Brandsch C, Friedl P, Lange K, Richter T, Mothes T
Institute of Clinical Chemistry and Pathobiochemistry, Medical Hospital II, Children's Hospital of the University of Leipzig, Germany.
Scand J Gastroenterol. 1998 Aug;33(8):833-8. doi: 10.1080/00365529850171495.
So far, no techniques are available for primary culture and efficient transfection of human small-intestinal enterocytes, which would provide a valuable tool to investigate intestinal function.
Human small-intestinal biopsy specimens were treated with collagenase and dispase. Resulting crypt units were cultured for several days. Using the intestinal epithelial cell lines Caco-2 and HT-29, we established optimal conditions for transfection of a control plasmid, which were then applied to primary cultured cells.
Cells growing out of crypt units formed monolayer-like sheets and proliferated for several days. Most of the cells could be stained with antibodies against epithelial markers. Among seven different transfection reagents tested, Lipofectamine was the most potent, with transfection efficiencies up to 25% for primary enterocytes.
An easy technique was developed providing viable small-intestinal enterocytes that can be efficiently transfected.
迄今为止，尚无用于人小肠肠上皮细胞原代培养和高效转染的技术，而这将为研究肠道功能提供一个有价值的工具。
用人小肠活检标本进行胶原酶和中性蛋白酶处理。将得到的隐窝单位培养数天。利用肠上皮细胞系Caco-2和HT-29，我们建立了对照质粒转染的最佳条件，然后将其应用于原代培养细胞。
从隐窝单位长出的细胞形成单层样薄片并增殖数天。大多数细胞可用抗上皮标志物的抗体染色。在测试的七种不同转染试剂中，Lipofectamine最有效，原代肠上皮细胞的转染效率高达25%。
开发了一种简单的技术，可提供能够高效转染的活的小肠肠上皮细胞。
