Objective: Overexpression of the HER2/neu oncogene is a frequent molecular event in multiple human cancers. Being a cancer antigen, p185 erbB2 is an ideal target for immunotherapy. In order to decrease the immunogenicity of mouse anti-p185 erbB2 monoclonal antibody in human cancer therapy, we constructed the eukaryotic expression vector of anti-p185 erbB2 chimeric monoclonal antibody and verified expression of the chimeric antibody in CHO-dhfr - cell. Methods: The variable regions of light chain and heavy chain were amplified with RT-PCR and inserted into the chimeric antibody vector pWSD2. After CHO-dhfr - cells were transfected with recombination plasmid by lipofectAMINE, the chimeric antibody expressing level was identified with RT-PCR, indirect-ELISA, and Western blot. The specificity of the anti-p185 erbB2 chimeric antibody was testified with ELISA assay and immunoprecipitation. Moreover, the effects of chimeric antibody on the proliferation of breast cancer cell line SKBR3, which is overexpressing p185 erbB2 , were measured with MTT assay in vitro. Results: The anti-p185 erbB2 chimeric antibody eukaryotic expression vector was constructed successfully and the expression of the chimeric antibody in CHO-dhfr - was verified by RT-PCR, indirect-ELISA, and Western blot. ELISA assay showed that chimeric antibody reacted with cells overexpressing p185 erbB2 specifically, but did not react with that non-overexpressing p185 erbB2 . Immunoprecipitation test confirmed that the chimeric antibody could bind to p185 erbB2 specifically. The MTT assay demonstrated that the chimeric antibody could inhibit the growth of SKBR3 cells overexpressing p185 erbB2 . Conclusion: The anti-p185 erbB2 mouse/human chimeric antibody that was expressed in CHO-dhfr - cells can bind to p185 erbB2 specifically and inhibit proliferation of SKBR3 cells overexpressing p185 erbB2 . It has a potential application in biotherapy of cancer.

The clinical data of 4 patients with peripheral vascular rupture admitted in the Hetian Regional People′s Hospital from March 2012 to October 2017 were retrospectively analyzed. The 4 patients were all males, aged 17-59 years. Among them, 2 cases caused by firearm injury, 1 case was caused by a car accident and 1 case was caused by a sharp injury; 2 cases were incompletely ruptured in femoral artery, 1 case was completely ruptured in popliteal artery, and 1 case was completely ruptured in axillary artery. All patients underwent successful reconstructive surgical procedures, and the skin temperature, skin color, and arterial pulse of the affected limb returned to normal after operation. The followed-up time was 28-95 months, with an average of 47.8 months. Blood flow in the reconstructed vessel was unobstructed. There were no complications. It is indicates that for peripheral vascular rupture if the diagnose and treatment are timely, the satisfactory results can be achieved.

Hsc70-SNCG fusion protein cDNA fragment containing signal peptide sequence of Igkappa, MMP9, and P37 was inserted into the vector pVAX1 to construct recombinant plasmid pVAX-Igkappa-Hsc70-SNCG, pVAX-MMP9-Hsc70-SNCG, and pVAX-P37-Hsc70-SNCG. Three eukaryotic vectors were constructed and verified by restriction enzyme digestion and sequencing. After transfection with recombination plasmids in QM-7 cells, the transient expression and secretion of three fusion proteins were detected by ELISA and Western Blot. The results suggested that Hsc70-SNCG carrying three different signal peptides could be expressed and secreted by transfected cells, and three signal peptides effectively directed secretion of fusion protein by QM-7 cells. BALB/c mice were immunized by three plasmids using gene gun system. The serum levels of anti-SNCG antibodies in mice were measured by ELISA. The results showed that three secreted plasmids could stimulate humoral immune responses to SNCG in mice, which depended on the secreted expression levels induced by signal peptides.
Animals ; Genetic Vectors ; Humans ; Immunoglobulin G ; biosynthesis ; Mice ; Mice, Inbred BALB C ; Neoplasm Proteins ; biosynthesis ; genetics ; immunology ; Plasmids ; metabolism ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; immunology ; Transfection ; gamma-Synuclein ; biosynthesis ; genetics ; immunology

Mycobacterium tuberculosis is the causative agent of tuberculosis(TB),which is still the leading cause of mortality from a single infectious disease worldwide.The development of novel anti-TB drugs and vaccines is severely hampered by the complicated and time-consuming genetic manipulation techniques for M.tuberculosis.Here,we harnessed an endogenous type Ⅲ-A CRISPR/Cas10 system of M.tuberculosis for efficient gene editing and RNA interference(RNAi).This simple and easy method only needs to transform a single mini-CRISPR array plasmid,thus avoiding the introduction of exogenous protein and minimizing proteotoxicity.We demonstrated that M.tuberculosis genes can be efficiently and specifically knocked in/out by this system as con-firmed by DNA high-throughput sequencing.This system was further applied to single-and multiple-gene RNAi.Moreover,we successfully performed genome-wide RNAi screening to identify M.tuberculosis genes regulating in vitro and intracellular growth.This system can be extensively used for exploring the functional genomics of M.tuberculosis and facilitate the development of novel anti-TB drugs and vaccines.

Objective: To analyze the etiology and epidemiological characteristics of influenza in Wenzhou from 2009 to 2014, so as to provide the scientific basis for control and prevention of influenza. Methods: Throat swab specimens of influenza like illness (ILI) were collected from national influenza surveillance sentinel hospitals for nucleic acid detection with real-time PCR and virus isolation, culture and sequencing, and the results were analyzed with statistical methods. Results: During the 8 years, a total of 10 577 089 cases from outpatient and emergency department were monitored in sentinel hospitals. There were 337 896 ILI cases with an average ILI treatment rate of 3.19%. A total of 4 046 ILI samples were detected in children, 511 were positive for influenza, the positive rate was 12.63%. Among the detected influenza types, type B had the highest proportion, followed by H3N2. Among the 6 age groups, the number of flu patients was the highest in 0-3 years old group, the positive rate in 10-12 years old group was the highest (35.03%). There were 28 and 45 amino acid sequence mutations of HA fragment in influenza A and B, respectively, which included multiple mutation of 391 and 145 amino acids. The phylogenetic analysis showed that the strains of type B were different in different years, and Yamagata evolved into Y1 and Y2 two branches. Conclusions The prevalence peaks of influenza in children occurred in winter and spring in Wenzhou city, accompanied by small peaks in summer. Three subtypes of serotypes B, H3N2 and A(H1N1) dominated alternatively in Wenzhou during the 8 years. We should focus on strengthening the prevention and control of influenza in preschool children and primary and secondary school students.