anti mhc class ii  (Thermo Fisher)


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    Structured Review

    Thermo Fisher anti mhc class ii
    DC-STAMP silencing does not induce phenotypic changes in mBMDCs . Murine BMDCs were infected with shScr and shST1 lentivirus at day 6 and day 7 of culture. (A) Light field microscopy image of mBMDCs in culture 4 days post-infection. (B) Lentiviraly transduced mBMDCs stained with antibody against calreticulin (green) to visualize ER and DAPI staining of nucleus (blue) analyzed by CLSM. (C) Flow cytometric analysis of CD80, <t>CD86</t> and <t>MHC</t> class II surface expression in CD11c + immature (-LPS) and LPS-matured (+LPS) non-infected (no virus), shScr and shST1 mBMDCs. Isotype controls are indicated by shaded area. Data shown are representative of two or three independent experiments.
    Anti Mhc Class Ii, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 93/100, based on 40 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Images

    1) Product Images from "DC-STAMP knock-down deregulates cytokine production and T-cell stimulatory capacity of LPS-matured dendritic cells"

    Article Title: DC-STAMP knock-down deregulates cytokine production and T-cell stimulatory capacity of LPS-matured dendritic cells

    Journal: BMC Immunology

    doi: 10.1186/1471-2172-12-57

    DC-STAMP silencing does not induce phenotypic changes in mBMDCs . Murine BMDCs were infected with shScr and shST1 lentivirus at day 6 and day 7 of culture. (A) Light field microscopy image of mBMDCs in culture 4 days post-infection. (B) Lentiviraly transduced mBMDCs stained with antibody against calreticulin (green) to visualize ER and DAPI staining of nucleus (blue) analyzed by CLSM. (C) Flow cytometric analysis of CD80, CD86 and MHC class II surface expression in CD11c + immature (-LPS) and LPS-matured (+LPS) non-infected (no virus), shScr and shST1 mBMDCs. Isotype controls are indicated by shaded area. Data shown are representative of two or three independent experiments.
    Figure Legend Snippet: DC-STAMP silencing does not induce phenotypic changes in mBMDCs . Murine BMDCs were infected with shScr and shST1 lentivirus at day 6 and day 7 of culture. (A) Light field microscopy image of mBMDCs in culture 4 days post-infection. (B) Lentiviraly transduced mBMDCs stained with antibody against calreticulin (green) to visualize ER and DAPI staining of nucleus (blue) analyzed by CLSM. (C) Flow cytometric analysis of CD80, CD86 and MHC class II surface expression in CD11c + immature (-LPS) and LPS-matured (+LPS) non-infected (no virus), shScr and shST1 mBMDCs. Isotype controls are indicated by shaded area. Data shown are representative of two or three independent experiments.

    Techniques Used: Infection, Microscopy, Staining, Confocal Laser Scanning Microscopy, Flow Cytometry, Expressing

    2) Product Images from "An Epithelial Integrin Regulates the Amplitude of Protective Lung Interferon Responses against Multiple Respiratory Pathogens"

    Article Title: An Epithelial Integrin Regulates the Amplitude of Protective Lung Interferon Responses against Multiple Respiratory Pathogens

