Journal: Neurochemistry International
Article Title: Expression and secretion of synaptic proteins during stem cell differentiation to cortical neurons
Figure Lengend Snippet: Neuronal and synaptic maturation during differentiation of hiPSC to cortical neurons. Representative images from immunocytochemistry staining during differentiation. (A and B) NANOG and OCT-4 were strongly stained on d0 and the stainings became weak on d4, while KI-67 had similar staining intensities on d0 and d4. (C) PAX-6, a primary neuro-progenitor expression decreased from d20 to d40 of differentiation. (D) Staining for neuron specific tubulin, TUJ-1 was weak on d30 but strong staining was observed on d60. (E) SV-2, a pre-synaptic protein, staining was weak on d60. On d120, a strong punctuate staining was observed (arrows). (F) PSD-95, a post-synaptic density protein, staining was weak on d60. On d120, the staining had intensified and became punctate (arrows). Green or red = protein of interest, blue = nuclei (DAPI), Scale bar = 20 μm.
Article Snippet: Primary antibodies, OCT-4 (1:400; Cell Signaling, D73G4), NANOG (1:800, Cell Signaling, C30A3), TUJ-1 (1:2000; Abcam ab14545), SV-2 (1:500; DSHB), PSD-95 (1:100; NeuroMab, P78352), KI-67 (1:600; BD Pharmingen™, 550609), PAX-6 (1:600; BioLegend, 901301), nestin (1:50; R & D Systems, MAB1259), CTIP-2 (1:300; Abcam, ab18465), TBR-1 (1:300; Abcam, ab31940), BRN-2 (1:400; Santa Cruz, sc-6029), CUX-1 (1:300; Santa Cruz, sc-13024), GAP-43 (1:1000; Abcam, ab75810), tau (1:1000; Biorbyt, orb175815), NRGN (1:100; Upstate Biotechnologies, 07–425), SNAP-25 (1:400; Sigma Aldrich, S9684-100UL) and SYT-1 antibody (1:200; Synaptic systems, 105,011) were diluted in block buffer and incubated at 4 °C overnight.
Techniques: Immunocytochemistry, Staining, Expressing