mscv ires ngfr  (Addgene inc)

Journal: STAR Protocols

Article Title: Protocol to assess cell-intrinsic regulatory mechanisms using an ex vivo murine T cell polarization and co-culture system

doi: 10.1016/j.xpro.2022.101543

Figure Lengend Snippet: Schematic representation of the retroviral vectors described in this protocol Top: MSCV-IRES-NGFR (NGFR, Addgene #27489). Bottom: MSCV-IRES-Thy1.1 (Thy1.1, Addgene #17442).

Article Snippet: MSCV-IRES-NGFR (NGFR) , Addgene , RRID: Addgene_27489.

Techniques: Retroviral

Journal: STAR Protocols

Article Title: Protocol to assess cell-intrinsic regulatory mechanisms using an ex vivo murine T cell polarization and co-culture system

doi: 10.1016/j.xpro.2022.101543

Figure Lengend Snippet: Schematic overview of the murine splenic T-cell retroviral transduction and co-culture protocol Step 1: Production of Thy1.1 and NGFR-expression construct-carrying retroviruses in the Plat-E packaging cell line. Step 2: Isolation, activation, and transduction of splenic naïve CD4 + T cells from RORγt WT and RORγt S182A mice. Step 3: Co-culture, polarization, and harvest of retrovirally transduced RORγt WT and RORγt S182A CD4 + T cells marked by cell surface Thy1.1 or NGFR.

Article Snippet: MSCV-IRES-NGFR (NGFR) , Addgene , RRID: Addgene_27489.

Techniques: Retroviral, Transduction, Co-Culture Assay, Expressing, Construct, Isolation, Activation Assay

Journal: STAR Protocols

Article Title: Protocol to assess cell-intrinsic regulatory mechanisms using an ex vivo murine T cell polarization and co-culture system

doi: 10.1016/j.xpro.2022.101543

Figure Lengend Snippet: Comparison of ex vivo co-culture and in vivo intestinal lamina propria T lymphocyte populations Top: Representative flow cytometry plots of transduced RORγt WT (NGFR + ) and RORγt S182A (Thy1.1 + ) CD4 + T cells co-cultured and polarized toward multiple T cell subsets, including Th17 (RORγt + Foxp3 - , blue), RORγt + Treg (RORγt + Foxp3 + , red) and cTreg (RORγt - Foxp3 + , green). Bottom: Representative flow cytometry plots from the small intestinal and colonic lamina propria (siLP and cLP), respectively. Lamina propria T lymphocyte populations were isolated as previously described by ( ; Valle-Noguera et al., 2020 ). Note that IL-10 staining on ex vivo (top) and in vivo (bottom) samples was performed with clone JES5-16E3 (Alexa Fluor 488) and JES5-16E3 (Alexa Fluor 700), respectively.

Article Snippet: MSCV-IRES-NGFR (NGFR) , Addgene , RRID: Addgene_27489.

Techniques: Comparison, Ex Vivo, Co-Culture Assay, In Vivo, Flow Cytometry, Cell Culture, Isolation, Staining

Journal: STAR Protocols

Article Title: Protocol to assess cell-intrinsic regulatory mechanisms using an ex vivo murine T cell polarization and co-culture system

doi: 10.1016/j.xpro.2022.101543

Figure Lengend Snippet: Expected transduction efficiency in cultured murine CD4 + T cells Top: Flow cytometry gating strategy for evaluating transduction efficiency in live CD4 + T cells. Bottom: Proportions of live NGFR + or Thy1.1 + CD4 + T cells 72 h post-polarization. Data are represented as mean ± SD.

Article Snippet: MSCV-IRES-NGFR (NGFR) , Addgene , RRID: Addgene_27489.

Techniques: Transduction, Cell Culture, Flow Cytometry

Journal: STAR Protocols

Article Title: Protocol to assess cell-intrinsic regulatory mechanisms using an ex vivo murine T cell polarization and co-culture system

doi: 10.1016/j.xpro.2022.101543

Figure Lengend Snippet:

Article Snippet: MSCV-IRES-NGFR (NGFR) , Addgene , RRID: Addgene_27489.

Techniques: Recombinant, Modification, Infection, Transfection, Staining, Plasmid Preparation, Cell Isolation, Bacteria, Retroviral, Functional Assay, Software