    Journal: PLoS Pathogens

    doi: 10.1371/journal.ppat.1005804

    Exogenous TGF-β1 reverses macrophage phenotype and reverses protection from influenza infection. (A) Active TGF-β1 was quantified by ELISA in WT (Jackson labs) or β6 KO mice at homeostasis. (B) Experimental design describing the treatment of mice with exogenous TGF-β1. (C) Macrophages were isolated from the lungs of uninfected WT or β6 KO mice treated with or without TGF-β1 using the gating strategy shown in S2A Fig . Data reflects 2 independent experiments with n = 2–6 mice per group. (D) Quantification of CD11c + CD11b - macrophages in mice treated with exogenous TGF-β1. Data reflects 2 independent experiments with n = 2–6 mice per group. ****p
    Figure Legend Snippet: Exogenous TGF-β1 reverses macrophage phenotype and reverses protection from influenza infection. (A) Active TGF-β1 was quantified by ELISA in WT (Jackson labs) or β6 KO mice at homeostasis. (B) Experimental design describing the treatment of mice with exogenous TGF-β1. (C) Macrophages were isolated from the lungs of uninfected WT or β6 KO mice treated with or without TGF-β1 using the gating strategy shown in S2A Fig . Data reflects 2 independent experiments with n = 2–6 mice per group. (D) Quantification of CD11c + CD11b - macrophages in mice treated with exogenous TGF-β1. Data reflects 2 independent experiments with n = 2–6 mice per group. ****p

    Techniques Used: Infection, Enzyme-linked Immunosorbent Assay, Mouse Assay, Isolation

    Altered alveolar macrophage phenotype in the lungs of β6 KO mice. (A) Cells were isolated from the lungs of uninfected WT or β6 KO mice and gated on F4/80 + cells then divided into macrophage and DC populations by CD11b and CD11c expression. (B) Quantification of the percentage of autofluorescent F4/80 + CD11c + CD11b + cells. p
    Figure Legend Snippet: Altered alveolar macrophage phenotype in the lungs of β6 KO mice. (A) Cells were isolated from the lungs of uninfected WT or β6 KO mice and gated on F4/80 + cells then divided into macrophage and DC populations by CD11b and CD11c expression. (B) Quantification of the percentage of autofluorescent F4/80 + CD11c + CD11b + cells. p

    Techniques Used: Mouse Assay, Isolation, Expressing

    3) Product Images from "MMP-9 Inhibits IL-23p19 Expression in Dendritic Cells By Targeting Membrane Stem Cell Factor Affecting Lung IL-17 Response"

    Article Title: MMP-9 Inhibits IL-23p19 Expression in Dendritic Cells By Targeting Membrane Stem Cell Factor Affecting Lung IL-17 Response

    Journal: Journal of immunology (Baltimore, Md. : 1950)

    doi: 10.4049/jimmunol.1303183

    MMP-9 enzymatic activity reduces mSCF leading to lower DC IL-23p19. Lung CD11c + cells were isolated by enzymatic digestion and magnetic bead separation after CT or CpG. Alveolar macrophages were gated out using combined autofluorescence and Siglec F staining. ( A ) Remaining DCs were analyzed for the indicated markers to delineate major sub-populations (left hand panels). The right-hand panels demonstrate that c-kit + DCs are MAR-1 + /CD64 + inflammatory moDCs. ( B ) IL-23p19 mRNA expression in sorted MAR-1 + /CD64 + c-kit + DCs. ( C ) Intracellular cytokine staining for IL-23p19 protein in MHC Class II high /CD11b + /c-kit + DCs. ( D ) Immunoblotting for mSCF on CD11c + lung cells that would include both DCs and lung macrophages from WT and MMP-9 −/− mice. ( E ) Gel zymography of total lung extracts following CT or CpG treatment of MMP-9 −/− or WT animals. Data in panels A-D are derived from pooled groups of 3 mice each. Each experiment was performed a minimum of 3 times yielding similar results.
    Figure Legend Snippet: MMP-9 enzymatic activity reduces mSCF leading to lower DC IL-23p19. Lung CD11c + cells were isolated by enzymatic digestion and magnetic bead separation after CT or CpG. Alveolar macrophages were gated out using combined autofluorescence and Siglec F staining. ( A ) Remaining DCs were analyzed for the indicated markers to delineate major sub-populations (left hand panels). The right-hand panels demonstrate that c-kit + DCs are MAR-1 + /CD64 + inflammatory moDCs. ( B ) IL-23p19 mRNA expression in sorted MAR-1 + /CD64 + c-kit + DCs. ( C ) Intracellular cytokine staining for IL-23p19 protein in MHC Class II high /CD11b + /c-kit + DCs. ( D ) Immunoblotting for mSCF on CD11c + lung cells that would include both DCs and lung macrophages from WT and MMP-9 −/− mice. ( E ) Gel zymography of total lung extracts following CT or CpG treatment of MMP-9 −/− or WT animals. Data in panels A-D are derived from pooled groups of 3 mice each. Each experiment was performed a minimum of 3 times yielding similar results.

