title: Subcellular protein extraction from human pancreatic cancer tissues.
abstract: Proteins are the major class of effector molecules in cellular systems. For the identification of functional differences between normal and diseased tissues, a reliable analysis of their protein content is essential. Reproducible isolation and fractionation of intact proteins are important in this respect, but their complexity in structure and concentration, their close interaction, and their instability represent major challenges. For protein isolation in tissues, the breakdown of cell-cell and cell-matrix connections within a tissue without affecting protein quality is a critical factor. We compared different processes for a compartmental protein preparation from pancreatic tissue, one of the most challenging tissues for protein isolation because of its high protease content. Success of the different procedures varied greatly. Based on a scheme of tissue-slicing and subsequent cell isolation, we established a reliable workflow for the fractional extraction of cytosolic proteins, membrane and organelle proteins, nuclear proteins, and cytoskeletal filaments. The tissue slices also allow for a representative confirmation of individual samples' cellular status by histochemical processes, and a proper separation or mixing of cellular material from across a tumor if required.
authors.affiliation: Functional Genome Analysis, Deutsches Krebsforschungszentrum, Heidelberg, Germany. a.boerner@dkfz.de
authors.forename: Anette
authors.initials: A
authors.lastname: Börner
authors.forename: Uwe
authors.initials: U
authors.lastname: Warnken
authors.forename: Martina
authors.initials: M
authors.lastname: Schnölzer
authors.forename: Jörg von
authors.initials: Jv
authors.lastname: Hagen
authors.forename: Nathalia
authors.initials: N
authors.lastname: Giese
authors.forename: Andrea
authors.initials: A
authors.lastname: Bauer
authors.forename: Jörg
authors.initials: J
authors.lastname: Hoheisel
mesh_terms: D000818:Animals; D002458:Cell Fractionation / Q000379:methods*; D045744:Cell Line, Tumor; D017364:Collagenases / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional; D004591:Electrophoresis, Polyacrylamide Gel; D016591:Histocytological Preparation Techniques; D006801:Humans; D009363:Neoplasm Proteins / Q000032:analysis / Q000302:isolation & purification*; D010190:Pancreatic Neoplasms / Q000737:chemistry* / Q000473:pathology; D051381:Rats; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization; D013347:Subcellular Fractions / Q000737:chemistry*; D013552:Swine
chemical_list: D009363:Neoplasm Proteins; D017364:Collagenases