Balb/c mice were immunized by intraperitoneal iniection of group A erythrocytes. Immunized spleen cells were fused with Sp2/0 murine myeloma cells from the solid tumour. After 10～15 days of incubation at 37℃ CO_2, the supernatants were screened for the agglutinabal antibodies. Three hybridoma cell lines secreting high titer and specific monoclonal anti-A antibodies were established.These hybridoma cells have the ability of constant seceting antibodies which belong to the IgM class. The specificity monocloal anti-A sera was the same as the human anti-A sera.

0.05), which were lower than that with manual compression. The failure rates were 8.3% and 9.2% respectively, but the vascular complications were less than 1%. Hematoma and femoral artery infection were seen in the PCI group, but happened less compared with manual compression. The removing of the arterial sheaths will not be limited by the anticoagulation and antiplatelet therapies, and it could be performed immediately after CAG and PCI, making hemostasis more easy to archeive. It could also reduce the burden of the medical staff, and be accepted by the patients. Conclusion Femoral arterial closure following PCI using regular and intensive anticoagulation and antiplatelet therapies could be safe and effective with vascular complication rates similar to or lower than with manual pressure.

Objective To fractionate phosphoproteome of mouse liver by two-dimensional (2D) liquid phase chromatography fractionation. Methods Phosphoproteins were extracted from lysates of normal mice livers by phosphate metal affinity chromatography (PMAC) resin. The phosphoproteins were exchanged by start buffer and separated by chromatofocusing in the first dimension. Then the fractions between pH 8.5 and pH 4.0 were separated by non-porous silica (NPS) reverse-phase high performance liquid chromatography (RP-HPLC). Finally, the UV maps were converted into gel-like maps by ProteoVue software. Results Phosphoproteins of mouse liver were successfully extracted and fractionated by two dimensional liquid phase chromatographic fractionation after concentration and desalt. Then pI/UV map of mouse liver phosphoproteome was successfully set-up. There are 16 fractions between pH 8.5 and pH 4.0 after chromatofocusing in the first dimension and the UV maps of each fraction were converted into pI/UV gel-like maps. Conclusions Combination of technique of phosphoproteins enrichment and 2-D liquid phase chromatographic fractionation is an effective approach to research phosphoproteome and the key base for further identification and investigation of phosphoproteins.

The expression of microRNA (miRNA) is closely related to radio-chemosensitivity in glioma stem cells (GSCs).Moreover,the growth of glioma stem cells could be inhibited comprehensively by increasing radio-chemosensitivity and apoptosis,simultaneously with the regulation of a single miRNA,which has been confirmed by some researches.Thereby microRNA is prospective for the adoption as a specific agent in targeted therapy of glioma,so as to increase the radio-chemosensitivity in glioma stem cells.

Objective: To investigate whether FasL expression by exogenous FasL gene transfer had regulatory effect on drug sensitivity of tumor cells. Methods: Exogenous Fasl, gene transfer was mediated by lipofectAMINE. The FasL expression was detected bv Immunohistochemistry and RT-PCR. The growth inhibition of cytotoxic drugs on tumor cells was measured by MTT assay. Fas mRNA expression was analyzed bv Semi-quantitative RT-PCR. Results: The eukaryotic expression vector, PcDNA3-FasL and PcDNA3 were successfully transteeted into MCF-7 cells. The transfectant cells was named as MCF-7/FL or MCF-7/Vt respectively. FasL was expressed on MCF-7/FL but not on MCF-7/Vt or MCF-7. The cytolysis ot MCF-7/FL induced by adriamycin at concentration of 0.625 ~ 2.5 ?g/ml, cisplatin of 1 .25 ~ 10 ?g/ml, or methotrexate of 3 ~ 6 ?g/ml respectively were all significantly enhanced compared with the cytolysis of MCF-7. Blocking FasL by using F(ab' )2-anti APO-1 antibody markedly reduced this up-regulated cytolysis induced by these drugs The significant up-regulation of Fas mRNA on MCF-7/FL was observed by Semi-quantitative RT-PCR after these drugs treatment. Conclusion: These findings indicated that FasL expression by gene transfer had up-regulation effect on drug sensitivity of tumor cells, this effect was associated with enhanced Fas expression induced by cytotoxic drugs.

