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IntroductionPhotomodifiable azopolymer nanotopographies represent a powerful means of assessing how cells respond to rapid changes in the local microenvironment. However, previous studies have suggested that azopolymers are readily photomodified under typical fluorescence imaging conditions over much of the visible spectrum. Here we assess the stability of azopolymer nanoridges under 1-photon and 2-photon imaging over a broad range of wavelengths. MethodsAzopolymer nanoridges were created via microtransfer molding of master structures that were created using interference lithography. The effects of exposure to a broad range of wavelengths of light polarized parallel to the ridges were assessed on both a spinning-disk confocal microscope and a 2-photon fluorescence microscope. Experiments with liveDictyostelium discoideumcells were also performed using alternating cycles of 514-nm light for photomodification and 561-nm light for fluorescence imaging. Results and DiscussionWe find that for both 1-photon and 2-photon imaging, only a limited range of wavelengths of light leads to photomodification of the azopolymer nanotopography. These results indicate that nondestructive 1-photon and 2-photon fluorescence imaging can be performed over a considerably broader range of wavelengths than would be suggested by previous research. 

Abstract Nanotopographic surfaces are a powerful tool for studying and controlling cell behavior. However, the fabrication of nanotopographic master patterns using conventional photolithography is expensive, which limits the range of designs that can be explored. In this study, a method is demonstrated for the photoreshaping of large‐area patterns of nanoridges. The original master pattern is created using conventional lithography, and an azopolymer replica is prepared using soft lithography. The manipulation of the nanoridges is achieved by projecting light with specific polarizations and exposure times, resulting in controllable widening, buckling, or removal of the ridges. The reprogrammed azopolymer master patterns can then be replicated, creating reproducible new nanotopographies that can be transferred into other materials using a molding procedure. Diffraction can be used for in situ monitoring of the reprogramming during exposure. Image‐analysis methods are used to characterize buckled ridges as a function of exposure time. The response of MCF10A epithelial cells are investigated to buckled nanoridges. A substantial impact of buckling on the dynamics and location of actin polymerization, as well as on the distribution and lengths of contiguous polymerized regions is also observed.