV-NA potency was established with cytomegalovirus immunoglobulin national standard substance. The repeatability, intermediate precision, specificity, accuracy, durability and reproducibility of cytopathic inhibition assay were investigated with CMV-NA plasma and CMV-IVIG.Results　The relative standard deviations of the repeatability validation of CMV-NA potency for plasma and CMV-IVIG were 12% and 19%, respectively. The CMV-NA potency of plasma and CMV-IVIG were detected 6 times by 2 experimentalists, with F values of 0.84 and 0.03, respectively, and P > 0.05. With 6 repetitions of plasma and CMV-IVIG for CMV-NA potency tested on different dates, the F values were 0.89 and 0.55, respectively, with P > 0.05. Plasma, CMV-IVIG and national standard substance were free of cytopathy. The recovery rates of both plasma and CMV-IVIG labeled samples were within 100% ± 40%. The CMV-NA potencies of plasma and CMV-IVIG were tested at various neutralization time, and the F values were 0.48 and 0.04, respectively, with P > 0.05. The CMV-NA potencies of plasma and CMV-IVIG were detected in different viral titer ranges with F values of 0.49 and 0.11, respectively, and P >0.05. The CMV-NA potency of CMV-IVIG was detected by 2 laboratories with a t value of 0.69 and P >0.05.Conclusion　The cytopathic inhibition assay for CMV-NA potency has good repeatability, intermediate precision, specificity, accuracy, durability and reproducibility.

Infants and young children are at high risk of influenza, and vaccination is the most cost-effective way to prevent influenza. Currently, trivalent and quadrivalent influenza vaccines are available for children in China. The dosage forms include half dose (7.5 μg hemagglutinin/0.25 ml) and full dose (15 μg hemagglutinin/0.5 ml). Studies have found that the age of infants and young children is positively correlated with the level of immune response, and the immune effect of influenza vaccine for infants and young children can be improved by increasing the antigen content. Full-dose quadrivalent influenza vaccine for infants and young children has lately come to market in the Chinese mainland. This article reviews the epidemic status of influenza virus and the application progress of full-dose quadrivalent influenza vaccine for Chinese infants and young children from the aspects of disease burden of influenza, vaccination status, protective effect, immunogenicity and safety of influenza vaccine, and vaccination intention.

Objective　To validate a quantitative ELISA method for the detection of residual host cell protein (HCP) in recombinant hepatitis B vaccine (Saccharomyces cerevisiae).Methods　The S. cerevisiae HCP ELISA Kit was used to detect HCP residues in recombinant hepatitis B vaccine (Saccharomyces cerevisiae), and the linearity, accuracy, repeatability, limit of quantification, specificity and intermediate precision of the method were verified.Results　The ELISA standard curve had good linearity (coefficient of determination ≥ 0.97), with the lowest absorbance (A_405/492) value <0.300, and the highest A_405/492 value >1.000. With same experimenter, samples added with low, medium and high concentration standards were detected 3 times, and the recovery rates were 80%-120%. Medium concentration standard-added sample was detected 6 times, and the relative standard deviation (RSD) was <10%, indicating good accuracy and repeatability of the method. The A_405/492 values of the sample background solution and the sample diluent were both smaller than the lowest point of the curve, indicating that the method was highly specific. The medium concentration standard-added sample was detected by different experimenter groups at different time for 6 times, and the RSD was <10%, indicating good intermediate precision of the method.Conclusion　The ELISA method for the detection of HCP residues in recombinant hepatitis B vaccine (Saccharomyces cerevisiae) has good linearity, accuracy, repeatability, limit of quantification, specificity and intermediate precision, thus can be used for the process control of hepatitis B vaccine production.

