Senecavirus A (SVA) is a major viral pathogen causing disease in pigs, and effective monitoring of SVA infection is critical for disease control. In this study, we aimed to develop a reliable ELISA method for rapidly detecting neutralizing antibodies against SVA. We used HEK293F cells to express an SVA-specific porcine Fab antibody and verified the biological activity of the Fab antibody by indirect ELISA, immunofluorescence assay, virus neutralization test, and Western blotting. The Fab antibody was biotinylated and used as a competitive antibody to establish a competitive ELISA (C-ELISA) for detecting neutralizing antibodies against SVA. We then evaluated the C-ELISA in terms of sensitivity, specificity, repeatability, and result agreement rate with the VNT. The results showed that we successfully prepared an SVA-specific porcine Fab antibody, which showed high affinity for SVA. We named this antibody 1M33Fab and designated it as Bio-1M33Fab after biotin labeling. The assay conditions were optimized as follows: the coating concentration of SVA particles being 1 μg/mL, the working concentration of Bio-1M33Fab being 0.5 μg/mL, the optimal serum dilution of 1:10, and the optimal dilution of enzyme-labeled avidin being 1:30 000. At a percent inhibition (PI) of 47%, the assay demonstrated the highest sensitivity (96.88%) and specificity (100%), with no cross-reactivity observed with the positive sera of major porcine viral diseases. The intra-assay coefficient of variation ranged from 1.12% to 7.34%, while the inter-assay coefficient of variation ranged from 1.10% to 8.97%, indicating good repeatability. In the detection of 224 clinical pig serum samples, C-ELISA and VNT showed a result agreement rate of 93.75%. In conclusion, we successfully develop a C-ELISA method for detecting neutralizing antibodies against SVA by using a porcine-derived Fab antibody, which lays a foundation for the development of detection kits.
Animals ; Swine ; Antibodies, Neutralizing/immunology* ; Enzyme-Linked Immunosorbent Assay/methods* ; Immunoglobulin Fab Fragments/immunology* ; Antibodies, Viral/immunology* ; Picornaviridae/immunology* ; Humans ; HEK293 Cells ; Swine Diseases/diagnosis* ; Picornaviridae Infections/diagnosis*

Objective To explore the application value of artificial intelligence in medical research assistance, and analyze the key paths to achieve precise execution of model instructions, improvement of model interpretation completeness, and control of hallucinations. Methods Taking esophageal cancer research as the scenario, five types of literature including research articles, case reports, reviews, editorials, and guidelines were selected for model interpretation tests. The model performance was systematically evaluated from five dimensions: recognition accuracy, format accuracy, instruction execution accuracy, content reliability rate, and content completeness index. The performance differences of Ruibin Agent, GPT-4o, Claude 3.7 Sonnet, DeepSeek V3, and DouBao-pro models in medical literature interpretation tasks were compared. Results A total of 15 studies were included, with 3 studies of each type. The five models collectively conducted 1 875 tests. Due to the poor recognition accuracy of the editorial type, the overall recognition accuracy of Ruibin Agent was significantly lower than other models (92.0% vs. 100.0%, P＜0.001). In terms of format accuracy, Ruibin Agent was significantly better than Claude 3.7 Sonnet (98.7% vs. 92.0%, P=0.002) and GPT-4o (98.7% vs. 78.9%, P＜0.001). In terms of instruction execution accuracy, Ruibin Agent was better than GPT-4o (97.3% vs. 80.0%, P＜0.001). In terms of content reliability rate, Ruibin Agent was significantly lower than Claude 3.7 Sonnet (84.0% vs. 92.0%, P=0.010) and DeepSeek V3 (84.0% vs. 94.7%, P＜0.001). In terms of content completeness index, the median scores of Ruibin Agent, GPT-4o, Claude 3.7 Sonnet, DeepSeek V3, and DouBao-pro were 0.71, 0.60, 0.85, 0.74, and 0.77, respectively. Conclusion Ruibin Agent has significant advantages in terms of formatted interpretation of medical literature and instruction execution accuracy. In the future, it is necessary to focus on optimizing the recognition ability of editorial types, strengthening the coverage ability of core elements of various types of literature to improve interpretation completeness, and improving content reliability through optimizing the confidence mechanism to ensure the rigor of medical literature interpretation.