    Techniques Used: Activity Assay, Isolation, Staining, Expressing, Mouse Assay, Zymography, Derivative Assay

    4) Product Images from "Maturation of dendritic cells by pullulan promotes anti-cancer effect"

    Article Title: Maturation of dendritic cells by pullulan promotes anti-cancer effect

    Journal: Oncotarget

    doi: 10.18632/oncotarget.10183

    Pullulan promotes antigen presentation and antigen-specific T cell proliferation in vivo A. C57BL/6 mice were injected with PBS, OVA and combination of OVA and pullulan for 24 hours. The expression levels of MHC class I and II on the gated Lineage − CD8α + CD11c + and Lineage − CD8α − CD11c + cDCs in splenocytes from these mice were analyzed. B. Purified CD8 T cells from OT-I or CD4 T cells from OT-II mice were labeled with CFSE and transferred into CD45.1 congenic mice; 24 hours later, mice were injected with PBS, OVA, pullulan, and combination of OVA and pullulan. After 3 days of treatment, splenocytes from these mice were stained for CD45.2 to identify the donor OT-I or OT-II cells. The proliferation of these cells was determined by CFSE dilution. C and D. C57BL/6 mice were treated i.v. with PBS, 50 μg OVA, 25 mg/kg pullulan, and the combination of OVA and pullulan on days 0, 7, and 14. (C) On day 21 after treatment, frequencies and functional activities of OVA (257-264)-specific spleen CD8 T cells were analyzed ex vivo by tetramer staining (left panel). Mean percentage of tetramer positive cells in CD8 T cells (right panel). D. SIINFEKL-specific IFN-γ-producing cell were analyzed by ELISPOT assay. All data are from analyses of 6 individual mice each group (2 mice per experiment, total 3 independent experiments).
    Figure Legend Snippet: Pullulan promotes antigen presentation and antigen-specific T cell proliferation in vivo A. C57BL/6 mice were injected with PBS, OVA and combination of OVA and pullulan for 24 hours. The expression levels of MHC class I and II on the gated Lineage − CD8α + CD11c + and Lineage − CD8α − CD11c + cDCs in splenocytes from these mice were analyzed. B. Purified CD8 T cells from OT-I or CD4 T cells from OT-II mice were labeled with CFSE and transferred into CD45.1 congenic mice; 24 hours later, mice were injected with PBS, OVA, pullulan, and combination of OVA and pullulan. After 3 days of treatment, splenocytes from these mice were stained for CD45.2 to identify the donor OT-I or OT-II cells. The proliferation of these cells was determined by CFSE dilution. C and D. C57BL/6 mice were treated i.v. with PBS, 50 μg OVA, 25 mg/kg pullulan, and the combination of OVA and pullulan on days 0, 7, and 14. (C) On day 21 after treatment, frequencies and functional activities of OVA (257-264)-specific spleen CD8 T cells were analyzed ex vivo by tetramer staining (left panel). Mean percentage of tetramer positive cells in CD8 T cells (right panel). D. SIINFEKL-specific IFN-γ-producing cell were analyzed by ELISPOT assay. All data are from analyses of 6 individual mice each group (2 mice per experiment, total 3 independent experiments).