Objective　To establish and verify the verification method of virus inactivation of Sabin strain inactivated poliovirus vaccine (Vero cell)(sIPV).Methods　Poliovirus types Ⅰ, Ⅱ and Ⅲ were inoculated into Vero cells by a series of gradient dilution, and then amplified by blind cell passage twice. The cytopathic changes were observed, the minimum detection limit of this method and cell sensitivity to virus were explored, and the precision (repeatability and intermediate precision) and applicability of this method were verified.Result　The detection limits of the established virus inactivation validation method for poliovirus types Ⅰ, Ⅱ and Ⅲ were ﹣2, ﹣1 and 0 lgCCID₅₀/mL, respectively. The minimum inoculation concentration of poliovirus types Ⅰ, Ⅱ and Ⅲ to Vero cells was 2 lgCCID₅₀/mL, and the Vero cell lesions were all observed on day 7 .Thus, the sensitivity of the cell was 2 lgCCID₅₀/mL. The same inspector repeatedly tested the same batch of samples 6 times, and the test results were consistent. The same batch of samples were tested 6 times by different inspectors at different time, and the test results were consistent. The result of sample stored at ﹣70 °C or below for 7 d was consistent with that of unfrozen sample.The inactivation verification test was carried out on 3 batches of types Ⅰ, Ⅱ and Ⅲ monovalent inactivated virus solution and bulk from sIPV production process, and the inactivation verification test results of the samples tested on the day of sampling and after 7 d of storage at ﹣70 °C or below were consistent, indicating that the precision and applicability of the method were good.Conclusion　The sIPV virus inactivation validation method effectively detects the inactivation status of the sample, which can be used to evaluate the virus inactivation effect and monitor key steps such as intermediates in the process.

Drug marketing authorization holder takes the main responsibility for drug quality and safety. This paper reviews the implementation of the drug marketing authorization holder system at home and abroad, studies the regulations on the supervision and administration of drug marketing authorization holder implementing the main responsibility of drug quality and safety, which was implemented on March 1, 2023, and further comments on the responsibilities and requirements of drug marketing authorization holder in carrying out drug quality management.

Objective　To benchmark the quality control indicators of the homemade rabbit derived type 5 pneumococcal specific serum by comparing with the commercial reference products, in order to detect the process samples in the production of 23-valent pneumococcus polysaccharide vaccine.Methods　The non-competitive turbidity mode of the dual path rate turbidity immunoassay system was used for detection. The working concentration, linearity, crossover, comparability with commercially available serum, detection difference both between and within laboratories， stability under working concentration of type 5 sera were investigated in turn.Results　The maximum dilution multiples of the working concentrations for the 4 batches of home made type 5 sera were 75, 20, 30 and 8 times, respectively. Under the corresponding working concentrations, the polysaccharide concentration showed a good linear relationship with the response value in the range of 0.625-4.000 μg/ml, and the linear correlation coefficients were all > 0.985. There was no crossover between 4 batches of type 5 sera reference samples and the other 22 types, and the specificity was good. In the comparability test with commercially available serum, the coefficient of variation (CV) of type 5 polysaccharide content in 3 different manufacturers' marketed vaccines was less than 5% for all 5 batches of sera including commercially available serum. There was no apparent difference in the detection results between the 4 batches of type 5 sera and commercially available serum as the weight-average molecular weight decreased. The CV values of between- and within-department outdoor testing results were less than 10%. Four batches of type 5 sera with working concentration maintained good stability for at least one month under 2—8 ℃.Conclusion　The homemade rabbit derived type 5 pneumococcal specific serum has high potency, good linearity,no crossover, comparability to commercially available serum， and good stability at working concentrations， and can be used for polysaccharide content detection of type 5 pneumococcus polysaccharide vaccine.

Objective　To establish an environmental bacterial bank in clean area by monitoring, collection and identification of environmental bacteria in pilot clean area of research and development.Methods　Static and dynamic environmental monitoring was carried out in the pilot clean area of research and development, and the collected environmental bacteria (settling microbe, airborne microbe and surface microbe) were identified by morphological observation, staining microscopy and genotypic microbial identification (bacterial 16S ribosomal RNA sequencing analysis).Results　A total of 89 strains of environmental bacteria were isolated, belonging to 16 genera, among which Bacillus sp. accounted for 29%, Staphylococcus sp. accounted for 27% and Micrococcus sp. accounted for 12%.Conclusion　The environmental bacterial bank in pilot clean area of research and development is initially established, which provides technical support for microbial contamination traceability.

