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Targeting hepatocytes from liver tissue by laser capture microdissection and proteomics expression profiling
Author(s) -
MarkoVarga György,
Berglund Magnus,
Malmström Johan,
Lindberg Henrik,
Fehniger Thomas E.
Publication year - 2003
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.200305645
Subject(s) - laser capture microdissection , proteome , proteomics , microdissection , liver tissue , gel electrophoresis , two dimensional gel electrophoresis , chromatography , biology , microbiology and biotechnology , protein purification , hepatocyte , chemistry , gene expression , biochemistry , in vitro , gene , endocrinology
Abstract A tissue proteomics process is presented where hepatocyte cell isolation in combination with two‐dimensional (2‐D) gel electrophoresis and mass spectrometric identification were used to annotate the liver proteome. Laser microdissection of 8 μm liver tissue sections was performed and protein expression profiling was compared using a variety of quantities of input cells, and gel separation conditions. The 30 μm diameter laser generated the highest protein yields from the polymer coated caps following microsolubilization. We found that 6000 laser pulses (approximately 7200 hepatocytes) were required in order to generate high‐resolution gel maps. Within homogeneous tissue samples, this could be accomplished in a total cycle time of 20 min using an automated dissection procedure. Close to 1000 high‐quality gel annotations were generated from the corresponding 2‐D gel expression profiles which matched closely the corresponding patterns of analytical‐scale liver preparations detected by silver staining.