Mab 1621

Immunohistochemistry was performed on whole-mount retinas. Antibodies to neurofilament-M (1∶1000, MAB-1621; Millipore, Billerica, MA), melanopsin (1∶250, PA1-781; Thermo Scientific, Waltham, MA) and RNA binding protein with multiple splicing (RBPMS, 1∶1000) were used. The RBPMS polyclonal antibodies were generated against the N-terminus of the RBPMS polypeptide, GGKAEKENTPSEANLQEEEVR, in guinea pig by a commercial vendor (ProSci, Poway, CA), and affinity purified and characterized in our laboratory [8] . Retinas were mounted on cellulose filter paper (Millipore) with the GCL up and fixed in 4% PFA for 10 minutes. Whole-mounted retinas were incubated in 10% normal goat serum at 4°C overnight. The retinas were subsequently incubated in primary antibody for 5–7 days at 4°C, washed three times in phosphate buffer (PB) 0.1 M pH = 7.4 and then incubated overnight at 4°C in the appropriate secondary antibody (1∶500, coupled to Alexa Fluor 488, 633 or Cy3, Invitrogen, Carlsbad, CA). After three final washes in PB, the retinas were mounted in Vectashield Mounting Medium (Vector Laboratories, Burlingame, CA). Coverslips were sealed with nail polish for prolonged storage. Slides were stored at 4°C and protected from light.

Mouse anti-TUJ1 1:1000 (MMS-435P-250, Covance), rabbit anti-CHAT 1:1000 (AB 143, Millipore), mouse anti-NFM 1:500 (MAB1621, Millipore), rabbit anti-ISLET1 1:1000 (20670, Abcam), rabbit anti-GFAP 1:1000 (AB5804, Millipore), rabbit anti-AQP4 1:100 (AB3594, Millipore), rabbit anti-EAAT2 1:100 (ab41621, Abcam), mouse anti-GS 1:100 (MAB302, Millipore), mouse anti-O4 1:1000 (MAB345, Millipore), rabbit anti-IBA1 1:500 (019-19741, Wakochemical), rat anti-CD11b 1:200 (550282, BD pharmingen), mouse anti-MAP2 1:500 (MAB3418, Millipore), mouse anti-TDP-43 1:300 (gift from Dr J-P Julien’s laboratory), mouse anti-NeuN 1:500 (MAB377, Millipore), mouse anti-CNPase 1:200 (NE1020, Millipore), mouse anti-Nestin 1:200 (AB6142, Abcam), mouse anti-SOX2 1:500 (MAB4343, Millipore), mouse anti-Vimentin 1:1000 (AB28028, Abcam), rabbit anti-OLIG2 1:500 (AB9610, Millipore), rat anti-CD31 1:500 (550274, BD Biosciences), rabbit anti-cleaved caspase 3 1:200 AB3623, Millipore), Alexa fluor 488 goat anti-rabbit (A-11008, Life Technologies), Alexa fluor 594 goat anti-mouse (A-11005, Life Technologies)