Alexa fluor 488 donkey anti chicken igg

Primary antibodies were used at the following concentrations for immunohistochemistry: rabbit anti-NF200 (1:500, Sigma, St. Louis, MO, #N4142), IB4-biotin conjugate (1:200, Sigma, #L2140), rabbit anti-CGRP (1:2000, Peninsula Labs, San Carlos, CA, #T4032), chicken anti-GFP (1:1000, Avés Labs Inc, Davis, CA, #GFP-1020), and rabbit anti-GFAP (1:500, Agilent, Santa Clara, CA, #N1506). Secondary antibodies used were Alexa Fluor 647-goat anti-rabbit IgG (1:400, Invitrogen, Indianapolis, IN, #31573), fluorescein (FITC)-conjugated goat anti-rabbit IgG (1:400, Millipore, Temecula, CA, AP307F), Alexa Fluor 568-conjugated goat anti-mouse IgG1 (1:400, Invitrogen, A21124), Alexa-Fluor 568-conjugated goat anti-rabbit (1:400, Invitrogen, #A11011), rhodamine (TRITC) streptavidin (1:400, Jackson ImmunoResearch Labs Inc, West Grove, PA, #016-020-084), or Alexa Fluor 488-donkey anti-chicken IgG (1:400, Jackson ImmunoResearch Labs Inc, #703-545-155). DAPI nucleic acid stain (1:1000, Thermo Fisher Scientific, Pittsburgh, PA, #D1306) for cell nuclei or Nissl substance stain (1:200, Thermo Fisher Scientific, #N21482) for neuronal cells was used to counterstain prior to final wash steps in PBS.

Anti-Pax6 (rabbit; Medical & Biological Laboratories #PD022), Pax7 (mouse; DSHB, PAX7-s), FoxA2 (mouse; DSHB, 4C7), β-galactosidase (β-gal: chicken; abcam, ab9361) and GFP (rabbit; MBL, 598) were used as the primary antibody at dilution of 1:1000 (Pax6), 1:10 (Pax7), 1:20 (FoxA2), 1:1000 (β-gal) and 1:1000 (GFP), respectively. For the secondary antibody, Alexa Fluor 488 donkey anti-chicken IgG (Jackson Immuno Research, 703-545-155), 647 donkey anti-mouse IgG (abcam, ab150111) and 647 donkey anti-rabbit IgG (abcam, ab150075) were utilized at dilution of 1:600. Immunostaining was performed according to the standard methods. DAPI (1:10000; Invitrogen) staining was performed to visualize nuclei. Fluorescent images were acquired under BZ-X700 (KEYENCE).