ADSORPTION AND ELUTION OF EXTRACELLULAR-MATRIX PROTEINS ON NONTISSUE CULTURE POLYSTYRENE PETRI DISHES

We have devised a simple procedure for directly measuring the adsorption of fibrinogen, type IV collagen, and fibronectin on non-tissue culture polystyrene petri dishes using proteins labeled with 125I and removable dish sections. Our approach provides a rapid method for obtaining quantitative adsorption isotherms for these large extracellular matrix (ECM) proteins in situations applicable for studies of cell culture and cell adhesion migration. We found that the surface density of adsorbed ECM protein depends not only on solution protein concentration but also on ECM protein type, with roughly one and two orders of magnitude more fibronectin and type IV collagen, respectively, adsorbing at an equivalent solution concentration compared to fibrinogen. Adsorption isotherms for each of these proteins were compared with theoretical bounds for monolayer density based on random sequential adsorption and molecular close-packing. We also observed that exposure to 1% sodium dodecyl sulfate in 3 M NaOH for 25 h was effective at eluting fibrinogen from our dishes over a wide range of protein concentration, but that this same detergent treatment was ineffective at completely desorbing type IV collagen adsorbed at high density. Our results, useful to researchers examining the role of substratum-bound molecules in the control of cell behavior, demonstrate that estimates of the adsorbed molecular density of ECM proteins obtained from indirect methods, such as elution and enzyme-linked immunosorbent assays, or inferred from solution coating concentrations may provide erroneous estimates of the true number of molecules actually adsorbed. © 1992.