cd8 α ly 2 icrobeads  (Miltenyi Biotec)

Journal: Cell host & microbe

Article Title: SMAC mimetics induce autophagy-dependent apoptosis of HIV-1-infected resting memory CD4+ T cells

doi: 10.1016/j.chom.2018.09.007

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: α-human CD8 microbeads , Miltenyi Biotec , Cat# 130-045-201.

Techniques: Virus, Recombinant, Marker, Enzyme-linked Immunosorbent Assay, Cell Isolation, shRNA, Transduction, Software

Journal: Nature Communications

Article Title: Rational combination of oncolytic vaccinia virus and PD-L1 blockade works synergistically to enhance therapeutic efficacy

doi: 10.1038/ncomms14754

Figure Lengend Snippet: B6 mice were intraperitoneally inoculated with 5 × 10 5 MC38-luc and treated with VV and/or α-PD-L1 as described. Single cells were made from primary tumours collected from tumour-bearing mice at day 5 post first treatment, blocked with α-CD16/32 Ab and then stained with antibodies against CD45, CD8, CD4, PD-1, ICOS, PD-1, CTLA-4, TIM-3, LAG-3, TIGIT and Foxp3 to determine the quantities of CD8 + T cells ( a ), CD8 + T-cell activation ( b – d ), CD8 + T-cell exhaustion ( e – h ), Treg cells ( i ), CD8 + /CD4 + Foxp3 + T cells ( j ) and CD4 + Foxp3 − T cells ( k , l ) in the TME. Of note, the anti-PD-L1 antibody clone 10F.9G2 was used for therapy while clone MHI5 was used for subsequent phenotypic analysis. Data were analysed using Student's t- test (* P <0.05; ** P <0.01; *** P <0.001).

Article Snippet: Splenic CD8 + T cells were isolated from naive and MC38-luc-bearing B6 mice received different treatment 18 days post tumour cell injection using α-mouse CD8 microbeads following vendor's protocols (Miltenyi Biotec, San Diego, CA, USA).

Techniques: Staining, Activation Assay

Journal: Nature Communications

Article Title: Rational combination of oncolytic vaccinia virus and PD-L1 blockade works synergistically to enhance therapeutic efficacy

doi: 10.1038/ncomms14754

Figure Lengend Snippet: ( a ) B6 mice were intraperitoneally inoculated with 5 × 10 5 MC38-luc cancer cells and treated with VV and/or α-PD-L1 as described. Splenic CD8 + T cells (4 × 10 5 ) were isolated from naive and MC38-luc-bearing mice that received different treatments 18 days post tumour cell injection and restimulated with mitomycin C-treated MC38-luc or B16 cancer cells (4 × 10 4 cells each) in the presence of 4000-rad-irradiated CD8-depleted naive B6 splenocytes (2 × 10 6 ) in 200 μl RPMI-1640 medium supplemented with 10% FBS at 37 °C, 5% CO 2 for 2 days. The concentration of IFN-γ in the culture supernatants was tested by ELISA. The statistical analyses were performed with t -test. ( b ) Naive or MC38-luc-bearing B6 mice with dual treatments, which survived for more than 60 days, were s.c. rechallenged with 1 × 10 6 MC38-luc cancer cells. The primary tumour size was measured and presented here. ( c ) In a separate experiment, B6 mice were inoculated with 5 × 10 5 MC38-luc cells i.p. and treated with VV plus α-PD-L1 or PBS at day 5 post tumour inoculation, α-PD-L1 Ab was injected every 2 days for a total of four times. α-CD8 Ab (250 μg per injection), α-CD4 Ab (150 μg per injection) or α-IFN-γ Ab (200 μg per injection) were intraperitoneally injected into mice to deplete CD8+ T cells, CD4+ T cells or neutralize circulating IFN-γ as scheduled in c , and the overall survival was monitored by Kaplan–Meier analysis and analysed using log rank test ( d ).

Article Snippet: Splenic CD8 + T cells were isolated from naive and MC38-luc-bearing B6 mice received different treatment 18 days post tumour cell injection using α-mouse CD8 microbeads following vendor's protocols (Miltenyi Biotec, San Diego, CA, USA).

