Objective To establish a novel flow cytometric crossmatch assay allowing detection of complement-fixing donor specific anti-HLA IgG alloantibodies (Flow cytometry complement-dependent crossmatch,Flow-CDC). Methods One hundred pretransplant crossmatchings were performed using Flow-CDC and NIH-CDC between 62 patients awaiting renal transplantation and 33 donor cells.These crossmatchings were divided into two groups according to PRA.Group 1 consisted of 25 sera with negative PRA,and group 2 consisted of 75 sera with positive PRA.All of the sera were pretreated with DTT to inactivate IgM. Lymphocytes were isolated from peripheral blood (or,in a few instances,from the spleen) of the cadaveric donors. The correlation between different techniques for detection of donor specific anti-HLA antibodies was evaluated.The effect of both methods on clinical transplantation outcome was observed. Results In group 1,NIH-CDC and Flow-CDC were negative for all 25 sera.In group 2,24 (32.0%) had a positive NIH-CDC,31 (41.3%) had a positive Flow-CDC.There was a significant difference between two methods (?2=5.14, P= 0.016 ).Overall concordance between both tests was 93% with 69 concordant negatives and 24 concordant positives. The correlation coefficient (r) was 0.80.In group 2,5 patients received transplantation. One of them with negative NIH-CDC and positive Flow-CDC suffered from acute rejection after transplantation and lost the graft,and the other patients with negative NIH-CDC and Flow-CDC had good outcome. Conclusions Flow-CDC can detect specifically complement-fixing IgG alloantibodies against donor HLA and is more sensitive than NIH-CDC.Additionally,the computer printouts represent a permanent record of the crossmatch for retrospective review.Flow-CDC may become the standard crossmatch method as a possible alternative to conventional NIH-CDC testing.

With the application of monoclonal antibody technology more and more widely, its production technology is becoming more and more perfect. Small molecule monoclonal antibody technology is becoming a hot research topic for people. The application of traditional Chinese medicine small molecule monoclonal antibody technology has been more and more widely, the technology for effective Chinese medicine component knockout provide strong technical support. The preparation of monoclonal antibodies and small molecule knockout technology are reviewed in this paper. The preparation of several steps, such as: in the process of preparation of antigen, hapten carrier coupling, coupling ratio determination and identification of artificial antigen and establishment of animal immunization and hybridoma cell lines of monoclonal antibody, the large-scale preparation; small molecule monoclonal antibody on Immune in affinity chromatography column method is discussed in detail. The author believes that this technology will make the traditional Chinese medicine research on a higher level, and improve the level of internationalization of Chinese medicine research.
Animals ; Antibodies, Monoclonal ; chemistry ; genetics ; immunology ; Humans ; Hybridomas ; metabolism ; Immunologic Techniques ; methods ; trends

This study examined the promoter methylation of APO-1/CD95 (Fas) gene in bladder urothelial carcinoma and analyzed the relationship between the Fas promoter methylation and the biological behavior of bladder cancer. Promoter methylation of Fas gene was detected by methylation-specific PCR (MSP) in 4 bladder cancer cell lines, 50 human bladder urothelial carcinoma samples and l0 normal bladder tissue samples. Correlation of the aberrant methylation of Fas promoter with the clinicopathological parameters was statistically analyzed. The results showed that Fas was down-regulated at both mRNA and protein level in bladder cancer cell lines and tissue samples of bladder urothelial carcinoma. The positive rate of Fas protein expression in bladder urothelial carcinoma was 34.0% (17/50), significantly lower than that in normal bladder tissues (70.0%, 7/10) (P<0.01). Fas promoter methylation was detected, and the positive rate of Fas promoter methylation in bladder urothelial carcinoma was 42.0% (21/50), which was obviously higher than that in normal bladder tissues (0.0%, 0/10) (P<0.01). The aberrant methylation of Fas promoter was reversely correlated with Fas protein expression (P<0.05). Furthermore, the positive rates of Fas promoter methylation in high-grade and low-grade bladder urothelial carcinoma were 73.3% (11/15) and 34.2% (12/35), respectively, with significant difference shown (P<0.05). No statistical significance was found in the Fas promoter methylation among different clinical stages of bladder cancer. It was concluded that Fas promoter hypermethylation plays an important role in the pathogenesis of bladder urothelial carcinoma and may serve as a prognostic indicator of bladder urothelial carcinoma.