    Techniques Used: In Vivo, Mouse Assay, Injection, Expressing, Purification, Labeling, Staining, Functional Assay, Ex Vivo, Enzyme-linked Immunospot

    5) Product Images from "Mycobacterium tuberculosis Rv2005c Induces Dendritic Cell Maturation and Th1 Responses and Exhibits Immunotherapeutic Activity by Fusion with the Rv2882c Protein"

    Article Title: Mycobacterium tuberculosis Rv2005c Induces Dendritic Cell Maturation and Th1 Responses and Exhibits Immunotherapeutic Activity by Fusion with the Rv2882c Protein

    Journal: Vaccines

    doi: 10.3390/vaccines8030370

    Recombinant Rv2005c induces dendritic cell (DC) maturation through the MAPK pathway. ( A ) The purified, recombinant Rv2005c protein was subjected to ( A ) Coomassie blue staining sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and ( B ) western blot analysis using a mouse anti-His Ab. DCs were activated with the indicated concentration of Rv2005c or LPS (100 ng/mL) for 24 h. ( B ) Activated DCs were stained with anti-CD40, anti-CD80, anti-CD86, or anti-MHC class II Ab, and the expression of these surface markers was analyzed. The median fluorescence intensity (MFI) of the positive cells is shown for each panel. The bar graphs show the mean ± SEM ( n = 5). ( C ) The TNF-α, IL-1β, and IL-12p70 levels in the culture medium were measured by ELISA. The data are presented as the mean ± SEM ( n = 5). ( D ) The protein production by DCs treated with Rv2005c for the indicated periods was analyzed by immunoblotting using each specific Ab: phospho-p38 (p-p38), p38, phospho-ERK1/2 (p-ERK1/2), phospho-IκB-α, and IκB-α. ( E ) DCs were pretreated with pharmacological inhibitors of p38 (SB203580, 20 μM), ERK1/2 (U0126, 10 μM), JNK (SP600125, 20 μM), Bay11-7082 (20 μM), or DMSO (vehicle control) for 1 h prior to treatment with 10 μg/mL Rv2005c protein for 24 h. The cytokine levels in the culture supernatants were measured by ELISA. The data shown are the mean ± SEM; * p
    Figure Legend Snippet: Recombinant Rv2005c induces dendritic cell (DC) maturation through the MAPK pathway. ( A ) The purified, recombinant Rv2005c protein was subjected to ( A ) Coomassie blue staining sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and ( B ) western blot analysis using a mouse anti-His Ab. DCs were activated with the indicated concentration of Rv2005c or LPS (100 ng/mL) for 24 h. ( B ) Activated DCs were stained with anti-CD40, anti-CD80, anti-CD86, or anti-MHC class II Ab, and the expression of these surface markers was analyzed. The median fluorescence intensity (MFI) of the positive cells is shown for each panel. The bar graphs show the mean ± SEM ( n = 5). ( C ) The TNF-α, IL-1β, and IL-12p70 levels in the culture medium were measured by ELISA. The data are presented as the mean ± SEM ( n = 5). ( D ) The protein production by DCs treated with Rv2005c for the indicated periods was analyzed by immunoblotting using each specific Ab: phospho-p38 (p-p38), p38, phospho-ERK1/2 (p-ERK1/2), phospho-IκB-α, and IκB-α. ( E ) DCs were pretreated with pharmacological inhibitors of p38 (SB203580, 20 μM), ERK1/2 (U0126, 10 μM), JNK (SP600125, 20 μM), Bay11-7082 (20 μM), or DMSO (vehicle control) for 1 h prior to treatment with 10 μg/mL Rv2005c protein for 24 h. The cytokine levels in the culture supernatants were measured by ELISA. The data shown are the mean ± SEM; * p

    Techniques Used: Recombinant, Purification, Staining, Polyacrylamide Gel Electrophoresis, SDS Page, Western Blot, Concentration Assay, Expressing, Fluorescence, Enzyme-linked Immunosorbent Assay

    6) Product Images from "Contributions of IFN-γ and granulysin to the clearance of Plasmodium yoelii blood stage"

    Article Title: Contributions of IFN-γ and granulysin to the clearance of Plasmodium yoelii blood stage

