Goat anti rabbit igg alexafluor647

CD45.2-V500 and TNFα-APC-Cy7 were purchased from BD Biosciences (San Jose, California). KLRG1-FITC, Eomes-PE, CD107a-PE, CD107b-PE, CD27-PE, CD127-PE-Cy5, CD127-PerCP-Cy5.5, Tbet-PerCP-Cy5.5, IFNγ-PerCP-Cy5.5, Eomes-PErCP-efluor710, CD45.1-PECy7, KLRG1-PE-Cy7, Tbet-PE-Cy7, IRF4-AlexaFluor647, CD44-AlexaFluor700, CD62L-APC-eFluor780, CD44-eFluor450, KLRG1-eFluor450, IFNγ-eFluor450, CD90.2-APC-eFluor780, CD45.1-APC-eFluor780, IL-2-PerCP-Cy5.5 were purchased from eBioscience (San Deigo, California). CD8-PE-TexasRed, GranzymeB-PE, GranzymeB-APC, Live-Dead-Violet, Live-Dead-Aqua and goat-anti-rabbit IgG-AlexaFluor647 and -AlexaFluor488 were purchased from Life Technologies (Grand Island, New York). H2D^b-GP33 monomers were prepared at UMMS; LCMV-specific (H2D^b-NP396, H2D^b-GP276) and Influenza A PR8-OVA_I-specific (H2K^b-OVA257) monomers were obtained from the NIH Tetramer Core Facility (Atlanta, Georgia). Intracellular TCF1 staining was performed using rabbit-anti-mouse TCF1 (Cell Signaling Technology, Danvers, Massachusetts) followed by staining with goat-anti-rabbit secondary (Life Technologies). Samples were analyzed on an LSRII flow cytometer (Becton Dickinson), and data were analyzed using FlowJo (Tree Star).

Immunostaining was carried out as described previously [4 (link), 21 (link)]. The following primary antibodies were used: mouse anti-FLAG® M2 (Sigma-Aldrich, cat. no. F1804; 1:200), rabbit anti-cardiac troponin I (Santa Cruz Biotechnology, cat. no. sc-15368; 1:100), mouse anti-cardiac troponin T (Thermo Scientific, cat. no. MS-295-P; 1:200), rabbit anti-Nanog (Cell Signaling Technology, cat. no. #4903; 1:50), rabbit anti-TRA-1–60 (Abcam, cat. no. ab174813; 1:100), chicken anti-beta III tubulin (Abcam, cat. no. ab41489; 1:1000), rabbit anti-SATB2 antibody (Abcam, cat. no. ab34735; 1:500), rabbit anti-TBR1 antibody (Abcam, cat. no. ab31940; 1:200), and chicken anti-tyrosine hydroxylase antibody (Abcam, cat. no. ab76442; 1:1000). Secondary antibodies used were as follows: goat anti-mouse IgG-Alexa Fluor® 546 (Life Technologies, cat. no. A-11030; 1:400), goat anti-rabbit IgG-Alexa Fluor® 647 (Life Technologies, cat. no. A-20991; 1:400), goat anti-chicken IgY-Alexa Fluor® 647 (Life Technologies, cat. no. A-21449; 1:400), and goat anti-chicken IgY-Alexa Fluor® 488 (Abcam, cat. no. ab150169; 1:400).
Hoechst 33342 (Life Technologies, cat. no. H3570; 1:10000) was used to counterstain nuclei. The stained cells and tissue sections were visualized using a confocal microscope (Olympus, FV1000) and fluorescence microscope (Keyence, BZ-X710).