The directed molecular evolution of enzyme in vitro can not only improve the efficiency of evolution, but also evolve enzyme according to the investigator's desire. This review summed up the feasible methods of this novel technique.

This paper introduces the principles of an Environmental Temperature Supervising Network System, explain its hardware and software structure and the function of network system. The whole system is proved stable and reliable by experiment. It may have some profit in the market and great future in the application.

In order to construct a prokaryotic expression vector of human receptor syt II N-fragment and to express recombinant MBP-Syt fusion protein in E.coli and to purify and identify its activity. According to codon preference of E.coli, a DNA fragment encoding human syt II N-fragment was synthesized, and then cloned into prokaryotic vector pMAL-c2x for sequencing. Then the recombinant plasmid pMAL-Syt was introduced into E.coli ER2566 by transformation for expression and the obtained engineered bacteria were induced by IPTG. The fusion protein was purified by amylose resin affinity chromatography and identified by SDS-PAGE and Western blot. The binding activity of the protein was determined by ELISA. It is concluded that MBP-Syt protein is of good binding activity.

BACKGROUND: Results from clinical trials suggested that clopidogrel and ticlopidine had side effects of granulopenia, and aspirin could inhibit endothelial progenitor cell proliferation. There is no report of effects of these drugs on human bone marrow mesenchymal stem cells (hBMSCs) in stem cell transplantation. OBJECTIVE: To investigate the effects of antiplatelet drugs including clopidogrel, ticlopidine and aspirin on hBMSC proliferation and secretion. DESIGN, TIME AND SETTING: The cytology in vitro observation was performed at the Laboratory of Toxicology, Shanghai Municipal Center for Disease Control and Prevention from March to December 2006.MATERIALS: The second passage of hBMSCs was kindly donated from Shanghai Tissue Engineering Research & Development Center, Shanghai Ninth People's Hospital. Clopidogrel (Lot number J20040006) and ticlopidine (Lot number H19980186) were obtained from Hangzhou Sanofi-Synthelabo Minsheng Pharmaceutical CO., Ltd. Aspirin (Lot number 20050059) was obtained from Bayer Vital GmbH. METHODS: The standard culture medium consisted of DMEM-LG, 10% heat-inactivated FBS, 100 U/mL penicillin and 100 μg/mL streptomycin. After being cultured in vitro expanded out to passage 6, hBMSCs were treated with antiplatelet drugs of different concentrations and compared with control group. MAIN OUTCOME MEASURES: Cell proliferation was assessed by 3- (4, 5-dimethylthiazol -2-yl)-2,5-diphenyltetrazolium bromide (MTT) colorimetric assay, level of vascular endothelial growth factor (VEGF) of culture medium was detected by enzyme-linked immunoadsordent assay (ELISA), and surface antigens of hBMSCs were analyzed by the flow cytometry. RESULTS: A570 values of hBMSCs treated by clopidogrel or ticlopidine of 0.02,0.1,0.4,2,10,40 μmol/L were higher than control group (P < 0.01), while A570 values of aspirin group of 60, 600, 2 000 μmol/L were lower than control group(P < 0.05). Antiplatelet drugs had no obvious effect on cell surface antigens(CD34, CD105, CD106)expressed by hBMSCs. Treated by high dose clopidogrel or ticlopidine (40 μmol/L), VEGF level from hMSCs was lower than that of control group(P < 0.01), but VEGF level of low dose (0.02 μmol/L) ticlopidine group was higher than control group(P < 0.01), and there was no significantly difference of VEGF level among low dose clopidogrel group (0.02 μmol/L), aspirin group (5, 2 000 μmol/L), and control group(P > 0.05). CONCLUSION: Clopidogrel and ticlopidine improve proliferation of hBMSCs, but aspirin inhibits proliferation of hBMSCs. High dose of clopidogrel and ticlopidine suppress VEGF secretion of hBMSCs, while low dose of ticlopidine promote it. Antiplatelet drugs have no obvious effect on hBMSCs differentiation.

Objective To study the diagnostic value of double-balloon endoscopy (DBE) and capsule endoscopy (CE) for small intestinal bleeding. Methods Overall detection rates of small intestinal bleeding with DBE, CE and the whole alimentary tract barium meal were compared. Positive rates of bleeding detection with DBE and CE were compared within the same patients. Influence of CE on one-procedure rate of DBE was analyzed. Results In 105 cases of small intestine bleeding, DBE detected 24 cases of Crohn's disease, 15 adenocarcinoma, 12 chronic nonspecific inflammation, 10 small intestinal ulcer of unknown reason, 8 entero-mesenchymoma, 8 polypus, 6 vascular deformation hemorrhage, 5 ancylostomiasis, 5 Mechel's diverticula ( including multiple diverticula), 3 lymphoma and 9 of no evident abnormalities. The positive detection rate of DBE is 91.4% (96/105). Disease detection rates of CE and whole alimentary tract barium meal were 75.0% (30/40) and 33.3% (25/75), respectively. The one-procedure rate of DBE is 90% (36/40) based on CE results, but it was only 69. 2% (45/65) according to clinic features and the whole alimentary tract barium meal. Conclusion The main causes of small intestinal bleeding are benign ulcers (including Crohn's disease) and tumor, as well as chronic inflammation. Polyps, vascular deformation, parasitosis, Mechel's diverticulum and lymphoma are the secondary causes.DBE is superior to CE in diagnosis of small intestine bleeding, but CE can increase the one-procedure rate of DBE.

