Areal density measurement is a convenient method for the determination of porcine islet equivalents without counting and sizing individual islets
Areal density measurement is a convenient method for the determination of porcine islet equivalents without counting and sizing individual islets
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DOI：
10.3727/000000003783985214
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发表时间：
2003-01-01
影响因子：
3.3
通讯作者：
Ammon, HPT
中科院分区：
文献类型：
--
作者：
Lembert, N;Wesche, J;Ammon, HPT
胰岛质量的测定对于实验室胰岛实验的标准化以及用于移植的胰岛的精确剂量非常重要。常见的微观分析基于单个胰岛的大小、频率分布的计算以及胰岛当量 (IEQ) 的转换，IEQ 是直径为 150 克的球形胰岛的体积。然而，胰岛的形状不规则，这使得该确定依赖于用户，并且一旦丢弃原始样本，分析就不可重复。将这种胰岛定量的常规技术与面密度测量的分析进行了比较。假设胰岛占据的整个面积可以用 IEQ 表示，而无需对单个胰岛进行大小调整和计数。通过连续消化/过滤分离猪胰岛并通过梯度离心纯化。纯化的胰岛用双硫腙染色，并在显微镜下随机选择区域重复拍照。从 11 种不同的纯化中总共拍摄了 51 张照片，并通过数字图像分析检测染色的胰岛。两项分析之间的相关系数 (r) 为 0.977，通过面密度检测低估了胰岛产量（斜率：0.75 +/- 0.03）。每张图片的面密度分析大约需要 1 分钟，比传统方法快约 10 倍，且不会增加方法误差（CV 2.1% vs. 2.7%）。总之，面密度测量可以快速、可重复地估计 IEQ，而无需计算单个胰岛。它可以在无需计算机编程的情况下进行单步分析，对于移植前在线测定胰岛产量非常有价值。
The determination of islet mass is important for the normalization of islet experiments in the laboratory and for the precise dosing of islets for transplantation. The common microscopical analysis is based on individual islet sizing, calculation of the frequency distribution, and conversion into islet equivalents (IEQ), which is the volume of a spherical islet with a diameter of 150 gm. However, islets are of irregular form, which makes this determination user dependent, and the analysis is irreproducible once the original sample is discarded. This routine technique of islet quantification was compared with the analysis of areal density measurements. It was assumed that the entire area occupied by islets can be expressed in IEQ without sizing and counting individual islets. Porcine islets were isolated by continuous digestion/filtration and purified by gradient centrifugation. Purified islets were stained with dithizone and were repeatedly pictured under the microscope with random area selection. A total of 51 pictures was taken from 11 different purifications and stained islets were detected by digital image analysis. The correlation coefficient (r) between both analyses was 0.977 with an underestimation of islet yield by areal density detection (slope: 0.75 +/- 0.03). Areal density analysis per picture took about 1 min, which is about 10 times faster than the traditional method without increasing the method error (CV 2.1% vs. 2.7%). In summary, areal density measurements allow a rapid and reproducible estimation of IEQ without counting individual islets. It can be performed in a single step analysis without computer programming and is valuable for online determinations of islet yield preceding transplantation.