Quantifying cellular adhesion to extracellular matrix components by single-cell force spectroscopy

• Published in: Nature protocols Vol. 5; no. 7; pp. 1353 - 1361
• Main Authors: Muller, Daniel J, Friedrichs, Jens, Helenius, Jonne
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 01-07-2010; Nature Publishing Group
• Subjects: 631/1647/328/1262; 631/80/79/750; Adhesion; Adhesives; Analytical Chemistry; Atomic force microscopy; Biological Techniques; Biomedical and Life Sciences; Cell adhesion; Cell adhesion & migration; Cell Adhesion - physiology; Cell research; Collagen; Collagen Type I; Computational Biology/Bioinformatics; Extracellular matrix; Extracellular Matrix - metabolism; Extracellular Matrix - physiology; Extracellular Matrix - ultrastructure; HeLa Cells; Humans; Integrins - metabolism; Integrins - ultrastructure; Life Sciences; Ligands; Methods; Microarrays; Microscopy; Microscopy, Atomic Force - instrumentation; Microscopy, Atomic Force - methods; Observations; Organic Chemistry; Physiology; Probes; Protein Binding - physiology; Proteins; protocol; Quantitative research; Spectroscopy; Spectrum analysis; Germany
• ISSN: 1754-2189; 1750-2799
• DOI: 10.1038/nprot.2010.89

Atomic force microscopy (AFM)-based single-cell force spectroscopy (SCFS) enables the quantitative study of cell adhesion under physiological conditions. SCFS probes adhesive interactions of single living cells with substrates such as extracellular matrix (ECM) proteins and other cells. Here we present a protocol to study integrin-mediated adhesion of HeLa cells to collagen type I using SCFS. We describe procedures for (i) functionalization of AFM cantilevers with the lectin concanavalin A and supports with collagen, (ii) cell handling and attachment to the AFM cantilever, (iii) measurement of adhesion forces and (iv) data analysis and interpretation. Although designed to measure HeLa cell adhesion to collagen, the protocol can be modified for other cell lines and ECM proteins. Compared with other SCFS assays (for example, optical tweezer, biomembrane force probe), AFM-based SCFS has a more versatile force detection range, and it can therefore be used to address a broader range of biological questions. The protocol can be completed in 2–3 d.