Immunofluorescence was performed using sections of formalin fixed, paraffin embedded tissue generated as described above. Antigen retrieval in tissue sections was achieved using pressure-cooking in citrate buffer pH6 for 20 minutes. Tissue sections were incubated with primary antibodies for 1 hour at room temperature.
Primary antibodies included: Ki67 (1:200; Abcam, ab15580 and 1:500; Thermo Fisher Scientific, 14-5698-82), Calbindin (1:50; Millipore, AB1778), Contactin2 (1:75; R&D Systems, AF4439), Lef1 (1:500; Abcam, ab137872), Pax6 (1:200; Biolegend, 901301), G3BP1 (1:100; Proteintech, 66486-1-Ig), Gfap (1:500; Dako, Z0334), NeuN (1:200; Millipore, MAB377), Irf1 (1:100; Abcam, ab240299), Iba1 (1:500; Wako, 013-27691) and Synaptophysin (1:100; Cell Signaling, 9020). Following washing, tissue sections where then incubated for 1 hour at room temperature in secondary antibody. Secondary antibodies included Alexa 488, 594 or 680 (1:200; Invitrogen, A-21206, A-21207 or A10043). Sections were then counterstained using DAPI (Cell Signaling, 4083) and mounted using ProLong Gold antifade mountant (Thermo Fisher Scientific, P36930). Digital images of tissue sections were captured using a Zeiss ImagerM2 and Apotome microscope.
Primary antibodies included: Ki67 (1:200; Abcam, ab15580 and 1:500; Thermo Fisher Scientific, 14-5698-82), Calbindin (1:50; Millipore, AB1778), Contactin2 (1:75; R&D Systems, AF4439), Lef1 (1:500; Abcam, ab137872), Pax6 (1:200; Biolegend, 901301), G3BP1 (1:100; Proteintech, 66486-1-Ig), Gfap (1:500; Dako, Z0334), NeuN (1:200; Millipore, MAB377), Irf1 (1:100; Abcam, ab240299), Iba1 (1:500; Wako, 013-27691) and Synaptophysin (1:100; Cell Signaling, 9020). Following washing, tissue sections where then incubated for 1 hour at room temperature in secondary antibody. Secondary antibodies included Alexa 488, 594 or 680 (1:200; Invitrogen, A-21206, A-21207 or A10043). Sections were then counterstained using DAPI (Cell Signaling, 4083) and mounted using ProLong Gold antifade mountant (Thermo Fisher Scientific, P36930). Digital images of tissue sections were captured using a Zeiss ImagerM2 and Apotome microscope.