Micro-printing of molecular patterns on deformable substrate for the study of cell adhesion and traction forces

MARÍA CLAUDIA MARCHI; PRISCILA COLOMBO; LORENA SIGAUT; GONZALO GIORDANO; LÍA PIETRASANTA

Rosario

Congreso; L Reunión Anual de la Sociedad Argentina de Biofísica; 2022

Sociedad Argentina de Biofísica

Micropatterning techniques allow direct control over the spatial organization of molecules at the micrometric scale and provide a powerful tool to study biological processes such as migration, differentiation, and proliferation. In these processes, cellular responses are influenced by the composition and spatial distribution of extracellular proteins as well as mechanical properties of the substrate. The aim of this project is to generate molecular micropatterns over deformable substrates of tunable stiffness. Such substrates will provide a controlled microenvironment –adhesive micropattern surface and adjustable mechanical properties– to study cell behavior, control cell adhesion and will also enable to quantify cellular forces by traction force microscopy1.To generate micropatterns, we use a microcontact printing technique2 that consists of transferring molecules from an elastomeric stamp to a substrate by conformal contact between them. As deformable substrate, we employ polyacryilamide hydrogels due to their mechanical tunability, elastic material behavior and optical translucency. Here, we present the different stages needed to obtain deformable substrates with a molecular micropatterned surface that include: the design, fabrication and characterization of the master for the elastomeric stamps by electron beam lithography3, together with deformable substrate fabrication and microcontact printing optimization. Briefly, the designed micropatterns are written employing the electron beam from a scanning electron microscope on a glass or silicon wafer covered by a polymethylmethacrylate (PMMA) film generated by spin coating. We characterize the thickness of the film that determines the height of the relief patterns. The electron dose is optimized for the adequate writing of micropatterns on PMMA. Once the master is obtained, the polydimethylsiloxane (PDMS) stamp is fabricated. To optimized microcontact, the PDMS stamp is incubated with fluorescent molecules, and printed on glass or on polyacryilamide hydrogels. The obtained micropatterns are visualized by confocal microscopy. We found that reliable and reproducible printing of the designed micropatterns are achieved in both glass and polyacryilamide substrates.