He, Kuan; Huang, Xiang; Wang, Xiaolei; Yoo, Seunghwan; Ruiz, Pablo; Gdor, Itay; Ferrier, Nicola; Scherer, Norbert; Hereld, Mark; Katsaggelos, Aggelos; Cossairt, Oliver
Realizing both high temporal and spatial resolution across a large volume is a key challenge for 3D fluorescent imaging. Towards achieving this objective, we introduce an interferometric multifocus microscopy (iMFM) system, a combination of multifocus microscopy (MFM) with two opposing objective lenses. We show that the proposed iMFM is capable of simultaneously producing multiple focal plane interferometry that provides axial super-resolution and hence isotropic 3D resolution with a single exposure. We design and simulate the iMFM microscope by employing two special diffractive optical elements. The point spread function of this new iMFM microscope is simulated and the image formation model is given. For reconstruction, we use the Richardson-Lucy deconvolution algorithm with total variation regularization for 3D extended object recovery, and a maximum likelihood estimator (MLE) for single molecule tracking. A method for determining an initial axial position of the molecule is also proposed to improve the convergence of the MLE. We demonstrate both theoretically and numerically that isotropic 3D nanoscopic localization accuracy is achievable with an axial imaging range of 2um when tracking a fluorescent molecule in three dimensions and that the diffraction limited axial resolution can be improved by 3-4 times in the single shot wide-field 3D extended object recovery. We believe that iMFM will be a useful tool in 3D dynamic event imaging that requires both high temporal and spatial resolution.