Objective:To explore the purification method of non- affinity chromatography for leptin expressed in Pichia Pastoris.Methods:Ion exchange chromatography (Sepharose Q fast flow) and hydrophobic interaction chromatography (Phenyl Sepharose 6 fast flow ) were used to purify human leptin in pH 7.5. Results: After purification by Q column,the purity of leptin increased from 42.3% to 89.6%.Subsequently, after hydrophobic interaction chromatography ,its purity reached 96.2%. In SDS-PAGE, leptin was shown as one specific band.Conclusion:The human recombinant leptin expressed by Pichia Pastoris yeast can be successfully purified by ion exchange chromatography plus hydrophobic interaction chromatography.

Objective:To develop a high cell-density fermentation process in micro DCU-400 fermentor of Pichia pastoris GS115 containing a recombinant vector pPICZ ?/endostatin.Methods:Cells were grown in the basal salts medium(BSM) at 30℃,pO 2≥30%,pH 5.3(growing) or pH 5.0(inducing),and the feed operation using 50% glycerol was performed when dissolved oxygen(pO 2) increased(after approximately 16-18h).It lasted 3-5hrs.When cell density(OD 600 ) reached 450,the inducement was began.After the transition from glycerol-methanol mixture to methanol,methanol from 0.03ml/min/L to 0.08ml/min/L in 8h was used to induce the endostatin expression.The inducement lasted for approximately 60h.Then centrifugation,the supernatant was collected.The expression was examined using 12% SDS-PAGE and ELISA.Results:In inducing phase,OD 600 reached 580(wet wt 275g/L).The production of endostatin is 27.2mg/L,exceeding that in spinner flask(16.3mg/L).Conclusion:The human endostatin had been successfully expressed in 15L fermentor.

The key to deepen quality education in university is to integrate the education of humanities into speciality instruction.Research shows that the ways to combine them are varied: starting from knowledge,mental enlightenment, aesthetic pentration, interaction, classroom atmosphere creation, and anthropocentrism ideal promotion,etc.

Considering the character of biochemistry,this paper discussed teaching method and accounted for the importance of experiment teaching in biochemistry by some demonstrations including analyzing carefully for catching common ground;integrating theory with practice for helping understand and memorize;agitating the students' study interest etc.In addition,the importance of students was not to be ignored in teaching process.

BACKGROUND: Leptin is a hormone produced predominantly by adipocytes and has a variety of physiological functions. It has been a hot spot in energy metabolism research. However, leptin presently used is usually produced by E coli, in which leptin cDNA is expressed and is in the form of insoluble inclusion body. Therefore, extremely complicated reactivating procedure is needed to obtain biologically activated human leptin.OBJECTIVE: To explore the condition for purification of leptin in non-affinity chromatography in order to obtain soluble human leptin.DESIGN: An observational experiment.SETTING: Molecular laboratory of biochemical department in a medical college.MATERIALS: The strong anion exchanger sepharose Q and hydrophobic phenyl sepharose 6 were used in different conditions for removal of as many contaminants as possible.METHODS: The supernatant of pichia pastoris yeast culture solution was first purified through Q column chromatography, the protein was collected and was further purified through hydrophobic support with 1 mol/ L (NH4) 2SO4.MAIN OUTCOME MEASURES: Gel scanning revealed that the purity of human leptin was 42. 3% before purification, 89.6% after Q column chromatography and 96.2% after hydrophobic interaction chromatography.RESULTS: The post-purification product presented a s. ingle band in SDS-PAGE. Gel scanning revealed that the purity of human leptin was 42.3% before purification, 89.6% after Q column chromatography and 96.2% after hydrophobic interaction chromatography.CONCLUSION: The combined use of strong anion exchange chromatography and hydrophobic interaction chromatography can effectively purify leptin expressed by pichia pastoris yeast and the purity is identical to that of nickel affinity column chromatography. It provides reliable evidence and method for possible manufacture of human leptin and lays experimental basis for leptin-related research.

Objective: To study early diagnosis and treatment of multi-primary esophageal cancer and esophageal and cardiac double primary cancer. Methods: The data of 71 cases patients of multi-primary esophageal, double primary esophageal and cardiac cancer were collected. The diagnosis was made by dye staining through gastroscopy and X-ray. Pathological examination after operation was analyzed. Results: 14 patients were diagnosed by X-ray(14/71), 69 by endoscopy (69/71), 54 early foci and 6 early stage patients were found. All of them were operated. The resection rate is 100% with no operative death. 3-year survival rate was 40 5%. Conclusion: Routine X-ray examination of esophagus or stomach, and esophageal dye-staining and/or biopsy through endoscopy are important measures for early diagnosis of multi-primary cancer or esophageal and double primary cardiac cancer prompt surgery is advised.

