Rabbit anti darpp32

Cells were fixed with paraformaldehyde (4% in PBS) for 20 min, followed by permeabilization (0.2% Triton X-100 in PBS for 10 min) and blocking (3% bovine serum albumin in 0.2% Triton X-100 in PBS for 10 min). Human iPSCs/NPCs were incubated in primary antibody for 2 h at room temperature (Mouse anti-polyQ (Millipore, MAB1574, 1:50), Mouse anti-OCT4 (Stem Cell Technologies, #60093, 1:200), Rabbit anti-PAX6 (Stem Cell Technologies, #60094, 1:300), Mouse anti-FUS (Abcam, #154141, 1:500), Rabbit anti-G3BP1 (MBL, #RN048PW, 1:500), Rabbit anti-DARPP32 (Abcam, #40801, 1:50), Rabbit anti-GABA (Sigma, #A2052, 1:100) and Chicken anti-MAP2 (Abcam, #5392, 1:500). Then, cells were washed with 0.2% Triton-X/PBS and incubated with secondary antibody (Alexa Fluor 488 Goat anti-Mouse (ThermoFisher Scientific, #A-11029, 1:500), Alexa Fluor 568F(ab’)2 Fragment of Goat Anti-Rabbit IgG (H+L) (ThermoFisher Scientific, #A-21069, 1:500)), Alexa Fluor 647 Donkey anti-Chicken (Jackson ImmunoResearch, #A-703-605-155, 1:500) and Hoechst 33342 (Life Technologies, #1656104) for 1 h at room temperature. PBS and distilled water wash were followed before the cover slips were mounted on Mowiol (Sigma, #324590).

Mice were anesthetized and perfused with PFA 4%. Brains were dissected, embedded in optimal cutting temperature (OCT, Tissue-Tek) and sectioned (10 μm) using a cryostat. For immunohistochemistry, slides underwent citrate buffer antigen retrieval and were then incubated with blocking solution (5% goat serum, 1% BSA, 0.05% Triton-X in PBS) for 1 hr. Then, slides were incubated overnight at 4°C with the following primary antibodies: rabbit anti -GFAP (1:200, Invitrogen), rabbit anti-DARPP-32 (1:50, Abcam), and mouse anti-TH (1:200, Sigma). Then, sections were incubated with the secondary antibody Alexa Fluor 488 and 568 (1:500; Invitrogen) for 1 hr. The nuclei were counterstained with 4, 6-diamidino-2-phenylindole (DAPI; 1:1000; Sigma-Aldrich).