As a protein expression vector,the baculovirus demonstrates many advantages over other vectors.With the development of biotechnology,baculoviral vectors have been genetically modified to facilitate high level expression of heterologous proteins in both insect and mammalian cells.These modifications include utilization of different promoters and signal peptides,deletion or replacement of viral genes for increasing protein secretion,integration of polycistronic expression cassette for producing protein complexes,and baculovirus pseudotyping,promoter accommodation or surface display for enhancing mammalian cell targeting gene delivery.This review summarizes the development and the current state of art of the baculovirus expression system.Further development of baculovirus expression systems will make them even more feasible and accessible for advanced applications.

[ ABSTRACT] AIM: To investigate the role of TNF-αand NF-κB in the mechanism of neuropathic pain due to chronically compressed dorsal root ganglion ( CCD) .METHODS:Based on the CCD model, von Frey filaments were used to quantify behavior test.The expression changes of TNF-αand NF-κB were determined by Western blotting, and the cor-relation between the expression of TNF-αand the 50%paw withdrawal threshold was also analyzed.Moreover, the location of TNF-αin dorsal root ganglion ( DRG) was observed with immunofluorescence double staining.RESULTS: We found 50%paw withdrawal threshold of CCD decreased at the first day after operation.The mechanical allodynia was the most ob-vious at postoperative 7~14 d and lasted longer than 35 d.The expression of TNF-αand NF-κB increased significantly in DRG after operation (P<0.01), especially at 7~14 d, and then restored gradually.Moreover, there was a correlation between the protein expression of TNF-αand the changes of neuropathic behavior ( P<0.05 ) .CONCLUSION: TNF-αand NF-κB are involved in the mechanism of mechanical allodynia after chronically compressed DRG.

BACKGROUND: There are differences in physical and biological activity between the antibody from mammals and egg yolk antibody (IgY) from chicken. IgY is acid- and heat-resistant, and can prevent and cure the infectious diseases in animals and human being, which is also benefit to develop routine diagnostic immunoassays. Conventional ELISA assay for IgY takes much more time than dot-immunobinding assay.OBJECTIVE: To detect the IgY stability byusing dot-immunobinding assay.DESIGN: Open trail.SETTING: Department of Transfusion, Kunming General Hospital of Chinese PLA.MATERTALS: The experiment was completed in the Kunming General Hospital of Chengdu Military Area Command of Chinese PLA from January to June 2006. Two White Leghorn hens (30 weeks old) were selected. HLA-A*0201 α chain served as the antigen. The total protein concentration of the purified antigen was 0.04 g/L with the molecular mass of 32 000(self-prepared); nitrocellulose filter (NC, import and divided); nonfat dry milk (Anyi Corp. No. 20051220); DAB (Boshide Corp.);caprylic acid (made by Shanghai Xinghuo Chemical Factory); ammonium sulfate (Shantou Guanghua Chemical product).METHODS: ①HLA-A*0201 α chain with the total protein concentration of 0.04 g/L was purified with egg yolk antibody,and identified by SDS-PAGE. ②1 μL antigen was spotted into the center of NC membrane and dried in the incubator at 37 ℃. Then the NC membrane was blocked in 1 mL PBST and put in the incubator at 37℃ with shaking in 90 r per minute for 15 minutes. Then the liquid was exchanged with 1 mL PBST and added the primary antibody at a final concentration of 10 mg/L. After 30 minutes shaking in the incubator at 37 ℃, the NC membrane was washed in PBST for three times. The second antibody, mouse anti-chicken IgY conjugated to horseradish peroxidase (HRP) was added and after 30 minutes incubation, the NC membrane was washed three times in PBST. Binding was revealed by incubation with a DAB reagent. A positive reaction was represented by adeep brown spot,Irdlcating that IgY had better activity; if the spot became lighter IgY lost part activity, and when the spot disappeared, the IgY lost a the activty.According to intensity (gray degree)of the dot compared tothe standard, the remained percent of activity of the IgY was calculated. ③IgY was adjusted to three different protein concentrations with PBS: 1, 0.1, 0.01 g/L and stayed at room temperature for four months. 10 μg lgY was taken out from each concentration sample every month to detect the activity by dot-immunobinding assay. ④IgY was put into seven EP tubes with 100 μL per tube and numbered 1-7. Number 1 to 3 was adjusted pH to 5, 3 and 2, respectively with 1 mol/L HCI; Number 4 to 6 was to 9, 11 and 12, respectively with 1 mol/L NaOH. The pH of number 7 was neutral without adding acid or base. The samples were stayed in incubator at 37 ℃ for 3 hours. 10 μg IgY from each tube was taken every hour to detect the stability at different pH by dot-immunobinding assay. ⑤IgY was added to six EP tubes (10 μL per tube) and numbered 1-6. Number 1-6 was put into waterbath at 30, 40, 50, 60, 70 and 80 ℃ for 15 minutes. After cooled in refrigerator at 4 ℃, 10 mg samples from each tube and standard sample (untreated sample) taken to check the thermal stability by dot-immunobinding assay.MAIN OUTCOME MEASURES: ①SDS-PAGE of IgY. ②IgY stability at room temperature. ③IgY stability at different pH. ④ Detection of IgY thermal stability.RESULTS: ①Purified IgY after SDS-PAGE had two major binds, the molecular mass of the heavy chain was 66.000,and the light chain was 25 000. ②1, 0.1, 0.01 g/L IgY still had partial activity after staying at room temperature for four months. ③When pH ranged from 5 to 9, IgY still had partial activity after staying in 37 ℃ for 3 hours. If pH was lower than 5 or higher than 9, it lost the whole activity in above condition. ④Purified IgY was added to six EP tubes, the number 1-4 still had partial activity, but number 5 and 6 showed some white precipitate, which was caused by protein denaturation at higher temperature.CONCLUSION: IgY stability is higher than others. The dot-immunobinding assay described a rapid and simple method for the demonstration and characterization of functional activity of egg yolk antibody. With only small volume antigen and antibody, and specific dot, the dot-immunobinding assay method could process many samples at the same time.

