title: Measurement of protein tyrosine phosphorylation in cell adhesion.
abstract: This chapter describes biochemical, immunochemical, and microscopic approaches to measure protein tyrosine phosphorylation after cell adhesion. We have outlined detailed procedures to biochemically examine the phosphotyrosine content of cellular proteins by Western blotting, which in some cases can be performed using phospho-specific antibodies. Furthermore, we have described in detail the examination of subcellular localization of phosphotyrosine-containing proteins in focal adhesions using immunofluorescence. Finally, a quantitative fluorescence microscopic technique using an SH2-containing phosphotyrosine reporter to monitor tyrosine phosphorylation in focal adhesions in live cells is described.
authors.affiliation: Department of Cell and Developmental Biology, University of North Carolina-Chapel Hill, Chapel Hill, NC, USA.
authors.forename: Vidhya V
authors.initials: VV
authors.lastname: Iyer
authors.forename: Christoph
authors.initials: C
authors.lastname: Ballestrem
authors.forename: Joachim
authors.initials: J
authors.lastname: Kirchner
authors.forename: Benjamin
authors.initials: B
authors.lastname: Geiger
authors.forename: Michael D
authors.initials: MD
authors.lastname: Schaller
mesh_terms: D016475:3T3 Cells; D000818:Animals; D002448:Cell Adhesion / Q000502:physiology*; D002642:Chick Embryo; D003598:Cytoskeletal Proteins / Q000378:metabolism; D005109:Extracellular Matrix / Q000502:physiology / Q000648:ultrastructure; D007150:Immunohistochemistry / Q000379:methods; D051379:Mice; D008856:Microscopy, Fluorescence / Q000379:methods; D051419:Paxillin; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism*
chemical_list: D003598:Cytoskeletal Proteins; D051419:Paxillin; D010750:Phosphoproteins; C496062:Pxn protein, mouse; D019000:Phosphotyrosine