Porcine reproductive and respiratory syndrome (PRRS) is a highly contagious disease caused by porcine reproductive and respiratory syndrome virus (PRRSV), which causes great economic losses. At the moment, no effective neutralizing antibody is available for scientific research and treatment. Therefore, developing a method for screening the neutralizing monoclonal antibodies is of great significance for the prevention and treatment of PRRSV and the screening of antigen sites. Monoclonal antibodies have been widely used in the treatment and diagnosis of many human and animal diseases. Therefore, screening effective neutralizing antibodies for different pathogens is an urgent task. Among the methods for monoclonal antibody screening, B cell immortalization is an effective method to obtain neutralizing monoclonal antibody. Specifically, in this study, the bcl-6 and bcl-xl genes were connected by f2a and then the yielded product was ligated to a vector for retrovirus packaging. The swine lymphocytes immunized with PRRSV were infected the yielded mature viruses and cultured in the complete medium containing CD40L and IL21 cytokines. Then, CD21 was used as the marker to screen B cells with the magnetic bead method. Finally, monoclonal B cells were obtained and the secretion of antibodies was tested. The results showed that the plasmid, either being transfected alone or with the packaged plasmids, could be expressed, and that the packaged retrovirus could infect the cells. Moreover, the infected lymphocytes secreted antibodies, so did the screened B cells. Therefore, the method for screening monoclonal antibody against PRRSV was successfully established.
Animals ; Antibodies, Monoclonal ; Antibodies, Neutralizing ; Antibodies, Viral ; Humans ; Porcine Reproductive and Respiratory Syndrome/prevention & control* ; Porcine respiratory and reproductive syndrome virus/genetics* ; Swine

Antigenic purity is important for quality control of the foot-and-mouth (FMD) whole virus inactivated vaccine. The recommended method for evaluation the antigenic purity of FMD vaccine is to check the serum conversion to non-structural protein (NSP) 3AB antibody after 2 to 3 times inoculation of animals with inactivated vaccine. In this study, we developed a quantitative ELISA to detect the amount of residual 3AB in vaccine antigen, to provide a reference to evaluate the antigenic purity of FMD vaccine. Monoclonal antibody (Mab) of NSP 3A and HRP-conjugated Mab of NSP 3B were used to establish a sandwich ELISA to quantify the NSP 3AB in vaccine antigen of FMD. Purified NSP 3AB expressed in Escherichia coli was serially diluted and detected to draw the standard curve. The detectable limit was determined to be the lowest concentration of standard where the ratio of its OD value to OD blank well was not less than 2.0. Results: The OD value was linearly corelated with the concentration of 3AB protein within the range between 4.7 and 600 ng/mL. The correlation coefficient R² is greater than 0.99, and the lowest detectable limit is 4.7 ng/mL. The amount of 3AB protein in non-purified inactivated virus antigen was detected between 9.3 and 200 ng/mL depending on the 12 different virus strains, whereas the amount of 3AB in purified virus antigen was below the lowest detectable limit. The amount of 3AB in 9 batches of commercial FMD vaccine antigens was between 9.0 and 74 ng/mL, whereas it was below the detectable limit in other 24 batches of commercial vaccine antigens. Conclusion: the sandwich ELISA established in this study is specific and sensitive to detect the content of 3AB protein in vaccine antigen of FMD, which will be a useful method for evaluation of the antigenic purity and quality control of FMD inactivated vaccine.
Animals ; Antibodies, Viral ; Enzyme-Linked Immunosorbent Assay ; Foot-and-Mouth Disease/prevention & control* ; Foot-and-Mouth Disease Virus ; Viral Nonstructural Proteins/genetics* ; Viral Vaccines