Objective To investigate the expression and clinical significance of the estrogen receptor(ER) subsets ER?,ER? and Ki-67 in tissues of different thyroid diseases.Methods The pathological data of 93 patients with thyroid diseases were collected from Jun.2003 to Jun.2007,including thyroid carcinoma(56 cases),thyroid follicular adenoma(13 cases),nodular goiter(14 cases) and normal thyroid tissue(10 cases).S-P immunohistochemistry was performed to determine the expressions of ER?,ER? and Ki-67 in the thyroid tissues.Results ER? and ER? were expressed in the plasma and nuclei of thyroid epithelial cells,and ER? was also expressed in the stroma.The positive rates of ER? expression in cases with thyroid carcinoma,thyroid follicular adenoma,nodular goiter and normal thyroid tissue was 87.5%,69.2%,42.9% and 30.0%,respectively,with significant difference between carcinoma and goiter or normal thyroid tissue(P0.05).Conclusion ER? and ER? may play an aggravating effect on cell proliferation in the disease course of the thyroid carcinoma.The expressions of ER? and Ki-67 are related to invasion and metastasis of thyroid carcinoma.

Objective To assess the value of thinprep liquid based cytology test (TCT) and Besthesda system (TBS) in diagnosis of cervial lesions. Methods 11000 cases of cervical lesions were detected with TCT and TBS.The samples of atypical squamous cells(ASC) and more severe than ASC lesions were classified into postive cytology,and then cytological results were compared with pathological examination results of cervical biopsies.Results The satisfactory rate of TCT samples was 96.36%, and the postive rate of abnormal epithelial cells diagnosed by TCT was 10.96%(1206/11000). Among 1206 cases, 15 cases(0.14%) of CA, 89 cases (0.81%) of HSIL, 449 cases (4.08%) of LSIL, 73 cases (0.66%) of ASC-H and 580 cases (5.27%) of ASC-US and AGUS. 1128 cases with postive cytologic results received pathological examination. The pathological positive rate of CA, HSIL, LSIL, ASC-H, and ASC-US and AGUS were 100%, 100%, 67.12%, 54.17% and 14.09%, respectively. The coincident rate of two examination methods was 100% for CA, 91.1% for HSIL,57.82% for LSIL and 27.78% for ASC-H. Conclusion TCT and TBS can accurately detect cervical cancer, precancerous lesions and infection of bacteria, virus and other pathogens, and at present is a means of screening cervical cancer.

Colon cancer is a common and deadly disease with high rising incidence rate.Proteomics has already become one of the hot topic and maken great progress in the research of colon cancer pathogenesis,early diagnosis,treatment and prognosis.In recent years,more and more colon cancer-associated biomarkers have been identified through proteomics-based approaches.This review will summarize the research progress of proteomics in colon cancer.

Objective To explore the effects of chlamydia and mycoplasma infections on the healing of wounds after surgical operation for haemorrhoids,fistula and anoschisis.Methods Sixty-two patients with haemorrhoids,fistula and anoschisis undergoing the surgical operation during 2000.1 to 2008.10 were collected,and wounds did not heal 40 days after operation and the wound's surface infection occurred after postoperative anti-inflammatory therapy and dressing change.All patients were positive at least for laboratory Ct or Uu,and those with infections caused by other fungi,bacteria,viruses and the other systemic diseases were excluded.The correlation between the infections in 62 cases and sex,surgical types was analyzed.Results In 62 patients,there were 30 cases (48.4%) positive for both Ct and Uu,22 cases for single Ct (35.5%),and 10 cases for single Uu (16.1%),respectively.The infection rate in females was higher than in males (P<0.01).The surgical types included:surgery for 8 cases of haemorrhoids (12.9%),for 18 cases of fistula(29.0%),and for 36 cases of anoschisis(58.1%).respectively.Conclusion There is the possibility of chlamydia and mycoplasma infections in the patients with delayed healing of wounds following the surgical treatment on the anal region.Early diagnosis,early treatment,avoidance of ineffective medicines and repeatedly management of the wounds can shorten the healing period.