Marselli Lorella, Sgroi Dennis C, Bonner-Weir Susan, Weir Gordon C
Section on Islet Transplantation and Cell Biology, Joslin Diabetes Center, One Joslin Place, Boston, MA 02215, USA.
Methods Mol Biol. 2009;560:87-98. doi: 10.1007/978-1-59745-448-3_8.
Human beta-cell gene profiling is a powerful tool for understanding beta-cell biology in normal and pathological conditions. The assessment is complicated when isolated islets are studied because of contamination by non-beta-cells and the exposure to the trauma of isolation that causes changes in gene expression. These limitations can be overcome by dissecting the beta-cells from the pancreatic tissue directly using the laser capture microdissection (LCM) technique. LCM allows the sampling of specific cell types from tissue sections. The technique requires morphological criteria or specific stains for targeted cells, and the protocols must preserve the condition of the sought-after macromolecules. We have developed a protocol of rapid tissue dehydration followed by identification of human beta-cells by their intrinsic autofluorescence, which allows laser microdissection for gene-profiling studies.
人类β细胞基因谱分析是了解正常和病理条件下β细胞生物学的有力工具。当研究分离的胰岛时，评估会变得复杂，因为存在非β细胞的污染以及分离创伤导致基因表达变化。通过使用激光捕获显微切割（LCM）技术直接从胰腺组织中解剖出β细胞，可以克服这些局限性。LCM允许从组织切片中采样特定细胞类型。该技术需要针对目标细胞的形态学标准或特定染色，并且方案必须保留所需大分子的状态。我们已经开发了一种快速组织脱水方案，随后通过其内在自发荧光识别人类β细胞，这使得能够进行激光显微切割以进行基因谱研究。
