On the base of the one step, operator independent method which was set up by Christophe A.E., the pancreas was infused with cold University of Wisconsin(UW) solution for the preservation, ...On the base of the one step, operator independent method which was set up by Christophe A.E., the pancreas was infused with cold University of Wisconsin(UW) solution for the preservation, digested by the collagenase P, circuited with HBSS+5%fetal calf serum(FCS)+10mmol/L Hepes solution, and separated with the stainless steel mesh. The number of the collected islets were 400000～1800000 per pancreas, i.e. about 12150/g pancreas. After purification, the recovery was 350000～1700000 per pancreas, i.e. about 10250/g pancreas, the recovery rate was above 80%, and the purity of the final preparation was above 95%. The insulin secretion in the response to the high concentration glucose (22 mmol/L) stimulation was apparently different on the 1,3,5 day of the cultural islets, which the high level of insulin was three times the low level (5.5 mmol/L) on the 5th day, and the insulin level of the double stimulation under perfusion conditions is apparently higher than low glucose. The result demonstrated that the purified islets were functionally alive. Histological studies also show that the shape of islets are complete, and the β cell was specially stained by the dithizone (DTZ). The Trypan Blue staining had shown the living cell was above 90%. In conclusion, the new method was highly practical and yielded higher concentration of active pancreatic islets.展开更多
文摘目的 应用性染色体双色间期荧光原位杂交技术检测异性间造血干细胞移植后,患者的嵌合状态,并对其应用价值进行分析.方法 对2006年1月～2007年12月在天津血液病医院行异性间allo-HSCT的32例患者,常规染色体核型分析采用不加任何刺激剂的骨髓24 h短期培养法制备染色体,火焰法滴片,R显带.应用X/Y性染色体双色着丝粒DNA探针对骨髓间期细胞行荧光原位杂交,检测移植后患者的嵌合状态.结果 32例患者染色体核型分析115例次,其中111例次为正常供者核型,1例次受者核型,3例次分析失败,未能检测到混合核型.双色性染色体间期FISH检测163例次,129例次为完全嵌合,34例次为混合嵌合,检测成功率为100%.移植后2～3 w 32.2%患者为混合状态,移植后1 mo 76.7%患者为完全嵌合,移植后半年94.1%患者为完全嵌合.结论 性染色体双色间期荧光原位杂交能够简便、快速、灵敏地检测出异性间allo-HSCT患者移植后的嵌合状态.
文摘On the base of the one step, operator independent method which was set up by Christophe A.E., the pancreas was infused with cold University of Wisconsin(UW) solution for the preservation, digested by the collagenase P, circuited with HBSS+5%fetal calf serum(FCS)+10mmol/L Hepes solution, and separated with the stainless steel mesh. The number of the collected islets were 400000～1800000 per pancreas, i.e. about 12150/g pancreas. After purification, the recovery was 350000～1700000 per pancreas, i.e. about 10250/g pancreas, the recovery rate was above 80%, and the purity of the final preparation was above 95%. The insulin secretion in the response to the high concentration glucose (22 mmol/L) stimulation was apparently different on the 1,3,5 day of the cultural islets, which the high level of insulin was three times the low level (5.5 mmol/L) on the 5th day, and the insulin level of the double stimulation under perfusion conditions is apparently higher than low glucose. The result demonstrated that the purified islets were functionally alive. Histological studies also show that the shape of islets are complete, and the β cell was specially stained by the dithizone (DTZ). The Trypan Blue staining had shown the living cell was above 90%. In conclusion, the new method was highly practical and yielded higher concentration of active pancreatic islets.