This article introduces a new educational model-Ability Standard Teaching Mode,which emphasizes that students should participate in teaching and promotes the students to change their thinking style from memory and imitation style to thought and creativity style.

Objective To construct the vector that expresses the fusion protein of HIV-Tat protein and red fluorescent protein(mCherry) in mammalian cells,and observe by fluorescence microscopy the intracellular transduction and localization of recombinant protein in cells,in order to obtain a useful tool for the study of the uptake mechanism and intracellular localization of HIV-TAT.Methods With the designed primer coding mCherry sequence,the mCherry gene was amplified by PCR with the vector pmCherry-C2 as template,and inserted into vector pET14b-His-TAT to construct the expression vector pET14b-His-TAT-mCherry.The constructed vector was then transformed into E.coli BL21(DE3),which had been identified by PCR and double digested with restriction endonuclease,followed by sequencing.After IPTG induction,the recombinant protein of His-TAT-mCherry was lyzed and analyzed with SDS-PAGE.Purified His-TAT-mCherry recombinant protein was added to Hela cells and the fluorescence was observed to evaluate the transduction efficiency.Results The results of identification by PCR,digestion with restriction endonuclease and sequencing indicated that the vector His-TAT-mCherry was correctly constructed.His-TAT-mCherry fusion protein was expressed in mammalian Hela cell line and purified successfully,and the fusion protein showed cellular transduction activity.It was found by fluorescence microscopy that the red fluorescence protein located mainly over the cytoplasm,and also the membrane to some extent.Conclusion The expression vector is successfully constructed for HIV-TAT labeled with mCherry sequence.Effective expression and purification of this fusion protein is achieved.It has been observed that the constructed vector may be expressed in mammalian Hela cell under active condition.Thus,it might be useful in the study of uptake mechanism and intracellular localization of HIV-TAT.

In order to study the role of cofactor engineering in enhancing the production of S-adenosylmethionine (SAM), we altered the form and concentration of cofactor in Saccharomyces cerevisiae through gene recombination. Effects of cofactor on product synthesis, carbon and energy metabolism were analyzed aiming to provide a theoretical basis for a successful metabolic engineering of SAM producing strains. Because NADPH metabolism in mitochondrion and cytoplasm of S. cerevisiae is relatively independent, the effect of intracellular NADPH availability on the production of SAM was studied in different compartments of S. cerevisiae BY4741. The expression of NADH kinase in mitochondria (POS5 encoded) and cytoplasm (POS5Δ17 encoded) was separately confirmed using a laser scanning confocal microscope. NADPH regulation strategy enhanced SAM production. Compared with the control strain, the intracellular SAM concentration of strain NBYSM-1 was increased by 3.28 times, and the intracellular SAM concentration of strain NBYSM-2 was increased by 1.79 times at 24 h fermentation. In addition, SAM titer and NADPH/NADP⁺ ratio in strain NBYSM-1 were significantly higher than that of strain NBYSM-2. Therefore, NADPH regulation strategy will be a valuable tool for SAM production and could further improve the synthesis of a large range of cofactor-driven chemicals.