Objective To evaluate the relationship between low vitamin D level and metabolic syndrome (MS) in maintenance hemodialysis (MHD) patients.Methods A total of 143 patients who had received MHD from Jan 2016 to Jan 2017 in the dialysis center of our hospital were enrolled.Their clinical and laboratory data were collected.The serum 25(OH)D3 levels were measured by chemiluminescence instrument.According to the levels of 25(OH)D3,patients were divided into three groups:sufficient group (＞ 30 μg/L),insufficient group (15-30 μg/L) and deficient group (＜ 15 μg/L) to explore how the 25(OH)D3 were associated with MS and abnormal metabolic parameters,including central obesity,raised triglycerides (TG),reduced high-density lipoprotein cholesterol (HDL-C),raised systolic blood pressure (SBP),raised diastolic blood pressure (DBP) and increased fasting blood glucose (FBG).The risk factors of MS and abnormal metabolic factors were analyzed by multivariate logistic regression model.Results Among the 143 MHD patients,the median of serum 25(OH)D3 was 24.30(12.90,29.50) μg/L and the prevalence of MS was 45.45％(65 cases).Among 3 groups the prevalence of MS,the abdominal circumference and the serum TG showed statistical differences,and they increased with the severity of 25(OH)D3 deficiency (all P ＜ 0.05).The body mass indexes of patients in the insufficient and deficient groups were elevated compared with that in the sufficient group (all P ＜ 0.05).SBP,TG and FBG in deficient group were significantly higher but HDL-C was lower than those in the other two groups (all P ＜ 0.05).The more abnormal metabolism existed,the lower 25(OH)D3 levels patients had (H=61.316,P＜0.001).Multivariate logistic regression analysis showed that in MHD patients low 25(OH)D3 negatively correlated with MS (OR=0.889,95％CI 0.846-0.934,P ＜ 0.001) and abnormal metabolic factors central obesity (OR=0.913,95％CI 0.874-0.953,P ＜ 0.001),raised TG (OR=0.932,95％ CI 0.894-0.971,P=0.001),reduced HDL-C (OR=0.901,95％ CI 0.845-0.959,P=0.001),raised SBP (OR=0.898,95％CI 0.847-0.953,P＜ 0.001) and raised FBG (OR=0.956,95％CI 0.920-0.994,P=0.024).Conclusions The prevalence of MS is high in MHD patients and low levels of 25(OH)D3 may be an independent risk factor for MS and abnormal metabolic factors.

Objective:To study the relationship between gingival thickness(GT)and the underlying alveolar bone thickness(BT)in maxillary anterior region and the distance from cemento-enamel junction(CEJ)to alveolar crest.Methods:30 young volunteers with healthy gingiva were included.GT was measured at 2mm below the CEJ,buccal BT were measured at 3 locations:2,4 and 6 mm below the alveolar crest respectively,the distance from CEJ to alveolar crest were measured by CBCT and clinical direct measure respectively. Results:The correlation coefficient (r)values between GT and BT at 2,4 and 6 mm below alveolar crest were 0.493,0.383 and 0.342 (P <0.001 )respectively,the r value between GT and the distance from CEJ to alveolar crest was -0.21 3(P <0.01 ).No statistically significant difference was observed between CBCT and clinical measurements(t =-0.521 ,P =0.603).Conclusion:There is positive correlation between GT and BT at 2,4 and 6 mm below alveolar crest and negative relation between GT and the distance from CEJ to alveolar crest.

