Skip to main content
[Preprint]. 2023 Jul 6:rs.3.rs-3121892. [Version 1] doi: 10.21203/rs.3.rs-3121892/v1

Figure 3. Multi-color volumetric CLEM enabled by scFv-assisted immunofluorescence.

Figure 3.

a, The high-resolution ssEM volume acquired from the cerebellar lobule, Crus 1 with multi-color immunofluorescence from scFv probes separated by linear unmixing. The multi-color fluorescence data was co-registered with the high-resolution ssEM data. Numbers 1–4 indicate approximate regions where the ultrastructure was examined at high resolution. Owing to the absence of detergent in immunofluorescence labeling, fine ultrastructure was preserved throughout the ssEM volume, such as in the molecular layer (1), in the Purkinje cell layer (2), in the glomeruli in the granule cell layer (3), and in the granule cell bodies (4). b, Demonstration of the overlay between fluorescence signals and EM ultrastructure. Left panel shows the multi-color six-channel fluorescent image of slice 250 (of 848) of the spatially transformed fluorescence image volume. Middle panel shows three fluorescence channels corresponding to the labeling of CB, GFAP, and Hoechst overlaid onto the EM micrograph of slice 250. Right panel shows four fluorescent channels corresponding to the labeling of VGluT1, Kv1.2, PV, and Hoechst overlaid onto the EM micrograph of slice 250.