Objective To analyze the metastatic rule of common hepatic artery lymph node of thoracic esophageal squamous cell carcinoma,and to investigate the strategies of common hepatic artery lymph node dissection.Methods The clinical data of 682 patients with esophageal squamous cell carcinoma who were admitted to the Cancer Hospital of Fudan University from May 2005 to December 2010 were retrospectively analyzed.The locoregional lymph node metastasis of thoracic esophageal squamous cell carcinoma,relationship between metastatic rates of common hepatic artery lymph node and clinicopathological factors and the postoperative complications were analyzed.The enumeration data were analyzed using the chi-square test.Results A total of 18 277 lymph nodes were dissected (27 lymph nodes per patient).The lymph node metastatic rate was 55.87％ (381/682),and the metastatic lymph node ratio was 7.87％ (1438/18 277).Lymph nodes adjacent to the cardia of stomach,laryngeal nerve,lesser curvature of stomach,cervical esophagus,left gastric artery had a higher metastatic rate,while common hepatic artery lymph node had a lower metastatic rate.All the common hepatic artery lymph node metastasis was accompanied with locoregional metastasis.A total of 1480 common hepatic artery lymph nodes were dissected (2 common hepatic artery lymph nodes per patient).Twenty-four patients had common hepatic artery lymph node metastasis,with the metastatic rate of 3.52％ (24/682) and the lymph node ratio of 2.16％ (32/1480).The common hepatic artery lymph node metastatic rates of upper,middle and lower esophageal squamous cell carcinoma were 2.33％ (1/43),3.76％ (16/425) and 3.27％ (7/217),with no significant difference (x2 =0.295,P ＞ 0.05).The common hepatic artery lymph node metastatic rates of patients in T1,T2 and T3 stages were 2.35％ (2/85),5.46％ (10/183) and 2.90％ (12/414),with no significant difference (x2 =2.850,P ＞ 0.05).The common hepatic artery lymph node metastatic rates of patients with high,moderate and poor differentiated esophageal squamous cell carcinoma were 0(0/63),3.50％ (16/457) and 4.94％ (8/162),with no significant difference (x2=3.259,P ＞ 0.05).The common hepatic artery lymph node metastatic rates of patients with diameter of tumor under 3 cm,3-5 cm and above 5 cm were 2.59％ (6/232),3.02％ (11/364) and 8.14％ (7/86),with significant difference (x2 =6.267,P ＜ 0.05).The common hepatic artery lymph node metastatic rates of patients in N0,N1,N2,N3 stages were 0(0/301),2.53％ (5/198),5.65％ (7/124) and 20.34％ (12/56),with significant difference (x2 =62.368,P ＜ 0.05).The common hepatic artery lymph node metastatic rates of patients in stage Ⅰ,Ⅱ,Ⅲ and Ⅳ were 0(0/62),1.78％ (6/337),5.06％ (13/257) and 19.23％ (5/26),with significant difference (x2=25.959,P ＜0.05).Two hundred and twenty-eight patients had postoperative complications with the complication rate of 33.43％ (228/682).The incidence of anastomotic fistula was the highest,which was 11.58％(79/682).Conclusions The metastatic rates of common hepatic artery lymph node in thoracic esophageal squamous cell carcinoma is the lowest.For patients suffered from esophageal cancer in stage I or the tumor diameter under 5 cm,the dissection of common hepatic lymph node can be ommitted in surgery.

Objective To investigate the effects of mir?483 on the proliferation,invasion and migration of nasopharyngeal carcinoma cells . Methods RT?qPCR was used to detect mir?483 expression level in different nasopharyngeal carcinoma cells(CNE?1,CNE?2)and the alteration of mir?483 expression level in these cells after radiation. CNE?1,one of the NPC cells,was selected. Cationic liposomes transient transfection was used to construct cells which overexpressed mir?483 or negative control . MTT assay was conducted to test the cell proliferation ability. Wound scratch assay and transwell migration assay were used to detect the effects of mir?483 on NPC invasion and migration ability. Results In terms of radiation resistance ,the expression level of mir?483 was higher in CNE?1. Overexpressing mir?483 can enhance the proliferation ,invasion and migration ability of CNE?1. Conclusions Mir?483 enhances the proliferation,invasion and migration ability of nasopharyngeal carcinoma cells.