Mode of adsorption and orientation of an extracellular matrix protein affect its cell-adhesion-promoting activity

Cell adhesion to extracellular matrix contributes to the organization of ti ssues and modulates cell behavior. In conventional cell adhesion assays, pl astic wells are coated with matrix proteins and assayed for their adhesion- promoting activity. We show here that factors such as sample composition, c oating buffers, and manufacturers' plastic treatment markedly affect cell a dhesion to the extracellular matrix protein laminin-5 (Ln-5). These factors were shown to affect adsorption efficiency as determined by measuring tota l adsorbed protein with a polyclonal anti-Ln-5 antiserum. They also influen ce the availability of the epitope for an adhesion-blocking anti-Ln-5 monoc lonal antibody, suggesting that coating conditions affect the orientation o f Ln-5. Generally. cell, adhesion correlates more strongly with the availab ility of the epitope for the adhesion-blocking antibody than with total ads orbed Ln-5. Our data further indicate that cell adhesion to other matrix pr oteins may be influenced by similar factors. Adding Ln-5 samples to plastic wells that had been precoated with non-adhesion-blocking anti-Ln-5 antibod ies made cell adhesion independent of factors such as sample composition, c oating buffers, and source of plastic. Thus, the control of adsorption effi ciency and orientation of extracellular matrix proteins is essential for cr eation of reliable and reproducible conditions in cell adhesion assays. (C) 1998 Academic Press.