UID:
almahu_9949697704202882
Umfang:
1 online resource (613 p.)
ISBN:
1-281-03829-6
,
9786611038298
,
0-08-055138-6
Serie:
Methods in enzymology ; v. 426
Inhalt:
An integrin, or integrin receptor, is an integral membrane protein in the plasma membrane of cells. It plays a role in the attachment of a cell to the extracellular matrix (ECM) and to other cells, and in signal transduction from the ECM to the cell. There are many types of integrin, and many cells have multiple types on their surface. Integrins are of vital importance to all metazoans, from humans to sponges. This volume in Methods in Enzymology presents methods for studying integrins.
Anmerkung:
Description based upon print version of record.
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Cover; Contents; Contributors; Preface; Volumes in Series; Chapter 1: Quantitative Measurements of Integrin-Mediated Adhesion to Extracellular Matrix; 1. Introduction; 2. Spinning Disc Measurements; 3. Protocols; 3.1. Preparation of the substrate; 3.2. Preparation of the cells; 3.3. Setup of the spinning disc device; 3.4. Counting the discs; 3.5. Generating adhesion plots using SigmaPlot; 4. Data Analysis; 4.1. Interpretation and sources of error; 4.2. Quantification of other parameters; 4.3. Equilibrium and rates; 4.4. Making comparisons; Acknowledgments; Appendix 1. Reagents
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Appendix 2. Spinning Disc DeviceReferences; Chapter 2: Investigating Integrin Regulation and Signaling Events in Three-Dimensional Systems; 1. Introduction; 1.1. Types of 3D matrices; 1.2. Challenges of working in 3D matrices; 2. Collagen Matrices of Different Densities; 3. Gel Contraction as a Measure to Quantify Cell Contractility; 4. Rho Activity Assay from Cells Cultured in 3D Collagen Gels; 4.1. Cell culture; 4.2. G-LISA assay; 5. Co-Immunoprecipitation of Integrin-Associated Proteins from Cells Cultured in 3D Collagen Gels; 6. beta1-Integrin Endocytosis Assays
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6.1. Time-lapse imaging of integrin trafficking in cells cultured on 2D surfaces6.2. Endosome protection biotinylation assay; 7. Imaging Cell-Matrix Interactions in 3D Collagen Gels; 7.1. Cell culture; 7.2. Stable transfection; 7.3. Transient transfection; 7.4. Three-dimensional collagen gels; 7.5. Nonlinear optical imaging of collagen-3D matrix adhesion interaction; References; Chapter 3: Integrins in Cell Migration; 1. Introduction; 2. Methods for Analysis of Integrin-Mediated Cell Migration; 2.1. Imaging of adhesion dynamics; 2.2. Imaging of podosomes; References
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Chapter 4: Integrin Cytoskeletal Interactions1. Introduction; 2. Models of Integrin Cytoplasmic Tails; 3. Expression Constructs for Integrin Tail Model Proteins; 4. Purification of Integrin Cytoplasmic Tails; 5. Preparation of the Affinity Matrix; 6. Binding Assays Using Cell Lysates; 6.1. Cell lysis; 6.2. Binding assays; 7. Direct Protein-Protein Binding Assays; 7.1. Production of GST-fusion proteins; 7.2. Binding assays with purified GST-fusion proteins; 8. Advanced Applications; 8.1. Competition assays; 8.2. Investigation of the role of integrin phosphorylation
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8.3. ELISA using model tail proteins8.4. Surface plasmon resonance assays; 8.5. Structural studies of model proteins; 9. Concluding Remarks; Acknowledgments; References; Chapter 5: Cell Survival in a Three-Dimensional Matrix; 1. Introduction; 2. Protocols; 2.1. Isolation of matched cell lines expressing differing levels of integrins; 2.2. Preparation of three-dimensional collagen culture; 2.3. Scoring apoptosis in collagen gels; 2.4. Immunofluorescence techniques for assessing cell survival; 2.5. Determining cell survival by caspase activation
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2.6. Adaptation of three-dimensional culture to existing immunodetection methods
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English
Weitere Ausg.:
ISBN 0-12-373924-1
Sprache:
Englisch
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