Objective　To validate the storage potency of phenol red-free M199 medium and explore the effect of phenol red-free M199 potency on the key quality attributes of phenol red-free Sabin strain inactivated poliovirus vaccine (sIPV).Methods　The appearance, clarity, pH, weight loss on drying, bacterial endotoxin, microbial limit and other physical and chemical properties of 3 batches of phenol red-free M199 powder culture medium stored for 0, 6, 12, 18, 24, 30, 36 and 42 months were monitored. Key quality attributes of final sIPV products prepared with phenol red-free M199 calibration qualified after 36 months of storage were detected. The sIPVs prepared with near- and far-period phenol red-free M199 were stored at 37 ℃ and 25 ℃, respectively, for stability monitoring and evaluation by detecting the D antigen content.The 2 groups of sIPVs were used to immunize rats, and the immunogenicity was evaluated by detecting the antibody potency of rat serum. Results　The appearance, clarity, pH, loss on drying, bacterial endotoxin, microbial limit of phenol red-free M199 dry powder culture medium stored at 2~8 ℃ for 42 months were all qualified. The pH, D antigen content, protein content, Vero cell DNA residue, bovine serum albumin residue, Vero cell protein residue, free formaldehyde content and other test results of sIPV produced using phenol red-free M199 stored for 36 months were in line with the requirements of the Chinese pharmacopoeia 2020 edition volume Ⅲ and the enterprise standards. The sIPV prepared with phenol red-free M199 stored up to 36 months was potent with good stability, and the trend of immunogenicity was consistent with that of the sIPV prepared with far-period phenol red-free M199, with no statistically significant differences in potency（t =﹣0.15-2.17, P＞0.05） or neutralizing antibody level（t =﹣1.46-2.02, P＞0.05）.Conclusions　The sIPV final product prepared with phenol red-free M199 stored up to 36 months has good stability. The product quality attributes, immunogenicity and stability of sIPV products prepared with near- and far-period M199 show the same trend, and near-period M199 has no effect on the product quality. Phenol red-free media can be valid for 36 months.

Objective To study toxic reactions and active systemic allergic reactions to live attenuated Japanese encephalitis vaccine (JEV) in animals after subcutaneous vaccination to examine vaccine safety.　Methods　The acute toxicity of JEV was evaluated by subcutaneous injection single administration toxicity test in Sprague-Dawley（SD） rats, and the sensitization of JEV was evaluated by active systemic anaphylaxis in guinea pigs.Results　JEV was administered subcutaneously to SD rats at a dose of 1.50 mL per animal, and no toxic reactions related to the test substance were observed. The maximum tolerable dose was 1.50 mL per animal. JEV was administered subcutaneously 3 times at 0.05 mL/piece （low dosage）and 0.50 mL/piece （high dosage） for sensitization, and intravenously at 0.10 mL/piece （low dosage）and 1.00 mL/piece （high dosage） for stimulation, respectively, 14 days after the last sensitization. Active systemic allergic reactions were observed in guinea pigs.Conclusion　After subcutaneous administration of JEV in animals, the toxic reactions and active systemic allergic reactions meet the requirements of animal safety evaluation.　

Objective　To investigate the leaching level of extracts in disposable liquid storage bags directly in contact with mumps virus stock solution under cryopreservation, in order to evaluate the safety of mumps virus stock solution cryopreservation in disposable storage bags.Methods　The mumps virus simulant liquid stored in disposable liquid storage bags and standard schott bottles was kept in -60 ℃ freezer. Three samples were taken out at 0, 6, 12 and 15 months, respectively. Extract detection and safety dose analysis were conducted using acetone, tert-butanol and isopropanol detected in the extraction experiment as target substances.Results　The highest detection peaks of acetone, tert-butanol and isopropanol were detected at 15 months, and the extracts concentrations reached 415.02, 275.82 and 936.92 μg/L, respectively. The extracts concentrations in disposable storage bags at each detection time point were lower than the allowable daily exposure or safety assessment threshold.Conclusion　The extract content of mumps virus simulated liquid in disposable liquid storage bag under cryopreservation does not exceed the safety range, and the safety risk is small.

