Robertson G S, Chadwick D R, Contractor H, James R F, London N J
Department of Surgery, Leicester Royal Infirmary, UK.
Acta Diabetol. 1993;30(2):93-8. doi: 10.1007/BF00578221.
The use of the COBE 2991 cell processor (COBE Laboratories, Colorado) for large-scale islet purification using discontinuous density gradients has been widely adopted. It minimizes many of the problems such as wall effects, normally encountered during centrifugation, and avoids the vortexing at interfaces that occurs during acceleration and deceleration by allowing the gradient to be formed and the islet-containing interface to be collected while continuing to spin. We have produced cross-sectional profiles of the 2991 bag during spinning which allow the area of interfaces in such step gradients to be calculated. This allows the volumes of the gradient media layers loaded on the machine to be adjusted in order to maximize the area of the gradient interfaces. However, even using the maximal areas possible (144.5 cm2), clogging of tissue at such interfaces limits the volume of digest which can be separated on one gradient to 15 ml. We have shown that a linear continuous density gradient can be produced within the 2991 bag, that allows as much as 40 ml of digest to be successfully purified. Such a system combines the intrinsic advantages of the 2991 with those of continuous density gradients and provides the optimal method for density-dependent islet purification.
使用COBE 2991细胞处理器（COBE实验室，科罗拉多州）通过不连续密度梯度进行大规模胰岛纯化已被广泛采用。它最大限度地减少了许多诸如壁效应等通常在离心过程中遇到的问题，并且通过在继续旋转的同时允许形成梯度并收集含胰岛的界面，避免了加速和减速过程中在界面处出现的涡旋现象。我们已经制作了2991袋在旋转过程中的横截面轮廓，这使得能够计算这种阶梯梯度中界面的面积。这允许调整加载到机器上的梯度介质层的体积，以便最大化梯度界面的面积。然而，即使使用可能的最大面积（144.5平方厘米），在这样的界面处组织堵塞仍将在一个梯度上可分离的消化液体积限制为15毫升。我们已经表明，可以在2991袋内产生线性连续密度梯度，这允许多达40毫升的消化液被成功纯化。这样的系统将2991的固有优势与连续密度梯度的优势结合起来，并为依赖密度的胰岛纯化提供了最佳方法。
