A Crossed Laser Phase Plate for CryoEM

Yu, Y.; Cheng, A.; Montabana, E.; Paz Soldan, N.; Cooper, E. S.; Zhang, J. T.; Axelrod, J. J.; Petrov, P. N.; Maisenbacher, L.; Potter, C. S.; Mueller, H.; Carragher, B.; Agard, D.; Olshin, P. K.

doi:10.64898/2026.06.05.730245

bioRxiv · 10.64898/2026.06.05.730245

A Crossed Laser Phase Plate for CryoEM

Yu, Y.·Cheng, A.·Montabana, E.·Paz Soldan, N.·Cooper, E. S.·Zhang, J. T.·Axelrod, J. J.·Petrov, P. N.·Maisenbacher, L.·Potter, C. S.·Mueller, H.·Carragher, B.·Agard, D.·Olshin, P. K.

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Abstract

The laser phase plate (LPP) enables phase-contrast imaging in cryogenic electron microscopy (cryoEM), enhancing image contrast without compromising high-resolution information. Here we report the implementation of a crossed laser phase plate (xLPP) comprising two optical cavities oriented orthogonally, installed in a ThermoFisher Scientific Krios G4 microscope equipped with a newly designed transfer lens module. We demonstrate the expected, strong contrast enhancement and stable, additive phase shifts of 90{degrees}, with a contrast transfer function (CTF) that closely matches theory. Single-particle analysis (SPA) of apoferritin, a standard benchmark sample, reached a resolution of 1.79 [A], demonstrating the system is capable of acquiring high-resolution cryoEM data. When imaging thick E. coli cells ([~]350 nm), the xLPP enhances contrast and increases low-frequency template-matching signal. Together, these results establish the feasibility of the xLPP and highlight its potential for high-contrast, high-resolution cryoEM imaging of biological systems.

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Yu, Y., Cheng, A., Montabana, E., Paz Soldan, N., Cooper, E. S., Zhang, J. T., Axelrod, J. J., Petrov, P. N., Maisenbacher, L., Potter, C. S., Mueller, H., Carragher, B., Agard, D., Olshin, P. K.. 2026-06-05. A Crossed Laser Phase Plate for CryoEM. https://doi.org/10.64898/2026.06.05.730245

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