The cell spreading assay was performed as described previously (65). Briefly, MEFs or C33A cells at a density of 1 × 105 cells/ml were seeded onto coverslips that were coated with FN and blocked with BSA, as described in the adhesion assay, for 0 to 1 hour at 37°C. Cells were stained with the primary antibody recognizing the focal adhesion protein paxillin, followed by Alexa 488–conjugated secondary antibody. The relative area of spreading was determined by measuring the area circumscribed by paxillin staining and quantitated using Fiji software. Cells of similar nuclear size were used for measurement.

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