(A) Appearance of a brain sample after FixEL with probe 1 (mGlu1) and 3DISCO. Scale bars, 2.5 mm.
(B) z stacking fluorescence imaging of the (A) sample. PBS(–) containing probe 1 (mGlu1) (4.5 μL, 100 μM: Cbrain is ca. 1 μM) was injected into mouse lateral ventricle. After 6 h of incubation, the mouse was transcardially perfused with 4% PFA. After 3DISCO treatment, z stacking fluorescence imaging of the whole brain was performed using a CLSM equipped with a 5× objective and a GaAsP detector (633 nm excitation for Ax647). Scale bars, 1 mm.
(C) 3D rendering of the whole brain in (B). Scale bars, 1 mm.
(D) z stacking fluorescence imaging of cerebellum region in (C) with a 40× objective. Scale bars, 20 μm.
(E) Enlarged 3D rendering image of the cerebellar molecular layer (ML) in (D). Scale bars, 5 μm.
(F) z stacking fluorescence imaging of the whole brain after FixEL with probe 3 (DRD2). PBS(–) containing 25 μM of probe 3 (DRD2) (4.5 μL) was injected into mouse bilateral ventricle respectively. After FixEL (20 h incubation), the lower 2 mm of the brain was cutoff and then the upper side of the brain was treated with 3DISCO. z stacking fluorescence imaging was performed using a CLSM equipped with a 5× objective and a GaAsP detector (633 nm excitation for Ax647). Scale bars, 1 mm.
See also Figures S9-S11.