期刊
TRAFFIC
卷 12, 期 7, 页码 806-814出版社
WILEY-BLACKWELL
DOI: 10.1111/j.1600-0854.2011.01199.x
关键词
correlative microscopy; high-pressure freezing; HUVEC; iLEM; immuno-transmission electron microscopy; lipid; MDCK-II; section fixation; THP-1; VIS2FIX
类别
资金
- Technology Foundation Stichting Technologische Wetenschappen
- Innovatiegerichte Onderzoekprogramma's Genomics (IOP Genomics, SenterNovem)
- Dutch FSHD Foundation
Immuno-transmission electron microscopy (TEM) is the technique of choice for high-resolution localization of proteins in fixed specimen. Here we introduce 2 novel methods for the fixation of sections from cryo-immobilized samples that result in excellent ultrastructural preservation. These high-speed fixation techniques, both called VIS2FIX, allow for a reduction in sample preparation time from at least 1 week to only 8 h. The methods were validated in immuno-TEM experiments on THP-1 monocytes, human umbilical vein endothelial cells (HUVECs) and Madin-Darby canine kidney (MDCK-II) cells. The fixation and retention of neutral lipids is demonstrated, offering unique prospects for the application of immuno-TEM in the lipidomics field. Furthermore, the VIS2FIX methods were successfully employed in correlative fluorescence and electron microscopy.
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