Aliasing mitigation in optical microscopy of dynamic biological samples by use of temporally modulated color illumination and a standard RGB camera

Significance: Despite recent developments in microscopy, temporal aliasing can arise when imaging dynamic samples. Modern sampling frameworks, such as generalized sampling, mitigate aliasing but require measurement of temporally overlapping and potentially negative-valued inner products. Conventional cameras cannot collect these directly as they operate sequentially and are only sensitive to light intensity. Aim: We aim to mitigate aliasing in microscopy of dynamic monochrome samples by implementing generalized sampling via the use of a color camera and modulated color illumination. Approach: We solve the overlap problem by spectrally multiplexing the acquisitions and using (positive) B-spline segments as projection kernels. Reconstruction involves spectral unmixing and inverse filtering. We implemented this method using a color LED illuminator. We evaluated its performance by imaging a rotating grid and its applicability by imaging the beating zebrafish embryo heart in transmission and light-sheet microscopes. Results: Compared to stroboscopic imaging, our method mitigates aliasing with performance improving as the projection order increases. The approach can be implemented in conventional microscopes but is limited by the number of available LED colors and camera channels. Conclusions: Generalized sampling can be implemented via color modulation in microscopy to mitigate temporal aliasing. The simple hardware requirements could make it applicable to other optical imaging modalities.