Fig. 3: 3D cell surface protein localization by MAxSIM combined with 2D-SIM imaging-based spatial mask. | Communications Biology

Fig. 3: 3D cell surface protein localization by MAxSIM combined with 2D-SIM imaging-based spatial mask.

From: MAxSIM: multi-angle-crossing structured illumination microscopy with height-controlled mirror for 3D topological mapping of live cells

Fig. 3

a, b Reconstructed MAxSIM 3D topology and zoomed-in images (marked by black boxes) of fixed neuronal dendrites obtained from scanning the incidence angle (19°, 53°) with a 0.5° step size. Primary cortical cultures from Long Evans rat embryos stained with a WGA conjugated to Alexa555 (WGA-555) with laser excitation at 560 nm or with b anti-GluA2 antibody visualized with a secondary Atto647N antibody with excitation at 647 nm. Exposure time = 200 ms. c. Merged 3D topology image of WGA (gray) and GluA2 (red). d–g. Diffraction-limited lateral image d and super-resolution 2D-SIM image with excitation numerical aperture = 1.15 e of GluA2 on the dendrite. A fast Fourier transform image was generated (f), and intensity thresholding was applied to create a mask g from the 2D-SIM image e. h Merged 3D topology image of WGA (gray) and GluA2 (red) incorporating the mask g. A median filter with kernel size = 3 was applied to reduce noise a-c, h. Scale bars = 2 µm a–e, g, h.

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