ECM

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Question 1:  Can I coat cells with fluorescent ECM proteins?

Answer 1:  Yes, cells can be coated with fluorescently-labeled ECM proteins such as fibronectin.    Example protocols are found in Pallis et al., 1997 (Peripheral Blood Lymphocyte Binding to a Soluble FITC-Fibronectin Conjugate. Cytometry. 28, 157-164) and Huveneers et al., 2008 (Binding of soluble fibronectin to integrin a5b1 – link to focal adhesion redistribution and contractile shape. J. Cell Sci. 121, 2452-2462).  Briefly, cells are incubated with a low concentration of soluble fluorescently-labeled fibronectin in an appropriate buffer.  Low concentrations of fibronectin are recommended to minimize non-specific binding.  The cells/fibronectin mixture is incubated in the dark for 30 min at room temperature.  Others have done the incubation at 4°C for 1 hour.  The cells are then rinsed in an appropriate buffer, resuspended in the same buffer, and then the coating/binding of ECM proteins can be quantified by flow cytometry or the coated cells can be used experimentally.