Objective To investigate the subtype distribution and sequence characteristics of HIV-1 strains prevalent among sexual infectors in Beijing. Methods We collected the blood samples from 100HIV sexual infectors in Beijing during 2008 and separated plasma specimens. RNA was extracted from the plasma and the gag gene was amplified by RT-PCR and nest-PCR. The PCR products were sequenced directly and phylogenetic analyses of gag gene was performed using the MEGA4 software. Results Among 100 HIV-1 plasma samples,84 gag gene fragments were amplified and analyzed. Eight HIV subtypes including B(22 strains), B'(8 strains),C( 1 strain) ,CRF01_AE (38 strains) ,CRF02_AG (2 strains) ,CRF07_BC(9 strains) ,CRF08_BC(3 strains) and C/CRF01_AE recombinant like strain( 1 strain) were identified circulating in Beijing. Conclusion CRF01 _AE and subtype B were predominant in Beijing account for 45.2% and 26.2% and the surveillance of HIV gene variation should be paid more attention.

Objective:To compare the efficacy of bilateral and unilateral metallic stenting in the treatment of unresectable malignant hilar biliary obstruction.Methods:From January 2012 to December 2018, a total of 300 patients with malignant hilar biliary obstruction (Bismuth type Ⅱ-Ⅳ) were treated with endoscopic metallic stenting in Oriental Hepatobiliary Surgery Hospital. Ninety-four patients with bilateral metallic stenting (bilateral metallic stent group) and 94 patients with unilateral metallic stenting (unilateral metallic stent group) were matched and analyzed by propensity score matching. The clinical success rate, the total number of interventions, the stent patency and overall survival time were compared between the two groups.Results:The clinical success rate in the bilateral metallic stent group was significantly higher than that of the unilateral metallic stent group [98.9% (93/94) VS 78.7% (74/94), χ2=19.352, P<0.001], and the mean number of intervention in the bilateral metallic stent group was significantly less than that of the unilateral metallic stent group (1.2±0.5 times VS 1.7±0.9 times, t=-4.345, P<0.001). The stent patency time in the bilateral metallic stent group was significantly longer than that of the other group [10.0 (8.0, 12.1) months VS 5.7 (5.2, 6.3) months, χ2=19.789, P<0.001]. The median survival time of patients in the bilateral group was numerically longer than that in the unilateral group, but did not reach statistical significance [7.6 (6.3, 8.9) months VS 4.6 (3.7, 5.7) months, χ2=3.628, P=0.057]. Conclusion:Endoscopic bilateral metallic stenting is superior to unilateral metallic stenting in the clinical management of malignant hilar biliary obstruction unsuitable for surgery.

Objective:To investigate the feasibility and efficacy of endoscopic placement of transpapillary parallel-style bilateral metal stents for malignant hilar biliary stricture.Methods:A total of 55 patients with malignant hilar biliary stricture (Bismuth Ⅱ-Ⅳ) were treated with an improved endoscopic drainage technique (endoscopic transpapillary parallel-style bilateral metal stenting) from January 2012 to December 2018 in Shanghai Oriental Hepatobiliary Surgery Hospital. The technical success rate, clinical success rate, incidence of complications, success rate of endoscopic re-intervention, stent patency and overall survival were analyzed.Results:The technical success rate of endoscopic placement of bilateral metal stents was 96.4% (53/55), the clinical success rate was 96.2% (51/53), and the early complication incidence was 13.2% (7/53). The median stent patency was 9.2 months (95% CI: 8.0-10.3 months), the success rate of endoscopic re-intervention of bilateral biliary systems was 92.3% (12/13), and the median overall survival was 6.7 months (95% CI: 4.7-8.8 months). Conclusion:This modified technique of bilateral metal stent placement is safe and effective for malignant hilar biliary stricture that is unsuitable for surgery.

12E7 Antigen ; Aged ; Antigens, CD ; metabolism ; Carcinoma, Transitional Cell ; pathology ; Carcinosarcoma ; pathology ; Cell Adhesion Molecules ; metabolism ; Cystectomy ; methods ; Diagnosis, Differential ; Follow-Up Studies ; Humans ; Male ; Nephrectomy ; Osteosarcoma ; metabolism ; pathology ; surgery ; Ureter ; surgery ; Ureteral Neoplasms ; metabolism ; pathology ; surgery ; Vimentin ; metabolism

OBJECTIVETo elucidate the significance of detecting cytokeratin gene (CK-20 mRNA) expression in metastatic lymph nodes of colon carcinoma patients.