We investigated the enhanced immune response of a recombinant T cell immunogen as an effective cellular immune adjuvant. The T cell immunogen named TI contained several T cell epitopes from the VP1, VP4, 3A and 3D proteins of foot-and-mouth disease virus (FMDV) and two pan-T helper (T(H)) cell sites to broaden the immunogenicity of the protein. Meanwhile, another fusion protein named OA-VP1 was expressed in bacteria, which contained two VP1 proteins of O and Asia1 type FMDV. Mice were vaccinated with commercially inactivated vaccine or OA-VP1 protein with or without the TI immunogen. The results show that mice inoculated with inactivated vaccine or OA-VP1 protein supplemented with TI immunogen produced significantly higher level of neutralizing antibodies (P < 0.01 or P < 0.05) than the mice only inoculated with inactivated vaccine or OA-VP1 protein by microneutralization assay. An obvious increase in T cell number by flow cytometric analysis and significantly higher concentration of IFN-gamma secreted in culture media of spleen lymphocytes were observed in groups supplemented with TI immunogen (P < 0.01). TI immunogen was an effective stimulator for humoral and cellular immunity and could help improve the immunogenicity of inactivated vaccine or protein subunit vaccine.
Adjuvants, Immunologic ; pharmacology ; Animals ; Capsid Proteins ; genetics ; immunology ; Epitopes, T-Lymphocyte ; genetics ; immunology ; Foot-and-Mouth Disease ; immunology ; prevention & control ; virology ; Foot-and-Mouth Disease Virus ; immunology ; Immunization ; Mice ; Viral Vaccines ; genetics ; immunology ; pharmacology

Objective To investigate the effect of reduced glutathione(rGSH)on the retina and optic nerve in rats with experimental optic neuritis,and the underlying mechanism.Methods Animal model of experimental allergic encephalomyelitis(EAE),which was as used for acute demyelinating optic neuritis were established in 48 female SD rats,and then the rats were randomly divided into EAE group(n=18)and GSH group(n=30).Another 6 female SD rats served as normal control.Rats of GSH group received an intraperitoneal injection of 100 mg?kg-1?d-1 GSH for 14 d from the beginning of immunization,while the rats of EAE group were given saline alternatively.To observe pathological changes in the optic nerve,retinal ganglion cells(RGCs)apoptosis,and measure malondialdehyde(MDA),glutathione peroxidase(GSH-PX),nitric oxide(NO)levels in the retina were observed and measured in the early period,peak period and recovery period(12 to 14 d,16 to 18 d,or 26 to 28 d after the immunization)of the neuritis.Results There were 18 rats(60%)from GSH group having the disease with a symptom score of 1 to 3.Compared with those of EAE group,their optic nerve fibers were in a comparatively good order,and more glial cells and not obvious demyelination were observed in morphology;Significant lesser apoptotic RGCs were observed in GSH group than in EAE group at the early and peak period(P

Objective To investigate the combination of compound anisodine and physical therapy in the treatment of amblyopia,and its possible mechanism.Methods Totally 300 outpatients with amblyopia(3 to 14 years old) were randomly and equally divided into treatment group and control group.In the treatment group,subcutaneous injection of compound anisodine(2 ml,once per day) to the superficial temporal artery of eye was given for a course of 14 d and followed by another course after 5 days'interval.Cover treatment was carried out at the same time.The control group was only treated with physical therapy.Vision,central retinal artery peak systolic velocity(PSV) and diastolic resistance index(RI) were measured during the 3 months' follow-up.Results The 3 to 6-year-old efficiency was 75.0%,7 to 9-year-old efficiency was 69.6%,and 10 to 14-year-old efficiency was 61.1% ;The difference of efficiency between therapy group and control group was very significant(P0.05).Conclusion Compound anisodine plus physical therapy for amblyopia at different ages and varying degrees are effective and safe.The mechanism may be due to enhanced retinal blood supply.

TLC method for the identification of triterpenoids polyphenols in Fructus Chebulae was established .The results showed that this method had a plentiful chromatogram and good specificity,and can distinguish Fructus Chebulae immaturus from Fructus Chebulae.Determination of gallic acid, one of the hydrolysates of Fructus Chebulae, by HPLC was also reported.The linear range of gallic acid is from 0.17?g to 1.36?g(r=0.9998),and the average recovery is 102.9 %. The results showed that the method is simple, accurate and with good reproducibility.It can be used to determine gallic acid in Chinese herbal medicine that contains hydrolysable tannins.