Extended Data Fig. 5: Three-dimensional image stack of fluorescent beads under SD-SIM and Sparse SD-SIM. | Nature Biotechnology

Extended Data Fig. 5: Three-dimensional image stack of fluorescent beads under SD-SIM and Sparse SD-SIM.

From: Sparse deconvolution improves the resolution of live-cell super-resolution fluorescence microscopy

Extended Data Fig. 5

(a,b) A maximum intensity projection (MIP) view (left) and a horizontal section (right) of fluorescent beads (100 nm in diameter) recorded by SD-SIM (a) and after the sparse deconvolution (b), respectively. Insets in the left-lower corner show a magnified view of the same fluorescent bead under different reconstruction methods. (c) The corresponding Gaussian fitted profiles in (a, left-lower corner), which indicate that the lateral FWHM of SD-SIM (red) and Sparse SD-SIM (blue) are 185 nm (calibrated resolution ~165 nm) and 110 nm (calibrated resolution ~90 nm), respectively (Supplementary Note 9.2). (d) Magnified horizontal sections from the white boxes in (a–b) are shown in the left and right panels, while the SD-SIM image is processed with a median filter to avoid a non-converged fitted result. (e) We used Gaussian functions to fit the intensity profiles along the axial direction of the fluorescent bead in (d), yielding axial resolutions of 484 nm and 266 nm for SD-SIM and Sparse SD-SIM, respectively. (f) The gradually improved axial resolution (FWHM) of a 100 nm bead while increasing the weight of sparsity. (g) Measuring the FWHM with fluorescent beads with a diameter of 45 nm. The fitted FWHMs (cross-sections between white arrows displayed with white profiles in the right) of SD-SIM and Sparse SD-SIM are 175 nm and 92 nm, respectively. As shown with yellow profiles (cross-sections between yellow arrows), the Sparse SD-SIM resolved adjacent two beads with a distance of 95 nm. Experiments were repeated five times independently with similar results. Scale bars: (b) 4 μm, (b, inset) 100 nm and (d, f, g) 200 nm.

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