    Journal: PLoS Pathogens

    doi: 10.1371/journal.ppat.1008840

    P . yoelii -infected Retics express MHC class I. (A) Representative histograms of MHC-I expression on RBCs at different stages of maturation in uninfected mice (blue) and P . yoelii -infected mice (black) at 12DPI. (B) P . yoelii -infected Retics (iRetics) from blood, spleen and bone marrow express MHC class I on their surface. Data are pooled from two independent experiments (n = 5, uninfected mice; n = 7, P . yoelii -infected mice). (C) iRetics (filled circles) from bloodstream express higher levels of MHC-I than uninfected Retics (uRetics; open circles) from P . yoelii -infected mice. (D) Imaging flow cytometry of Retics purified from the blood of P . yoelii -infected mice confirm the difference in MHC-I expression between iRetics and uRetics. (E, F) Circulating RBCs do not express MHC class II or CD86 in their surface (n = 3, uninfected mice; n = 4 P . yoelii -infected mice). (G, H) Erythroblasts express IFN-γR on their surface (n = 3, uninfected mice; n = 5 P . yoelii -infected mice). Statistical analysis of MHC-I and IFN-γR expression was carried out using unpaired t-test or Mann-Whitney U test, according to the data distribution. **p
    Figure Legend Snippet: P . yoelii -infected Retics express MHC class I. (A) Representative histograms of MHC-I expression on RBCs at different stages of maturation in uninfected mice (blue) and P . yoelii -infected mice (black) at 12DPI. (B) P . yoelii -infected Retics (iRetics) from blood, spleen and bone marrow express MHC class I on their surface. Data are pooled from two independent experiments (n = 5, uninfected mice; n = 7, P . yoelii -infected mice). (C) iRetics (filled circles) from bloodstream express higher levels of MHC-I than uninfected Retics (uRetics; open circles) from P . yoelii -infected mice. (D) Imaging flow cytometry of Retics purified from the blood of P . yoelii -infected mice confirm the difference in MHC-I expression between iRetics and uRetics. (E, F) Circulating RBCs do not express MHC class II or CD86 in their surface (n = 3, uninfected mice; n = 4 P . yoelii -infected mice). (G, H) Erythroblasts express IFN-γR on their surface (n = 3, uninfected mice; n = 5 P . yoelii -infected mice). Statistical analysis of MHC-I and IFN-γR expression was carried out using unpaired t-test or Mann-Whitney U test, according to the data distribution. **p

    Techniques Used: Infection, Expressing, Mouse Assay, Imaging, Flow Cytometry, Purification, MANN-WHITNEY

