*2.5. Cell Alignment Quantification*

The cell alignment on the NFM was quantified by a two-dimensional fast Fourier transform (2-D FFT) image analysis method [42]. Briefly, the fluorescent cell photographs were converted to 8-bit grayscale and then cropped a 1024 × 1024 pixel square. This was then masked with a transparent circular pattern, and the corners were filled with black. The processed photograph was analyzed by FFT function in ImageJ software [43]. Pixel intensities along the radian were summed using the oval profile plug-in and normalized by the lowest intensity value [44]. The normalized results represent the percentage of cells aligned along a certain direction.
