摘要
目的探讨几种小鼠胰岛的分离纯化方法及进行纯化后胰岛的计数和完整性的分析。方法选用ICR小鼠,采用不同胶原酶消化方法和不同的胶原酶种类,并用Ficoll-PaqueTMPLUS密度梯度离心法对胰岛进行纯化,并对获得的胰岛进行DTZ染色计数和计算当量,扫描电镜考察胰岛的完整性。结果采用胶原酶V和P胰管灌注、内外消化结合,消化需时较短,胶原酶V和P在胰岛纯化前后每只小鼠收获的胰岛细胞数量和当量无差别(P>0.05);扫描电镜结果显示消化较好的胰岛表面被膜完整,消化过度的胰岛导致被膜不完整,易致外层细胞损伤。结论小鼠逆行胰管灌注胶原酶、内外消化相结合的胰岛消化方法所需时间较短,但同时要注意防止消化过度。
Objective To investigate the difference among several methods for isolation and purification of mouse islets and to analyze the amount and integrity after purification.Methods Two kinds of collagenases and digestion methods were choosed in this study with ICR mice,and Ficoll-PaqueTMPLUS were used to purify islets.The islets amount and equivalents（IEQ） were calculated through DTZ dyeing,and the integrity of the islets was analyzed under scanning electron microscopy.Results It needed less time in pancreas digestion by infusing collagenase V and P inversely into pancreatic duct and combining with internal and external digestion than digested only by external.There was no significant difference in islets number and equivalents（P 0.05） between collagen enzyme V and P before or after purification.The results of scan electron microscopy showed that the islet capsule would be less integrity,and the outer cells of islet would be damaged if the degree of digestion was excessive.Conclusion We can get mouse islets quickly by infusing collagenase inversely into mouse pancreatic duct,and digestion with the combination of internal and external digestion,but we should pay attention to the degree of digestion.
出处
《肝胆胰外科杂志》
CAS
2012年第5期403-405,414,共4页
Journal of Hepatopancreatobiliary Surgery
基金
国家自然科学基金资助项目(81071277)
浙江省自然科学基金资助项目(Y2080915)
浙江省大学生科技创新项目(2011R413006)
温州医学院本专科学生科研立项资助(wyx201101032)
