Spatially resolved microrheology of heterogeneous biopolymer hydrogels using covalently bound microspheres.

Characterization of the rheological properties of heterogeneous biopolymers is important not only to understand the effect of substrate elasticity on cell behaviors, but also to provide insights into mechanical changes during cellular remodeling of the environment. Conventional particle-tracking microrheology (PTM) techniques are compromised by probe-network slippage and cage-hopping problems, and require a priori knowledge of network mesh size in order to determine a suitable probe size. We demonstrated here the usefulness of covalently bound probes for PTM of biopolymers to overcome the above limitations. We showed that, in a well-defined system like polyacrylamide gels, surface-modified probe particles using a zero-length crosslinker provided more reliable measurements of network mechanics as compared to standard carboxylated probes. We further demonstrated that appropriate surface modification of microspheres for PTM circumvented the requirement of using microspheres larger than the network mesh, an approach typically considered to be ideal. Using the method presented in this study, we found the local network at the leading edge of a typical C6 glioma cell to be stiffer as compared to the side. Our findings established that permanent interaction between the probe and network is crucial to reliably measure the local network mechanics in reconstituted, heterogeneous networks using PTM.