Techniques: Isolation, Injection, Irradiation, Concentration Assay, Enzyme-linked Immunosorbent Assay

Journal: PLoS Pathogens

Article Title: Th17 Cells Are More Protective Than Th1 Cells Against the Intracellular Parasite Trypanosoma cruzi

doi: 10.1371/journal.ppat.1005902

Figure Lengend Snippet: Polyclonal CD8 + T cells were transferred into RAG KO mice along with either Th1 or Th17 T . cruzi -specific T cells (>10/group). Mice were then systemically challenged with T . cruzi . (A) Parasitemia counts 19 days post-infection demonstrate that co-adoptive transfer of Th17 cells resulted in better control of infection than co-adoptive transfer of Th1 cells ***p<0.001, **p<0.01, *p<0.05 by two-tailed Student t test. (B) All groups receiving CD8 + T cells were significantly protected compared to no adoptive transfer. Co-adoptive transfer of Th17 cells led to 100% long-term survival that was significantly higher than survival in all other groups.***p<0.001 by Mantel-Cox log-rank test. (C) T-bet KO Th17 cells resulted in decreases in parasite load comparable to WT Th17 cells when transferred into RAG KO mice (>7/group) with CD8 + T cells. **p<0.01 by two-tailed Student t test. (D) T-bet KO Th17 cells resulted in equivalent long-term protection from mortality compared to WT Th17 cells.

Article Snippet: In vitro generated Th1 or Th17 cells were transferred i.v. into RAG KO mice, along with polyclonal CD8 + T cells purified from naïve BALB/c mice by positive selection using α-CD8 microbeads (Miltenyi Biotec, Auburn, CA).

Techniques: Infection, Adoptive Transfer Assay, Two Tailed Test

Journal: PLoS Pathogens

Article Title: Th17 Cells Are More Protective Than Th1 Cells Against the Intracellular Parasite Trypanosoma cruzi

doi: 10.1371/journal.ppat.1005902

Figure Lengend Snippet: (A-B) T . cruzi -specific Th17 cells were co-transferred with CD8 + T cells into RAG KO mice (5/group) prior to T . cruzi challenge. Neutralizing anti-IL-17A or control IgG1 antibodies were injected intraperitoneally every 48 hours. IL-17A neutralization did not reduce protection as measured by both parasitemia (A) and survival (B). *p<0.001 by two-tailed Student t test, **p<0.01 by log-rank test compared with negative controls. (C-D) Polyclonal CD8 + T cells were transferred intravenously (i.v.) into RAG KO mice prior to T . cruzi infection. Either control AdV or IL-17 AdV was injected 1 day prior to infection and 7 days post-infection at 5x10 9 PFU i.v. Protection was measured by parasitemia 18 days post-infection (C) and survival (D).

Article Snippet: In vitro generated Th1 or Th17 cells were transferred i.v. into RAG KO mice, along with polyclonal CD8 + T cells purified from naïve BALB/c mice by positive selection using α-CD8 microbeads (Miltenyi Biotec, Auburn, CA).

Techniques: Injection, Neutralization, Two Tailed Test, Infection

Journal: PLoS Pathogens

Article Title: Th17 Cells Are More Protective Than Th1 Cells Against the Intracellular Parasite Trypanosoma cruzi

doi: 10.1371/journal.ppat.1005902

Figure Lengend Snippet: (A) Th17 cells required co-adoptive transfer with polyclonal CD8 + T cells to confer optimal immunity as measured by parasite burden. **p<0.01, ***p<0.001 by two-tailed Student t test. (B) Th17 cells transferred alone were not able to protect mice from T . cruzi -related death, indicating that Th17 cells protect through helper effects on CD8 + T cells. **p<0.01 by Mantel-Cox log-rank test. (C) CFSE-labeled, polyclonal CD8 + T cells were sub-optimally activated with plate-bound α-CD3 (1 μg/ml) and increasing numbers of dendritic cells. CD8 + T cells were cultured with or without Th17 cells for 5 days. Proliferation was measured by CFSE dilution. (D) CD8 + T cells were sub-optimally activated with α-CD3 as in C (without added dendritic cells), in the presence of Th17 cells, Th17 SNs or Th17 cells plus IL-17A neutralizing antibody. CD8 + T cell proliferation, CD44 expression and MIP-1α/IFN-γ production were measured 5 days later by Flow Cytometry and ICS and shown as fold increases compared with α-CD3 activation alone. (E) Purified Th17 cytokines were individually added to sub-optimally α-CD3 activated CD8 + T cells and markers of activation were analyzed 5 days later. (F) High levels of IL-21 could be detected by ELISA in Th17 cell SNs, but not in Th1 cell SNs. Representative results from multiple experiments are shown.

Article Snippet: In vitro generated Th1 or Th17 cells were transferred i.v. into RAG KO mice, along with polyclonal CD8 + T cells purified from naïve BALB/c mice by positive selection using α-CD8 microbeads (Miltenyi Biotec, Auburn, CA).

