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bioRxiv · 10.1101/2024.07.18.604091

Vortex light field microscopy: 3D spectral single-molecule imaging with a twist

Zhang, B.·Daly, S.·Zhu, C.·Lenz, M. O.·Weiss, L. E.·Needham, L.-M.·Peters, R.·Lee, S.·O'Holleran, K.
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Abstract

We introduce vortex light field microscopy (VLFM), a novel method for snapshot 3D spectral single-molecule localization microscopy. Inspired by the azimuthal phase profile of optical vortices, we place an azimuthally oriented prism array immediately after the microlens array in a Fourier light field microscope (FLFM). This innovative arrangement causes the axial position and spectral peak for a point emitter to be encoded in the radial and azimuthal displacement of point-spread-function (PSF) respectively. This enables simultaneous detection of 3D position and emission peak of individual fluorophores with 25 nm spatial precision and 3 nm spectral precision over a 4 m depth of field (DOF). We illustrate the spectral scalability of our method by performing four-color 3D single particle tracking of freely diffusing fluorescent beads, and two-color 3D dSTORM imaging of microtubules and mitochondria in fixed COS-7 cells, without the need for spectrally distinct fluorophores.

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Zhang, B., Daly, S., Zhu, C., Lenz, M. O., Weiss, L. E., Needham, L.-M., Peters, R., Lee, S., O'Holleran, K.. 2024-07-22. Vortex light field microscopy: 3D spectral single-molecule imaging with a twist. https://doi.org/10.1101/2024.07.18.604091

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