Chemicon human plasma fibronectin purified protein fibronectin solution

All experiments were carried out in Cellvis 96 well plates (#P96–1.5P). Briefly, wells were coated with 50uL of MilliporeSigma^™ Chemicon^™ Human Plasma Fibronectin Purified Protein fibronectin solution diluted 100x in PBS and were incubated at 37 °C for 30 min. HEK 293T cells were seeded in wells at a density of 35,000 cells/well in 100 μL and were spun down at 20 × g for 1 minute. In experiments requiring starvation (for all experiments involving SOS and EGFR constructs), after 24 hr, cells were starved by performing 7 80% washes with starvation media (DMEM + 1% P/S). Experiments were performed after 3 hr of starvation.

All experiments were carried out in Falcon^™ 353219 Black 96 well plates. Briefly, wells were coated with 50 uL of MilliporeSigma^™ Chemicon^™ Human Plasma Fibronectin Purified Protein fibronectin solution diluted 100x in PBS and were incubated at 37 °C for 30 min. HeLa or HEK 293T cells were seeded in wells at a density of 60,000 cells/well in 100 μL and were spun down at 20 × g for 1 minute to ensure an even distribution of cells across the wells. For imaging ELPs, HEK 293T cells were transfected with GFP-ELP₅₃ and H2B-iRFP 12–24 hours post-seeding in 96 well plates using the calcium phosphate method. Transfection mix/media was removed 24 hours post transfection and replaced with fresh media. Cells were imaged within 4–8 hours of this media change.