Objective　To investigate the trends in HBV infection and serological dynamics in low birth weight infants born to HBV surface antigen (HBsAg)-positive mothers, and to analyze HBV infection and the effectiveness of immunization in this very special group of children.Methods　Low birth weight infants born to HBsAg-positive mothers in 24 hospitals or obstetrics departments in Lu'an City from 2016 to 2021 were selected, categorized and counted by name, gender, weight, preterm delivery, hepatitis B vaccine（HepB） timely vaccination, vaccination dose, etc. The concentrations of HBsAg and hepatitis B surface antibody (HBsAb) were quantified by magnetic particulate chemiluminescence, and the antibody positivity rate and geometric mean concentration (GMC) of antibodies were analyzed and compared.Results　A total of 172 low birth weight infants were included in this study, of which 121 completed blood sample collection, with a mean age of (3.7±1.7) years, a median weight of 2.30 kg, a proportion of 58.68% of preterm infants, a proportion of 75.21% of infants who received the first dose of HepB timely The number of subjects who completed HepB primary and booster immunizations were 117 and 4, respectively, and the prevalence of HBV infection was 5.79%. Among the uninfected subjects, there was a statistically significant difference between HBsAb GMC of the booster immunization group and the primary immunization group, with Z=-2.17, P=0.030. Among the uninfected and primarily vaccinated subjects,timely vaccination preterm delivery, and birth weight diden´t effect GMC. Among the HBsAb positive subjects, no statistically significant difference in the positivity rate was seen in the primary immunization group compared to the booster immunization group (χ²=2.45, P=0.118); but there was a statistically significant difference between the two groups in GMC (Z=-2.25, P=0.024). In primarily vaccinated and uninfected subjects, there was an overall trend of decreasing HBsAb positivity and GMC after vaccination.Conclusion　In Lu'an, the HBV infection rate of low birth weight infants born to HBsAg-positive mothers is high, the HepB vaccination rate is low, and the total levels of HBsAg and HBsAb are low, so it is necessary to strengthen the implementation of 0-1-2-7 months HepB immunization program for low birth weight infants.

Objective　To initially compare the effect of 23-valent pneumococcal polysaccharide vaccine large-scale production by the plant-derived and animal-derived culture media using Streptococcus pneumoniae type 6B as the experimental serotype.Methods　Three batches of scale production were carried out for pilot production using soybean peptone culture medium and pancreatic peptone culture medium. The growth of Streptococcus pneumoniae was tested by determining the content of polysaccharide by rate turbidimetry in each process stage and calculating the recovery rate, verifying the harvested refined polysaccharide according to the manufacturing and verifications regulations of the 23-valent pneumococcal polysaccharide vaccine, and taking the nuclear magnetic resonance (NMR) detection. Then a comparative analysis was conducted systematically.Results　Compared to the pancreatic peptone culture medium, in average， bacterial concentration and the polysaccharide content in fermentation broth of serotype 6B strain in soybean peptone culture medium were 22.5% and 88.7% higher, respectively, and the yield of crude polysaccharide and refined polysaccharide were 66.87% and 139.67% higher, respectively. All quality indexes of the refined polysaccharide met the requirements of the registration standards. The fingerprint areas of NMR spectra of both refined polysaccharides were consistent, and the chemical shifts and peak types were consistent with those reported in literatures.Conclusion　The qualities of refined polysaccharides produced by soybean peptone medium and pancreatic peptone medium are comparable and soybean peptone medium improves the yield of refined polysaccharide.

