Goat anti rat alexa fluor 633

Adult brains were dissected in 1× PBS and fixed at room temperature for 25-30 min in 4% formaldehyde in phosphate buffer [4% formaldehyde, 0.1 M phosphate buffer (pH 7.2), 0.2% Triton X-100]. Samples were then washed with 1× PBS and placed in blocking buffer (PBS, 0.2% Triton X-100, 0.1% normal goat serum) for 2 h at room temperature or 4°C overnight. Brains were incubated with primary antibodies diluted with blocking buffer at 4°C overnight, then washed twice with PBST (PBS+0.1% Triton X-100) and incubated in secondary antibodies diluted with blocking buffer for 4 h at room temperature, then washed twice again with PBST. DAPI was added in the final wash for 30 min at room temperature. Brains were mounted on slides in Vectashield (Vector Laboratories, Burlingame, CA). The antibodies used are as follows: rabbit anti-cleaved Dcp-1 (Cell Signaling Technology, Danvers, MA; #9578, 1:100), mouse anti-Repo (Developmental Studies Hybridoma Bank, University of Iowa, Iowa City, IA; 8D12, 1:50), Rat anti-Elav (Developmental Studies Hybridoma Bank, University of Iowa, Iowa City, IA; 7E8A10, 1:250), DAPI (Sigma-Aldrich, St Louis, MO; 1 mg/l), goat anti-mouse Alexa-Fluor-488 (Life Technologies, Carlsbad, CA; 1:200), goat anti-rabbit Alexa-Fluor-568 (Life Technologies, Carlsbad, CA; 1:200), goat anti-rat Alexa-Fluor-633 (Invitrogen, Carlsbad, CA; 1:200).

Brains were dissected, stained, and imaged as described in Sudmeier et al. (2015) (link). The antibodies used are as follows: Chicken anti-Green Fluorescent Protein (Life Technologies, Carlsbad, CA; 1:500), Mouse anti-Repo (Developmental Studies Hybridoma Bank, University of Iowa, Iowa City, IA; 8D12, 1:50), Rat anti-Elav (Developmental Studies Hybridoma Bank, University of Iowa, Iowa City, IA; 7E8A10, 1:250), DAPI (Sigma-Aldrich, St. Louis, MO; 1 mg/L), Goat anti-Chicken Alexa Fluor-488 (Life Technologies, Carlsbad, CA; 1:200), Goat anti-Mouse Alexa Fluor-568 (Life Technologies, Carlsbad, CA; 1:200), and Goat anti-Rat Alexa Fluor-633 (Invitrogen, Carlsbad, CA; 1:200).