Techniques: Adoptive Transfer Assay, Two Tailed Test, Labeling, Cell Culture, Expressing, Flow Cytometry, Activation Assay, Purification, Enzyme-linked Immunosorbent Assay

Journal: PLoS Pathogens

Article Title: Th17 Cells Are More Protective Than Th1 Cells Against the Intracellular Parasite Trypanosoma cruzi

doi: 10.1371/journal.ppat.1005902

Figure Lengend Snippet: (A) WT or IL-21R KO CD8 + T cells were sub-optimally stimulated with plate-bound α-CD3 as in . ICS assays indicated the addition of Th17 cell SNs could help activate WT CD8 + T cells but not IL-21R KO CD8 + T cells. Shown are fold increases compared to WT or IL-21R KO CD8 + T cells cultured with sub-optimal α-CD3 alone, without Th17 cell SNs. (B) Total spleen cells (TSCs) recovered from infected RAG KO mice adoptively transferred with Th17 cells and IL-21R KO CD8 + T cells had diminished responses to TS stimulation compared to mice receiving Th17 cells and WT CD8 + T cells prior to challenge, as measured by IFN-γ ELISPOT. (C) Mice transferred with Th17 cells (n = 4) and IL-21R KO CD8 + T cells (n = 8) were unable to control infection, as indicated by increasing parasitemia over time. **p<0.01 by two-tailed Student t test. (D) Transfer of IL-21R KO CD8 + T cells failed to protect mice from T . cruzi -mortality. *** p<0.001 by Mantel-Cox log-rank test.

Article Snippet: In vitro generated Th1 or Th17 cells were transferred i.v. into RAG KO mice, along with polyclonal CD8 + T cells purified from naïve BALB/c mice by positive selection using α-CD8 microbeads (Miltenyi Biotec, Auburn, CA).

Techniques: Cell Culture, Infection, Enzyme-linked Immunospot, Two Tailed Test

Journal: PLoS Pathogens

Article Title: Th17 Cells Are More Protective Than Th1 Cells Against the Intracellular Parasite Trypanosoma cruzi

doi: 10.1371/journal.ppat.1005902

Figure Lengend Snippet: RAG KO mice were adoptively transferred with polyclonal CD8 + T cells and Th1 or Th17 cells and then infected systemically with T . cruzi . (A) Greater absolute numbers of CD8 + T cells were recovered from mice co-adoptively transferred with Th17 cells at 7 days post-infection. (B) The mean fluorescence intensity (MFI) of T-bet expression in these same CD8 + T cells was increased over mice receiving CD8 + T cells alone or CD8 + T cells with Th1 cells, as measured by ICS. (C) Co-culture of sub-optimally stimulated CD8 + T cells with activated Th17 cell SN in vitro also induced higher T-bet expression than co-culture with Th1 cell SN. (D-E) Antigen-specific total and CD8 + T cell responses were studied 7 days post-challenge by IFN-γ ELISPOT (D) and intracellular cytokine staining (E), respectively. Similar results were detected in multiple experiments.

Article Snippet: In vitro generated Th1 or Th17 cells were transferred i.v. into RAG KO mice, along with polyclonal CD8 + T cells purified from naïve BALB/c mice by positive selection using α-CD8 microbeads (Miltenyi Biotec, Auburn, CA).

Techniques: Infection, Fluorescence, Expressing, Co-Culture Assay, In Vitro, Enzyme-linked Immunospot, Staining

Journal: PLoS Pathogens

Article Title: Th17 Cells Are More Protective Than Th1 Cells Against the Intracellular Parasite Trypanosoma cruzi

doi: 10.1371/journal.ppat.1005902

Figure Lengend Snippet: Total spleen cells (A) and purified CD8 + T cells (B) recovered from WT BALB/c mice receiving TS-CD4-Tg Th17 cell adoptive transfer developed 3- to 4-fold higher levels of IFN-γ producing T . cruzi -specific T cells detectable responses upon stimulation with either p7 or whole TS antigen. (C-D) Adoptive transfer of TS-CD4-Tg Th1 cells resulted in an approximately 3-fold increase in antibody titers, and adoptive transfer of Th17 cells resulted in an approximately 9-fold increase, as measured by endpoint titers.

Article Snippet: In vitro generated Th1 or Th17 cells were transferred i.v. into RAG KO mice, along with polyclonal CD8 + T cells purified from naïve BALB/c mice by positive selection using α-CD8 microbeads (Miltenyi Biotec, Auburn, CA).

Techniques: Purification, Adoptive Transfer Assay