METHODSThe expression of CK-20 mRNA was detected in 342 lymph nodes by reverse transcription-polymerase chain reaction (RT-PCR) in 49 colon carcinoma patients. Its incidence was compared with that by routine pathologic examination.

RESULTSThe positive rates of detecting metastasis in the lymph nodes were 21.9% by RT-PCR and 11.1% by routine pathologic examination.

CONCLUSIONThe detection of cancer metastasis in the lymph nodes in colon carcinoma is almost doubled (21.9% vs 11.1%) by CK-20 mRNA, which may provide a guiding significance in staging, treatment planning and prognosis in the prognostic estimation of colon carcinoma patients.

Adult ; Aged ; Colonic Neoplasms ; diagnosis ; genetics ; pathology ; Female ; Gene Expression ; Genetic Markers ; genetics ; Humans ; Intermediate Filament Proteins ; genetics ; Keratin-20 ; Lymphatic Metastasis ; diagnosis ; Male ; Middle Aged ; Neoplasm Staging ; Prognosis ; RNA, Messenger ; biosynthesis ; Reverse Transcriptase Polymerase Chain Reaction

AIMTo study the vasorelaxation action of oxyphenamone (Oxy) and its mechanism.

METHODSThe contractile response of isolated rabbit renal, femoral and mesentery artery preparations was determined.

RESULTSOxy was shown to inhibit the contractile force of renal, femoral and mesentery arteries induced by phenylephrine in a concentration dependent manner. The vasorelaxation produced by Oxy was not attenuated by removal of the endothelium. Oxy (10(-6)-10(-4) mol.L-1) relaxed the contractions induced by KCl 30 mmol.L-1 as well as KCl 80 mmol.L-1, but the contraction curve of KCl 80 mmol.L-1 was shifted significantly to the right. Oxy in lower concentration (10(-6) and 5 x 10(-6) mol.L-1) increased the contractions induced by Ang II, and in middle concentration (10(-5) mol.L-1) it did not affect the contractions induced by Ang II. Whereas in higher concentration (5 x 10(-5) mol.L-1) it obviously inhibited the contractions induced by Ang II.

CONCLUSIONOxy showed significant vasorelaxation to various vascular preparations, and its vasorelaxation action is endothelium independent. The mechanism of its vasorelaxations seems to be related with Ca2+ activated K+ channel (Kca channel) and Ca2+ channel in vascular smooth muscle cells but its true mechanism needs further study.

Animals ; Cardiotonic Agents ; pharmacology ; Female ; Femoral Artery ; drug effects ; physiology ; In Vitro Techniques ; Male ; Mesenteric Arteries ; drug effects ; physiology ; Organic Chemicals ; Phenylephrine ; antagonists & inhibitors ; Potassium Channels, Calcium-Activated ; metabolism ; Rabbits ; Renal Artery ; drug effects ; physiology ; Vasodilation ; drug effects ; Vasodilator Agents ; pharmacology

To investigate the method of RNA isolation from human embryonic tissues and the factors influencing the quality of RNA, the RNA from human embryonic tissues obtained with drug-induced labor or non-drug induced labor were isolated by using grind with liquid nitrogen or homogenizer without liquid nitrogen. The results showed that the positive rates of RNA integrity in grind with liquid nitrogen group and in homogenizer without liquid nitrogen group were 68.42% and 29.79% respectively, and there was significant difference between these two groups; however, there was no statistic difference in positive rate of RNA integrity, OD(260)/OD(280) ratio and beta-actin gene expression level between the drug-induced labor group and non-drug induced labor group. It is concluded that pulverize of tissue in liquid nitrogen remains the integrity of RNA isolated and may be applied for RNA isolation from human embryonic tissues. The quality of RNA is not affected by different methods of induction of maternal labor.
Embryo, Mammalian ; metabolism ; Freezing ; Humans ; Nitrogen ; pharmacology ; RNA ; isolation & purification ; RNA Stability ; drug effects