Mapping of the B Cell Neutralizing Epitopes on ED III of Envelope Protein from Dengue Virus / 病毒学报

Dengue virus (DENV) envelope [E] protein is the major surface protein of the virions that indued neutralizing antibodies. The domain III of envelope protein (EDIII) is an immunogenic region that holds potential for the development of vaccines; however, the epitopes of DENV EDIII, especially neutralizing B-cell linear epitopes, have not been comprehensively mapped. We mapped neutralizing B-cell linear epitopes on DENV-1 EDIII using 27 monoclonal antibodies against DENV-1 EDIII proteins from mice immunized with the DENV-1 EDIII. Epitope recognition analysis was performed using two set of sequential overlapping peptides (16m and 12m) that spanned the entire EDIII protein from DENV-1, respectively. This strategy identified a DENV-1 type- specific and a group-specific neutralizing epitope, which were highly conserved among isolates of DENV-1 and the four DENV serotypes and located at two regions from DENV-1 E, namely amino acid residues 309-320 and 381-392(aa 309-320 and 381-392), respectively. aa310 -319(310KEVAETQHGT319)was similar among the four DENV serotypes and contact residues on aa 309 -320 from E protein were defined and found that substitution of residues E309 , V312, A313 and V320 in DENV-2, -3, -4 isolates were antigenically silent. We also identified a DENV-1 type-specific strain-restricted neutralizing epitope, which was located at the region from DENV-1 E, namely amino acid residues 329-348 . These novel type- and group-specific B-cell epitopes of DENV EDIII may aid help us elucidate the dengue pathogenesis and accelerate vaccine design.

Expression of SARS-CoV nucleocapsid protein in Bac-to-Bac Baculovirus System and antigenic analysis / 中国免疫学杂志

Objective:To express SARS-CoV nucleocapsid protein in Bac-to-Bac Baculovirus Expression System and analyze the antigenicity of the recombinant protein.Methods: The SARS-CoV nucleocapsid gene was amplified by PCR.The PCR product digested with BamHⅠand SalⅠrestriction endonucleases was cloned into vector pFastBac HTC of Bac-to-Bac Baculovirus expression system.Recombinant plasmid was transformed DH 10Bac cells to obtain the recombinant Bacmid DNA.Recombinant Bacmid DNA was transferred into Sf9 cells which were inducted to express the recombinant protein in High Five cells.After purified by Ni affinity chroma-tography ,the antigenicity of the recombinant protein was analyzed by Western blot and ELISA.Results:Recombinant plasmid was con-structed successfully.The recombinant protein with the relative molecular mass of 48 kD was efficiently expressed in High Five cells and purified successfully by Ni affinity chromatography.Western blot and ELISA analysis showed that the recombinant protein could be spe -cifically recognized by the monoclonal antibody to SARS-CoV N protein and immune serum from rabbits ,respectively.The recombinant protein can specifically reacted with serum from SARS patients ,not with serum from healthy persons and patients infected with hCoV-229 E and hCoV-OC43.Conclusion: SARS-CoV nucleocapsid protein has been expressed successfully in the Bac-to-Bac Baculovirus Expression System ,and obtained good antigenicity.It is preliminary deemed that it can't reacted with serum from patients infected with hCoV-229E and hCoV-OC43.

Analysis of serum neutralizing antibody response in patients with primary dengue virus type 1 infection / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the characteristics and dynamic changes of serum neutralizing antibody response in patients with primary infection of dengue virus type 1 (DENV-1).</p><p><b>METHODS</b>Serum samples were obtained from the same patients with primary infection of DENV-1 within 2 weeks after symptom onset in 2006 and in 2010. A group-specific DENV NS1 capture ELISA-based micro-neutralizing test (ELISA-MNT) capable of detecting neutralizing antibodies against all the 4 serotypes of DENV was used to test the neutralizing antibody titers against DENV in the serum samples. The neutralizing antibody titers against a standard strain and 2 clinically isolated strains of DENV-1 were detected in serum samples collected in 2010.</p><p><b>RESULTS</b>Cross-reactive neutralizing antibody response against all the 4 serotypes of DENV was found in both of the serum samples collected in 2006 and 2010, but the samples collected in 2006 showed stronger cross-reactive neutralizing antibody responses. The neutralizing antibody against DENV-2, rather than the anticipated DENV-1 antibody, had the highest titer in the samples collected in 2006, whereas the antibody against homologous DENV-1 had the highest titer in the samples obtained in 2010. The neutralizing antibody titers against the homologous DENV-1 was significantly higher in samples collected in 2010 (U=86.500, P=0.000), which also demonstrated significantly different neutralizing antibody titers against the 3 different strains of DENV-1 (Χ(2)=12.123, P=0.002).</p><p><b>CONCLUSION</b>The production of cross-reactive neutralizing antibodies between the 4 serotypes of DENV is a characteristic of DENV infection, particularly during early infection, but only the homologous neutralizing antibody increases obviously over time. The titers of the neutralizing antibodies against different strains, even of the same serotype, may differ distinctly.</p>

