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Objective To establish a method to prepare anti-sodium estrone sulfate monoclonal antibody ( ESS-Mab) . Methods Balb/c mice were immunized by ESS. Immune methods were screened. The blood serum potencies were measured by indirect ELISA and the best consistence of antigen and the first antibody were confirmed with method of titration. Cell fusion was carried by using PEG method and McAb hybridoma was screened with the indirect ELISA. Results The best immunization method of mice was subcutaneously multi-point injection in mouse back with the dose of 200/100 μg ESS antigen five times. The fusion rate was 90. 2%. Hybridoma positive rate of ELISA screening was 4. 4%. Finally two cell lines 2C8 and 8A7 with good specificity and sensitivity were obtained. Conclusion The best immunization way is selected and indirect ELISA is set up effectively and reliably for screening and presenting ESS McAb. the hybridoma technique is able to prepare monoclonal antibody of anti-ESS successfully.
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Objective To screen the differentially expressed genes in esophageal squamous cell cancer (ESCC) and normal tissue of esophageal mucosa in Kazakh. Methods RNA was extracted from the ESCC sections in Kazakh patients, and was amplified to obtain cRNA. The gene expression profiles in ESCC and normal tissue of esophageal mucosa were detected by HG-U133 Plus 2.0 gene chip. The results were analyzed by bioinfor-matics. Results One hundred and seventy differentially expressed genes in ESCC and normal tissue of esophageal mucosa were found, with a difference of more than 10 times in expression levels. Of the 170 genes, 39 were up-regulated (signal log ratio > 3 ) and 131 down-regulated (signal log ratio < - 3). These factors such as cell cycle regulation, apoptosis, cytoskeleton; extracellular matrix, intracellular signal transduction, protein translation and synthesis, and immunological functions were correlated with the genes with abnormal expression. Conclusion The use of oligonucleotide microarray accurately and efficiently screen the 170 target genes in ESCC in Kazakh. It is suggested that these genes may be related to the carcinogenesis and development of ESCC in Kazakh.
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Objective To explore the effect of the DNA methyltransferase 5-aza-2'-deoxycytidine (5-aza-CdR)on human esophageal squamous cancer Ecal09 cells.nethods Human esophageal squamous cell cancer(ESCC)Eca109 cells were treated by 5-aza-CdR with 10-7,10-6,1O-5,10-4,0 mol/L. Respectively.Consequently,the growth rate of the cells was detected by MTT assay and morphological structure Was observed.Meanwhile,cell apoptosis and cell cycle were analyzed by flow cytometry method. (FCM).ResulIs The proliferation of Eea109 cells Was inhibited by 5-aza-CdR from 10-7 to 10-4 mol/L Moreover,the inhibition rate showed time-and-concentration-dependent manner(24～96 h,F=160.06,P=0.000,10-7～10-4 mol/L,F:60.95,P:0.000).The maximum rate of inhibitory was reached up to(15.70±0.75)% in the group treated by 10-4 mol/L 5-aza-CdR after 96 hours.An apoptosis peak appeared before diploid peak.The proportion of Go/G1 phase cells Was significantly increased(F=6479.46, P=0.000),especially up to(89.70±0.91)% in the group treated by 10-4 mo/L 5-aza-CdR after 96 h. However,the proportion of S phase cells Was obviously decreased(F=4222.26,P=0.000),especially down to(9.10±0.48)% in the group treated bv 10-4mol/L 5-aza-CdR after 96 h.Conclusions The proliferation of Eca109 cells is inhibited bv 5-aza-CdR in a time-and-concentration-dependent manner.Moreover,the 5-aza-CdR can inhibit cell growth by regulation of DNA cycle and apoptosis.
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Objective To study the effects of nicotinamide on the human skin melanocytes and try to explore the potential mechanism of nicotinamide on the calcium signal transduction,cytoskeleton.Methods The primary cultured human skin melanocytes from foreskin were selected as the target cells in the present study.0.00,0.05,0.25,1.25 and 6.25 mg/ml nicotinamide were applied respectively.Western blot,fluorescent probes(Flu-3AM),flow cytometry analysis and time-lampse microscope digital skill were used to evaluate the effects of nicotinamide on melanosome motility and the melanosome distribution in melanocytes.Results The results showed that nicotinamide had a potential effect on regulating free calcium concentration in a dose-dependent manner(y=76.461 2-5.435x,r=0.97);The activity of Na+-K+-Ca2+-ATPase was down regulated with the increasing concentration of nicotinamide.The expression of cellular dynein was also altered by nicotinamide;Na +-K+-Ca2+-ATPase activity was kept normal when given 0.05,0.25 mg/ml nicotinamide,while the dynein protein expression was inhibited(P