Objective To investigate the effect of Roy Adaptation Model(RAM)in patients with adolescent insanity.Methods One hundred patients with adolescent insanity during October 2011 to March 2012 were randomized in equal number into two groups by random digit table:the study group and the control group.The former were intervened with RAM and the latter received routine care and health education.Seven weeks after intervention,Hamilton Depression Rating Scale(HAMD),Hamilton Anxiety Scale(HAMA) and Observation Scale(NOSIE)were used for the assessment. Results After intervention,the scores on HAMD and HAMA in the study group were significantly lower,compared to the control group(P<0.01).The scores on social function,social interest,subjective support and use of social support were all significantly higher than those of the control group(all P<0.01). Conclusions RAM can improve their ability of the patients with adolescent insanity to adapt to the environment.It may improve their mental state and their quality of life.

Objective:To analyze the effect of intestinal fluid reinfusion after anterior resection of rectum and preventive ileostomy in patients with low rectal cancer.Methods:A prospective research method was used, and 60 patients with low rectal cancer in Zhejiang Jinhua Guangfu Tumor Hospital from January 2021 to June 2022 were enrolled. The patients were divided into control group (treated with anterior resection of rectum and preventive ileostomy) and observation group (treated with anterior resection of rectum and preventive ileostomy combined with intestinal fluid infusion) by random number table method with 30 cases each. On the next day and 3 months after surgery, the low anterior resection syndrome (LARS) scale was used to evaluate anal function, the patient-generated subjective global assessment (PG-SGA) method was used to evaluate nutritional status, the stoma-quality of life (stoma-QOL) was used to evaluate the quality of life. The complications were recorded.Results:Both groups of patients were successfully completed the scheduled surgery. There were no statistical differences in LARS score and incidence of LARS on the next day after surgery between the two groups ( P>0.05); the LARS score and incidence of LARS 3 months after surgery in observation group were significantly lower than those in control group: (18.63 ± 3.15) scores vs. (24.90 ± 6.11) scores and 23.33% (7/30) vs. 46.67% (14/30), and there were statistical differences ( P<0.01 or <0.05). There were no statistical differences in PG-SGA score and incidence of malnutrition on the next day after surgery between the two groups ( P>0.05); the PG-SGA score and incidence of malnutrition 3 months after surgery in observation group were significantly lower than those in control group: (3.07 ± 0.82) scores vs. (5.13 ± 1.01) scores and 26.67% (8/30) vs. 46.67% (14/30), and there were statistical differences ( P<0.01 or <0.05). There were no statistical differences in the scores of psychological burden, social interaction, stoma management and daily living of stoma-QOL on the next day after surgery between the two groups ( P>0.05); the scores 3 months after surgery in observation group were significantly higher than those in control group: (27.70 ± 4.28) scores vs. (21.47 ± 5.16) scores, (14.33 ± 2.03) scores vs. (11.90 ± 1.64) scores, (14.87 ± 1.92) scores vs. (11.57 ± 2.38) scores and (15.30 ± 1.03) scores vs. (12.37 ± 2.11) scores, and there were statistical differences ( P<0.01). The incidence of complications in observation group was significantly lower than that in control group: 30.00% (9/30) vs. 60.00% (18/30), and there was statistical difference ( χ2 = 5.45, P<0.05). Conclusions:For the patients with low rectal cancer, the anterior resection of rectum and preventive ileostomy combined with intestinal fluid reinfusion can reduce LARS, improve nutritional status, improve quality of life, and also reduce complications.