昆明医学院学报2012，Vol.33Issue(8)：6-9,4.
简便快捷的小鼠胰岛分离纯化方法研究
A Convenient and Efficient Method of Mouse Pancreatic Islet Isolation and Purification
摘要
Abstract
Objective To investigate a convenient and efficient method of mouse pancreatic islets isolation and purification. Methods The isolation of mouse pancreatic islets was carried out by stationary digestion after multi-point injection of collagenase IV solution into the pancreas. A discontinuous Histopaque solution (1119, 1110, 1080 and 1060) was applied for the purification of the islets. Islet cell purity and viability were determined by dithizone ( DTZ) and Trypan blue staining. The islets were cultured overnight and stimulated with different concentration of glucose (0, 2.8 and 22.2mmol/L) for 1 hour. Supernatants were collected for insulin assays by an insulin RIA kit. Results (1) By this optimized islets isolation and purification protocol, each mouse pancreas could yield (114 ± 15) islets, with an average purity of (77.12 ± 3.23) % and viability more than 90% ; (2) The insulin level in 0, 2.8 and 22.2 mmol/L glucose stimulation groups were (23.80 ± 3.52) mIU/L, (67.57 ± 4.04) mlU/L and (164.32 ± 10.75) mlU/L, respectively. There was a statistically significant difference among groups (P <0.05). Compared with 0 mmol/L glucose group, the insulin level in 2.8 mmol/L glucose group was 2.84 times higher (P < 0.05) and the insulin level in 22.2 mmol/L glucose group was 6.90 times higher (P < 0.05). Compared with 2.8 mmol/L glucose group, the insulin level in 22.2 mmol/L glucose group was 2.43 times higher (P<0.05). Conclusions Multi-point injection of collagenase IV solution into the pancreas, followed by discontinuous density gradient seperation and manually picking up is a convenient and efficient mouse islet isolation and purification method. The islets isolated and purified by this optimized protocol can maintain functional insulin secretion potential in response to glucose stimulation.关键词
小鼠胰岛/多点注射/分离纯化Key words
Mouse islets/Multi-point injection/ Isolation and purification分类
医药卫生引用本文复制引用
合湫,苏恒,李超,沈涛,严新民,薛元明..简便快捷的小鼠胰岛分离纯化方法研究[J].昆明医学院学报,2012,33(8):6-9,4.基金项目
云南省自然科学基金资助项目(2005C0002R) （2005C0002R）
云南省中青年学术技术带头人后备人才项目(2007PY01-32) （2007PY01-32）
