Journal: The Journal of Experimental Medicine

Article Title: Anergy in Peripheral Memory Cd4 + T Cells Induced by Low Avidity Engagement of T Cell Receptor

doi:

Figure Lengend Snippet: Anergized cells are able to expand in the presence of rIL-2. Splenocytes and lymph node cells from anergized and nonanergized mice were cultured without peptide, with peptide (10 μM) alone or with rIL-2 (10 U/ml) for 8 d. Cells were stained with HA-DR1-SA-PE oligomers and anti-CD4–FITC.

Article Snippet: 9 d after the second peptide injections, 5 × 10 6 cells of HLA-DR1 Tg mice were cultured either with peptide (10 μM) alone or with rIL-2 (10 U/ml) for 8 d. Cells were stained with HA-DR1-SA-PE oligomers (provided by T. Cameron and L. Stern, MIT, Cambridge, MA) for 3.5 h at 37°C followed by anti–mouse CD4-FITC or anti–mouse CD4-Cyc and anti–mouse CTLA-4–FITC (Southern Biotechnology Associates, Inc.).

Techniques: Cell Culture, Staining

Journal: Journal for Immunotherapy of Cancer

Article Title: In situ vaccination at a peripheral tumor site augments response against melanoma brain metastases

doi: 10.1136/jitc-2020-000809

Figure Lengend Snippet: Intracranial versus extracranial infiltrating immune cell analysis after ISV + α-CTLA-4 treatment. (A) Survival curve for immune memory mice depleted of T-cells (CD4 and CD8) prior to receiving intracranial injection of B78 (**p<0.01, Kaplan-Meier, n≥4) compared with control (rat IgG) and naïve, and successful brain tumor engraftments (‘N’ above bars). (B) Immunohistochemistry of ISV + α-CTLA-4 treated and untreated mice, comparing intracranial (brain met, BrMet) and extracranial (left flank, LF) B78 melanoma tumors (brown=positive immunolabeling). (C) Quantified immunohistochemistry (***p<0.001, **p<0.01, *p<0.05, mean±SE with marker representing each individual mouse (ie, average of three high-powered fields), ANOVA with post hoc Bonferroni, n≥5, at least two independent animal experiments). (D) Flow cytometric analysis of B78 tumors after ISV + α-CTLA-4 treatment comparing BrMet to extracranial tumors (**p<0.01, mean±SE with marker representing individual data points, ANOVA with post hoc Bonferroni, n≥8, at least two independent animal experiments). ANOVA, analysis of variance; ISV, in situ vaccination.

Article Snippet: For IHC sections, antibodies used were: CD4 (mAb Rat IgG2b, kappa; clone GK1.5; Tonbo Biosciences 70–0041 U500l, 1:1000 dilution), CD8a (mAb Rat IgG2a, kappa; clone 53–6.7; eBioscience 14-0081-85, 1:1000), FOXP3 (mAb Rat IgG2a, kappa; clone FJK-16S; eBioscience 14-5773-82, 1:500), F4/80 (mAb Rat IgG2a,κ; clone BM8; BioLegend 123101, 1:2000), CD11b (mAb Rat, clone M1/70.15, Invitrogen MA5-17857, 1:6000); specific for paraffin were CD4 (mAb Rat IgG 1 , kappa; clone 4SM95; eBioscience 14-9766-82, 1:500) and CD8a (mAb Rat IgG2a, lambda; clone 4SM15; eBioscience 14-0808-80, 1:250).

Techniques: Cell Analysis, Injection, Control, Immunohistochemistry, Immunolabeling, Marker, In Situ