Abbott Sarah K, Jenner Andrew M, Mitchell Todd W, Brown Simon H J, Halliday Glenda M, Garner Brett
Illawarra Health and Medical Research Institute, University of Wollongong, Wollongong, NSW 2522, Australia.
Lipids. 2013 Mar;48(3):307-18. doi: 10.1007/s11745-013-3760-z. Epub 2013 Jan 26.
We have developed a protocol suitable for high-throughput lipidomic analysis of human brain samples. The traditional Folch extraction (using chloroform and glass-glass homogenization) was compared to a high-throughput method combining methyl-tert-butyl ether (MTBE) extraction with mechanical homogenization utilizing ceramic beads. This high-throughput method significantly reduced sample handling time and increased efficiency compared to glass-glass homogenizing. Furthermore, replacing chloroform with MTBE is safer (less carcinogenic/toxic), with lipids dissolving in the upper phase, allowing for easier pipetting and the potential for automation (i.e., robotics). Both methods were applied to the analysis of human occipital cortex. Lipid species (including ceramides, sphingomyelins, choline glycerophospholipids, ethanolamine glycerophospholipids and phosphatidylserines) were analyzed via electrospray ionization mass spectrometry and sterol species were analyzed using gas chromatography mass spectrometry. No differences in lipid species composition were evident when the lipid extraction protocols were compared, indicating that MTBE extraction with mechanical bead homogenization provides an improved method for the lipidomic profiling of human brain tissue.
我们已经开发出一种适用于人脑样本高通量脂质组学分析的方案。将传统的 Folch 提取法（使用氯仿和玻璃-玻璃匀浆法）与一种高通量方法进行了比较，该高通量方法将甲基叔丁基醚（MTBE）提取与利用陶瓷珠的机械匀浆相结合。与玻璃-玻璃匀浆相比，这种高通量方法显著减少了样品处理时间并提高了效率。此外，用 MTBE 替代氯仿更安全（致癌性/毒性更低），脂质溶解在上层相中，便于移液操作且具有自动化（即机器人操作）的潜力。两种方法都应用于人枕叶皮质的分析。通过电喷雾电离质谱分析脂质种类（包括神经酰胺、鞘磷脂、胆碱甘油磷脂、乙醇胺甘油磷脂和磷脂酰丝氨酸），并使用气相色谱质谱分析甾醇种类。比较脂质提取方案时，脂质种类组成没有明显差异，这表明采用机械珠匀浆的 MTBE 提取法为脑组织脂质组分析提供了一种改进方法。
