Laboratory of Cell Therapy in Diabetes, Department of Pediatrics, Faculty of Medicine and Odontology, Cruces University Hospital, University of the Basque Country (UPV/EHU), Barrio Sarriena, s/n, 48940, Leioa, Vizcaya, Spain,
Cytotechnology. 2015 Mar;67(2):199-206. doi: 10.1007/s10616-014-9690-7. Epub 2014 Jan 19.
Pancreatic islet transplantation is a promising therapy for Type I Diabetes. For many years the method used worldwide for islet purification in both rodent and human islet isolation has been Ficoll-based density gradients, such as Histopaque. However, it is difficult to purify islets in laboratories with staff limitations when large scale isolations are required. We hypothesized that filtration could be a more simple and fast alternative to obtain good quality islets. Four separate islet isolations were performed per method, comparing filtration and Histopaque purification with handpicking as the gold standard method for islet purity. Different parameters of quality were assessed: yield in number of islets per pancreas, purity by dithizone staining, viability by Fluorescein Diacetate/Propidium Iodide vital staining and in vitro functionality assessed by Glucose Stimulated Insulin Secretion. Time efficiency and cost were also analyzed. The overall quality of the islets obtained both by Histopaque and filtration was good. Filtration saved almost 90 % of the time consumed by Histopaque purification, and was also cheaper. However, one-third of the islets were lost. Since human and rodent islets share similar size but different density, filtration appears as a purification method with potential interest in translation to clinic.
胰岛移植是治疗 1 型糖尿病的一种有前途的疗法。多年来,在啮齿动物和人类胰岛分离中,用于胰岛纯化的全球方法都是基于菲可(Ficoll)的密度梯度,如 Histopaque。然而,当需要大规模分离时,在人员有限的实验室中,很难纯化胰岛。我们假设过滤可能是一种更简单、更快的替代方法,可以获得高质量的胰岛。每种方法进行了 4 次独立的胰岛分离,比较了过滤和 Histopaque 纯化与手工挑选作为胰岛纯度的金标准方法。评估了不同的质量参数:每个胰腺的胰岛数量、二噻嗪染色的纯度、荧光素二乙酸酯/碘化丙啶活细胞染色的活力以及通过葡萄糖刺激胰岛素分泌评估的体外功能。还分析了时间效率和成本。通过 Histopaque 和过滤获得的胰岛的整体质量都很好。过滤节省了 Histopaque 纯化所消耗时间的近 90%,而且成本更低。然而,三分之一的胰岛丢失了。由于人和啮齿动物的胰岛大小相似但密度不同,过滤似乎是一种具有潜在临床转化价值的纯化方法。