Dynamic Microstructured Thermoresponsive Interfaces for Label-Free Cell Sorting Based on Nonspecific Interactions

Label-free cell sorting methods and materials are developed in this work. The microstructured thermoresponsive surfaces made of poly(glycidyl methacrylate) (PGMA) and poly(N-isopropylacrylamide-co-glycidyl methacrylate) (PNIPAM-co-GMA) are prepared by phase separation on the submicron scale in thin films and then cross-linked and covalently grafted to the substrate. PGMA domains are used for cell adhesion, while the PNIPAM-co-GMA matrix pushes cells off the surface at a temperature below the lower critical solution temperature (LCST). The microstructure formation and swelling-shrinking caused by changes in temperature are studied experimentally and by using dissipative particle dynamics computer simulations. Experiments with RAW 264.7 murine macrophage-like cells, NIH3T3/GFP murine fibroblasts, and HaCaT human skin keratinocytes (unlabeled and GFP-positive strains) demonstrate successful cell sorting based on weak and nonspecific interactions with reconfigurable thermoresponsive microstructured surfaces. Efficient sorting with a separation factor of >50 is achieved if the push-off force is adjusted to a level between the adhesive forces of the separating cells. This experimental finding is supported by Monte Carlo simulations of cell adsorption and detachment on the microstructured surfaces. The experiments and simulations show that efficient cell sorting is possible for weak to moderate cell adhesion to the surfaces. However, the method is not successful for very weak or very strong adhesion. We demonstrate that cell adhesion to the microstructured surfaces can be adjusted by changes in the conditions of the phase separation at the stage of film formation and by varying the incubation time of the cells on the microstructured surfaces.