Effect of lentiviral vector-mediated short hairpin RNA targeting survivin against endometriosis-like lesions in chick embryo chorioallantoic membrane / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the inhibitory effect of lentiviral vector-mediated short hairpin RNA targeting survivin (LV-survivin shRNA) on angiogenesis and growth of endometriosis-like lesions in chick en embryo chorioallantoic membrane.</p><p><b>METHODS</b>Eutopic endometrium from women with endometriosis was transplanted onto the non-vascular region of (CAM), where LV-survivin shRNA was delivered subsequently. The angiogenesis and the growth of endometriosis-like lesions in the CAM model were evaluated.</p><p><b>RESULTS</b>The angiogenesis and formation of endometriosis-like lesions were significantly suppressed in the CAM model by treatment with LV-survivin shRNA in comparison with those in the untreated CAM models and models treated with empty LV or DMEM (P<0.001). LV-survivin shRNA also caused a significantly higher cell apoptotic rate in the endometriosis-like lesions than the other treatments (P<0.001) and induced necrosis in the lesions.</p><p><b>CONCLUSION</b>LV-survivin shRNA can effectively inhibit angiogenesis induced by the eutopic endometrium and markedly suppress the formation of endometriosis-like lesions in the CAM model.</p>

Comparing of the cost of special nursing and application effect between two ways of deep veins catheter in patients with chemotherapy / 中国实用护理杂志

Objective To confirm the deviation between the present charge and real cost of nursing by comparing them between PICC and jugular veins catheter,and the advantages and disadvantages were weighed between them in tumor patients undergoing chemotherapy.Methods The cost of manpower,financial resources or materials for 106 patients with PICC,68 patients with jugular veins catheter were measured,accounted and statistically analyzed with the ladder sharing method for project cost.Comparing the cost between calculation cost and current charging standard,and the cost and clinical application was studied.Results The real cost of PICC was (2259.99±30.99)Yuan and current charging standard was 1532.79 Yuan,the deviation was -727.20 Yuan,and the real cost of jugular veins catheter was (393.86±33.93) Yuan,and current charging standard was 292.13 Yuan,the deviation was -101.73 Yuan.The complication rate was 12％ in PICC,17％ in jugular veins catheter.The real cost of nursing on PICC and jugular veins catheter was higher than current charging standard,and the real cost of single nursing on PICC was 5.74 times higher than jugular veins catheter.The cost remains unchanged with 4～6 course of chemotherapy treatments in a year.Conclusions The advantages of PICC are more than jugular veins catheter,so the preferred choice is PICC,and jugular veins catheter comes secondary.

Identification and priliminary application of monoclonal antibodies against inrisive Aspergillus / 中华检验医学杂志

Objective To screen monoclonal antibodies (mAbs) for early diagnosis of invisive Aspergillus. Methods Monoclonal antibodies against different antigens of Aspergillus fumigatus were produced. The two pairs of combinations of monoclonal antibodies were selected accoring the distinct epitopes and double-antibody sandwich ELISA based on mAbs above were established. The sensitivity and specificity of the methods were analyzed by detecting culture supernatants of clinical isolates and environmental isolatesof Aspergillus. spp, Penicillium Marneffei, Candidas, and serum from animal models and patients. The epitopes recognized by mAbs were identified by immunobotting. Results A total of 32 hybridoma cell lines that stably produced MAbs were obtained. Two double- antibody sandwich ELISAs were established. One method was specific for 19 clinical isolates and environmental isolates of Aspergillus. spp, whereas the other one was specific for the clinical and environmental isolates of Aspergillus fumigatus without cross-reation with other Aspergillus. spp. For the same kind of medium of Aspergillus fumigatus, the sensitivity of the first method was 10 fold higher than the second method. Conclusions The specific mAbs for early diagnosis of invisive Aspergillus were obtained. Antigen recognized by the specific mAbs was mannoprotein with molecular weights of approximately 25 000-75 000. This antigen was potential early diagnostic marker for invasive Aspergillus.

Identification and application of monoclonal antibodies against aspergillus fumigatus / 中国免疫学杂志

Objective:To produce monoclonal antibodies(McAb) against aspergillus fumigatus and to establish rapid assay for the measurement of aspergillus fumigatus antigen.Methods:Recombinant galactomannoprotein of aspergillus fumigatus(AFMP1) was used to immune BALB/c mice.Monoclonal antibodies against AFMP1 were produced from hybridoma.Results:Three hybridomas producing antibodies against AFMP1 were obtained.IgG isotypes of three McAb were IgG1,IgG2a and IgG2b.The affinity constants(K) were 1.2?10 10 ,4.56?10 9 and 1.81?10 10 mol/L.The antibodies were proved to be specific for aspergillus fumigatus by Western blot and recognized different epitopes on AFMP1 by the additivity assay.An sandwich enzyme-linked immunosorbent assay(ELISA) to detect AFMP1 was established to produce standard curve which showed linearity between 0.1～60.0 ng/ml with a sensitivity of 0.1 ng/ml.Conclusion:These results show three hybridomas producing high specificity and affinity monoclonal antibodies against AFMP1 and can provide for rapid assay for the measurement of aspergillus fumigatus antigen.