Measuring Cytoplasmic Stiffness of Fibroblasts as a Function of Location and Substrate Rigidity Using Atomic Force Microscopy

The cytoplasmic stiffness of cells plays a significant role during cell migration. As a cell migrates, differences in cytoplasmic properties occur that subsequently modulate migratory behavior. The properties of the substrate to which cells are adherent also play a role. To accurately measure the cytoplasmic stiffness of cells, we provide detailed instructions on how to assemble hydrogels that exhibit different elastic moduli, culturing cells on these substrates followed by a step-by-step process to measure and analyze the cytoplasmic properties of fibroblasts. In this study, we have measured the elastic moduli of cells at different locations to demonstrate how this property varies as a function of where the measurement is performed. The degree of anisotropy measured by the difference between cytoplasmic stiffness at the two edges of the cell also varied as a function of the elasticity of their underlying substrates. Larger differences in cytoplasmic stiffness between the leading and trailing edges were observed on substrates with a higher elastic modulus. The methods reported in this study can provide information on cellular properties, specifically, how the elastic modulus of cells can be probed and analyzed in vitro.