Fujioka T, Terasaki P I, Heintz R, Merideth N, Lanza R P, Zheng T L, Soon-Shiong P
Department of Surgery, UCLA School of Medicine.
Transplantation. 1990 Feb;49(2):404-7. doi: 10.1097/00007890-199002000-00035.
A simple, rapid method of islet purification is important in large-scale human islet isolation. We have previously identified monoclonal antibodies specific for acinar cells, but not islets, and described an immunologic method of purification by selective lysis of the acinar cells. An attractive alternative to lysis of the acinar cell is depletion by a magnetic immunomicrosphere technique. We report in this study a rapid, reproducible method of rat islet purification utilizing magnetic microspheres coated with acinar-cell-specific monoclonal antibodies. Pancreatic digestion with collagenase followed by depletion of acinar cells with the magnetic immunomicrospheres (MIMS) yields large numbers of intact islets. We compared the islets thus obtained with hand-picked (HP) islets (control) for yield, purity, in vitro insulin secretory capacities, and in vivo functional viability. The islet yield with the MIMS method (n = 35) was 72.7% that obtained with the HP method (n = 6) (378 +/- 8 vs. 519 +/- 31 islets per pancreas). The purity of the MIMS-isolated islets was 84 +/- 1.9%, ranging from 75-95%. Static glucose stimulation showed excellent function (2-3-fold increase of insulin release over basal levels) with no statistical difference in insulin secretion between MIMS and HP islets. Under microscopic examination, both groups revealed a well-preserved structure with healthy endocrine cells. When 1321 +/- 59 MIMS islets were transplanted into streptozotocin-induced diabetic rats (n = 10), normoglycemia (less than 200 mg/dl) was restored in all recipients following transplantation, and 100% of them remained normoglycemic on day 120 postgrafting. In summary, a rapid, consistent, and simple method of isolating viable, purified rat islets is described. The broad interspecies crossreactivity of the McAb suggests that this technique may be generally useful for islet purification in large mammalia, including man.
一种简单、快速的胰岛纯化方法对于大规模人类胰岛分离至关重要。我们之前已鉴定出对腺泡细胞而非胰岛具有特异性的单克隆抗体，并描述了一种通过选择性裂解腺泡细胞进行纯化的免疫方法。腺泡细胞裂解的一种有吸引力的替代方法是磁免疫微球技术去除。我们在本研究中报告了一种利用包被有腺泡细胞特异性单克隆抗体的磁性微球快速、可重复的大鼠胰岛纯化方法。用胶原酶消化胰腺，随后用磁免疫微球（MIMS）去除腺泡细胞，可获得大量完整的胰岛。我们将由此获得的胰岛与手工挑选（HP）的胰岛（对照）在产量、纯度、体外胰岛素分泌能力和体内功能活力方面进行了比较。MIMS方法（n = 35）的胰岛产量是HP方法（n = 6）的72.7%（每个胰腺分别为378±8个与519±31个胰岛）。MIMS分离的胰岛纯度为84±1.9%，范围在75%至95%之间。静态葡萄糖刺激显示功能良好（胰岛素释放比基础水平增加2至3倍），MIMS和HP胰岛之间的胰岛素分泌无统计学差异。在显微镜检查下，两组均显示内分泌细胞健康且结构保存良好。当将1321±59个MIMS胰岛移植到链脲佐菌素诱导的糖尿病大鼠（n = 10）中时，所有受体在移植后血糖恢复正常（低于200 mg/dl），并且100%的受体在移植后第120天仍保持血糖正常。总之，描述了一种分离有活力、纯化的大鼠胰岛的快速、一致且简单的方法。单克隆抗体的广泛种间交叉反应性表明该技术可能普遍适用于包括人类在内的大型哺乳动物的胰岛纯化。
