Publications & technical resources

We demonstrate single-molecule fluorescence imaging beyond the optical diffraction limit in 3 dimensions with a wide-field microscope that exhibits a double-helix point spread function (DH-PSF). The DH-PSF design features high and uniform Fisher information and has 2 dominant lobes in the image plane whose angular orientation rotates with the axial (z) position of the emitter. Single fluorescent molecules in a thick polymer sample are localized in single 500-ms acquisitions with 10- to 20-nm precision over a large depth of field (2 μm) by finding the center of the 2 DH-PSF lobes. By using a photoactivatable fluorophore, repeated imaging of sparse subsets with a DH-PSF microscope provides superresolution imaging of high concentrations of molecules in all 3 dimensions. The combination of optical PSF design and digital postprocessing with photoactivatable fluorophores opens up avenues for improving 3D imaging resolution beyond the Rayleigh diffraction limit.

We demonstrate three-dimensional tracking of fluorescent microparticles, with a computational optical system whose point spread function (PSF) has been engineered to have two twisting lobes along the optical axis, generating a three-dimensional (3D) double-helix (DH) PSF. An information theoretical comparison in photon limited systems shows that the DH-PSF delivers higher Fisher information for 3D localization than the standard PSF. Hence, DH-PSF systems provide better position estimation accuracy. Experiments demonstrate average position estimation accuracies under 14nm and 37nm in the transverse and axial dimensions respectively. The system determines the 3D position of multiple particles with a single image and tracks them over time while providing their velocities.

Rotating point spread functions (PSFs) present invariant features that continuously rotate with defocus and are important in diverse applications such as computational imaging and atom/particle trapping. However, their transfer function efficiency is typically very low. We generate highly efficient rotating PSFs by tailoring the range of invariant rotation to the specific application. The PSF design involves an optimization procedure that applies constraints in the Gauss-Laguerre modal plane, the spatial domain, and the Fourier domain. We observed over thirty times improvement in transfer function efficiency. Experiments with a phase-only spatial light modulator demonstrate the potential of high-efficiency rotating PSFs.

The accuracy of depth estimation based on defocus effects has been essentially limited by the depth of field of the imaging system. We show that depth estimation can be improved significantly relative to classical methods by exploiting three-dimensional diffraction effects. We formulate the problem by using information theory analysis and present, to the best of our knowledge, a new paradigm for depth estimation based on spatially rotating point-spread functions (PSFs). Such PSFs are fundamentally more sensitive to defocus thanks to their first-order axial variation. Our system acquires a frame by using a rotating PSF and jointly processes it with an image acquired by using a standard PSF to recover depth information. Analytical, numerical, and experimental evidence suggest that the approach is suitable for applications such as microscopy and machine vision.