Sleep quality is closely related to human health. It is very important to correctly discriminate the sleep stages for evaluating sleep quality, diagnosing and analyzing the sleep-related disorders. Polysomnography (PSG) signals are commonly used to record and analyze sleep stages. Effective feature extraction and representation is one of the most important steps to improve the performance of sleep stage classification. In this work, a collaborative representation (CR) algorithm was adopted to re-represent the original extracted features from electroencephalogram sig- nal, and then the kernel entropy component analysis (KECA) algorithm was further used to reduce the feature dimension of CR-feature. To evaluate the performance of CR-KECA, we compared the original feature, CR feature and readied CR feature (CR-PCA) after principal component analysis (PCA). The experimental results of sleep stage classification indicated that the CR-KECA method achieved the best performance compared with the original feature, CR feature, and CR-PCA feature with the classification accuracy of 68.74 +/- 0.46%, sensitivity of 68.76 +/- 0.43% and specificity of 92.19 +/- 0.11%. Moreover, CR algorithm had low computational complexity, and the feature dimension after KECA was much smaller, which made CR-KECA algorithm suitable for the analysis of large-scale sleep data.
Algorithms ; Electroencephalography ; Entropy ; Humans ; Polysomnography ; Principal Component Analysis ; Sleep Stages ; Sleep Wake Disorders ; diagnosis ; Software

Objective To study the severity of systemic inflammatory response to type B epidemic encephalitis and blood tumor necrosis factor complicated with respiratory failure.Methods Fourteen children with type B epidemic encephalitis were divided into two groups;6 cases in group one with respiratory failure were given mechanical ventilation and the other 8 cases had normal respiration.Breat-hing pattern,respiratory rate,GCS scores,daily dosage of sedatives were observed before endotracheal intubation;while heart rate,temperature,WBC,CRP,TNF were measured.Results Patients in respiratory failure group were given a small dosage of Luminal;there was no significant difference in GCS between two groups treated with compound Wintermin.The heart rate,temperature,WBC,CRP,TNF levels in respiratory failure group were significantly higher than those in normal respiratory group.Conclusions The severity of systemic inflammatory response is more severe in epidemic type B encephalitis with respiratory failure than the controls.Significant increase in blood TNF may be an important factors to cause peripheral respiratory failure.

Objective To investigate the relapse risk assessment for patients with acute promyelocytic leukemia(APL)through testing PML-RAR? fusion gene by real-time quantitative polymerase chain reaction(RQ-PCR).Methods Relative copies of PML-RAR? fusion gene were measured in 25 patients with APL in phases of first diagnosis,complete remission(CR)and relapse.The minimal residual disease(MRD)situations were also monitored in 6 of the 25 patients.Results Different PML-RAR? fusion gene expression levels were observed in patient groups of different phases of the disease.(P

Objective　To investigate the significance of TGF-β1, TGFRl and TGFR2 in the pathogenesis and prognosis in patients with myelofibrosis. Methods　The expression of TGF-β1 and its receptors (TGFR1 and TGFR2 ) in bone marrow tissues and the level of TGF-β1 in the blood of 23 patients with myelofibrosis were detected by SABC immunocytochemistry and ELISA repectively. Results　Expression of TGF-β1 and TGFR 1 was significantly higher in primary and secondary myelofibrosis patients than that of the control. No significant difference of TGFR2 expression was found between the groups of myelofibrosis and the control (P>0.05). The level of TGF-β1 in the blood of the patients with myelofibrosis was significantly higher than that of the control (P<0.01) and more obvious in secondary cases while TGF-β1 decreased nearly to the normal level when patients were in clinical remission. Conclusion　TGF-β1 and it's receptors may be involved in the pathogenesis of myelofibrosis and might be of importance for the prognosis of the patients with myelofibrosis.

To produce antibodies capable of neutralizing botulinum neurotoxin type B(BoNT/B),We cloned the carboxy-terminal end of Hc containing the major determinants responsible for specific toxin,induced and purifed.The heavy-chain and kappa light-chain variable region gene repertoire of immunoglobulin were amplified individually from the spleen cell mRNA by RT-PCR and joined as a single-chain Fv(scFv)DNA fragment.These fragment were cloned into the phagemid pCANTAB5E and the phage display library was constructed.Results showed that the high affinity scFv was obtained after 4 rounds of panning,with its DNA sequence conforming to that of mouse antibody.