The drug qualified persons（QPs） engage in the quality management activities of pharaceutical enterprises, take responsibilities for the product release，and ensure that the production and inspection of each batch of released products meet the relevant laws and regulations, drug registration requirements and quality standards.The professional ethics and legal consciousness of the drug QPs directly affect the quality and safety of drugs. This paper states the basic situation and existing problems of the implementation of the drug QP system in China, as well as puts forward professional ethics requirements for the drug QP, and sorts out the legal responsibilities and risks that the drug QP needs to bear, aiming to improve the professional quality and legal awareness of drug QPs and further ensure the ability of drug QPs to perform their duties.

Objective　To study the effect of ultrafiltration concentration of influenza B virus liquid based on MDCK cell culture using ultrafiltration membrane package and hollow fiber column under different process parameters.　Methods　Six batches of influenza B virus clarified liquid based on MDCK cell culture were prepared. Ultrafiltration membrane packages and hollow fiber columns were used to make 2× concentrated influenza B virus clarified liquid, which was then washed and filtered. Hemagglutination titers of the virus concentrates were measured using chicken erythrocyte suspensions, protein content was determined by Lowry method 2 for the caculation of antigen recovery and miscellaneous protein removal rate, in order to investigate the effects of different washing times and types of ultrafiltration membranes on the above indices. Ultrafiltration concentration was carried out on the virus filtration solution that was washed and filtered 10 times to analyze the virus residue under different membrane cleaning times after ultrafiltration concentration using two types of ultrafiltration membranes. The stability of the virus concentrates stored at various temperatures was evaluated. Results　After filtration through the ultrafiltration membrane package and hollow fiber column, the hemagglutination titer of the virus filtrate remained stable, and the miscellaneous protein removal rate was relatively high at 5 times of filtration, reaching more than 90%. The antigen recovery rate of virus concentrates treated by hollow fiber column (61.95%) was slightly higher than that of ultrafiltration membrane package (49.91%), while the overall protein removal rates of the virus concentrates using both types of membranes were around 96%. After the ultrafiltration membrane package and hollow fiber column were washed once following ultrafiltration concentration, residual virus antigen in the ultrafiltration system accounted for over 10% of the antigen content of the virus clarification liquid. The virus concentrates processed with two types of membranes showed high stability after being stored at 2-8 ℃ for 1 week, while the hemagglutination titer dropped dramatically after being stored at ﹣70 ℃ for 1 week. Conclusion　When the virus clarification liquid is washed and filtered using ultrafiltration membrane packages and hollow fiber columns 5 times and above, the virus solution obtained has high viral content and low heterogeneous protein content.

Objective To establish a double antibody sandwich ELISA for quantitative detection of recombinant porcine trypsin (RPT), and to perform methodological validation and preliminary application. Methods A double antibody sandwich ELISA for quantitative detection of RPT content was established.Paired mouse monoclonal antibodies against RPT were selected through antibody screening, optimal working concentrations of coating and enzyme-labeled antibodies as well as type and concentration of blocking agent were determined. The linear range, sensitivity, accuracy, precision and specificity of the method were determined. This established method was used to detect the content of RPT in viral background liquid during the vaccine production, and to verify the detection efficiency of this method for different brands of natural extracted porcine trypsin. Results The coating antibody was determined to be RPT-5C2D12 with optimal working concentration of 6 000 μg/mL， the enzyme-labled antibody was RPT-1E10A5 with optimal dilution of 1∶5 000, and the blocking solution was 1% bovine serum albumin. A four-parameter logistic fitting standard curve achieved correlation coefficient ≥0.990 and detection range of 0.156-40.000 ng/mL. Limits of quantification was 0.156 ng/mL. Spike recovery rate for accuracy ranged from 92.3% to 106.7%. Precision validation showed inter-assay coefficient of variation (CV) ≤6.7% and intra-assay CV ≤5.9%. Except for RPT, there was no cross-reactivity with other control proteins. The spike recovery rate for three batches of viral background fluid samples ranged from 87.4% to 98.2%. The relative detection efficiency for two brands of natural extracted porcine trypsin was 59% and 68% compared to RPT. Conclusion The established double antibody sandwich ELISA for quantitative detection of RPT has good linearity, sensitivity, accuracy, precision, and specificity, and can be used for assessing trypsin residues in vaccine type biological products.