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By engineering the point-spread function (PSF) of single molecules, different fluorophore species can be imaged simultaneously and distinguished by their unique PSF patterns. Here, we insert a silicon-dioxide phase plate at the Fourier plane of the detection path of a wide-field fluorescence microscope to produce distinguishable PSFs (X-PSFs) at different wavelengths. We demonstrate that the resulting PSFs can be localized spatially and spectrally using a maximum-likelihood estimation algorithm and can be utilized for hyper-spectral super-resolution microscopy of biological samples. We produced superresolution images of fixed U2OS cells using X-PSFs for dSTORM imaging with simultaneous illumination of up to three fluorophore species. The species were distinguished only by the PSF pattern. We achieved ∼21-nm lateral localization precision (FWHM) and ∼17-nm axial precision (FWHM) with an average of 1,800 - 3,500 photons per PSF and a background as high as 130 - 400 photons per pixel. The modified PSF distinguished fluorescent probes with ∼80 nm separation between spectral peaks.

We designed, fabricated, and characterized a flat multi-level diffractive lens comprised of only silicon with$\mathrm{d}\mathrm{i}\mathrm{a}\mathrm{m}\mathrm{e}\mathrm{t}\mathrm{e}\mathrm{r}=15.2\mathrm{m}\mathrm{m}$, focal$\mathrm{l}\mathrm{e}\mathrm{n}\mathrm{g}\mathrm{t}\mathrm{h}=19\mathrm{m}\mathrm{m}$, numerical aperture of 0.371, and operating over the long-wave infrared (LWIR)$\mathrm{s}\mathrm{p}\mathrm{e}\mathrm{c}\mathrm{t}\mathrm{r}\mathrm{u}\mathrm{m}=8\text{µ<\#comment/>}\mathrm{m}$to 14 µm. We experimentally demonstrated a field of view of 46°, depth of focus$><\#comment/>5\mathrm{m}\mathrm{m}$, and wavelength-averaged Strehl ratio of 0.46. All of these metrics were comparable to those of a conventional refractive lens. The active device thickness is only 8 µm, and its weight (including the silicon substrate) is less than 0.2 g.