    MHC class I expression on erythroblasts and Retics depends on IFN-γ. (A, B) Representative histograms of MHC-I expression on erythroblasts and Retics from P . yoelii -infected WT (grey) and IFN-γ KO (red) mice at 12 DPI. (C) Erythroblasts from spleen and bone marrow of P . yoelii -infected WT (black circle) express higher levels of MHC-I in comparison to IFN-γ KO (red circles) mice. Note that the frequency of erythroblasts in spleen of uninfected mice is below the limit of detection. (D) iRetics and uRetics from infected IFN-γ KO mice (red circles) express very low levels of MHC-I. (E) CD71 and MHC-I MFI correlation in erythroblast from uninfected mice (blue symbols) and P . yoelii -infected mice from bone marrow [filled black (WT) and red (IFN-γ KO) symbols] and spleen [half open black (WT) and red (IFN-γ KO) symbols]. (F) Retics from uninfected mice (blue symbols) and iRetics [filled black (WT) and red (IFN-γ KO) symbols] and uRetics [open black (WT) and red (IFN-γ KO) symbols] from P . yoelii -infected mice. Note that MHC-I, but not CD71 expression is impaired both in erythroblasts and Retics from infected IFN-γ KO mice. (G) Imaging flow cytometry of iRetics purified from blood confirm the difference in MHC-I expression between WT and IFN-γ KO mice. (H) Comparison of percentage parasitemia in RBCs from IFN-γ KO mice (red line) and WT mice (black line). (I) All IFN-γ KO mice (red line) succumbed to P . yoelii infection between 21 and 26 DPI. The statistical analysis for MHC-I was performed using unpaired t-test or Mann-Whitney U test, according to data distribution. Data are pooled from two independent experiments (n = 3–5, uninfected mice; n = 4–7, P . yoelii -infected). The statistical analysis of parasitemia was carried out using two-way ANOVA followed by Bonferroni post-hoc test. The statistical analysis of survival was performed using the log-rank test. Data are pooled from two independent experiments (n = 12, for each group). **p
    Figure Legend Snippet: MHC class I expression on erythroblasts and Retics depends on IFN-γ. (A, B) Representative histograms of MHC-I expression on erythroblasts and Retics from P . yoelii -infected WT (grey) and IFN-γ KO (red) mice at 12 DPI. (C) Erythroblasts from spleen and bone marrow of P . yoelii -infected WT (black circle) express higher levels of MHC-I in comparison to IFN-γ KO (red circles) mice. Note that the frequency of erythroblasts in spleen of uninfected mice is below the limit of detection. (D) iRetics and uRetics from infected IFN-γ KO mice (red circles) express very low levels of MHC-I. (E) CD71 and MHC-I MFI correlation in erythroblast from uninfected mice (blue symbols) and P . yoelii -infected mice from bone marrow [filled black (WT) and red (IFN-γ KO) symbols] and spleen [half open black (WT) and red (IFN-γ KO) symbols]. (F) Retics from uninfected mice (blue symbols) and iRetics [filled black (WT) and red (IFN-γ KO) symbols] and uRetics [open black (WT) and red (IFN-γ KO) symbols] from P . yoelii -infected mice. Note that MHC-I, but not CD71 expression is impaired both in erythroblasts and Retics from infected IFN-γ KO mice. (G) Imaging flow cytometry of iRetics purified from blood confirm the difference in MHC-I expression between WT and IFN-γ KO mice. (H) Comparison of percentage parasitemia in RBCs from IFN-γ KO mice (red line) and WT mice (black line). (I) All IFN-γ KO mice (red line) succumbed to P . yoelii infection between 21 and 26 DPI. The statistical analysis for MHC-I was performed using unpaired t-test or Mann-Whitney U test, according to data distribution. Data are pooled from two independent experiments (n = 3–5, uninfected mice; n = 4–7, P . yoelii -infected). The statistical analysis of parasitemia was carried out using two-way ANOVA followed by Bonferroni post-hoc test. The statistical analysis of survival was performed using the log-rank test. Data are pooled from two independent experiments (n = 12, for each group). **p

    Techniques Used: Expressing, Infection, Mouse Assay, Imaging, Flow Cytometry, Purification, MANN-WHITNEY

    7) Product Images from "SND1 promotes Th1/17 immunity against chlamydial lung infection through enhancing dendritic cell function"

    Article Title: SND1 promotes Th1/17 immunity against chlamydial lung infection through enhancing dendritic cell function

    Journal: PLoS Pathogens

    doi: 10.1371/journal.ppat.1009295

    Impaired function of conventional DCs in lung of SND1 -/- mice compared with that of control mice following Cm lung infection. Lung collected at day 7 p.i. were analyzed for population of conventional DCs by surface staining, as described in Materials and Methods . (A) MHC-II + CD11c + conventional DCs in lung. (B) Two pulmonary DC subsets of CD11b + CD103 - DCs and CD11b - CD103 + DCs gated at the population of CD11c + MHC-II + conventional DCs. (C) Population of IL-12-producing CD11c + DCs gated from CD103 + CD11b - pulmonary DC subset. (D) Population of IL-12-producing CD11c + DCs gated from CD103 - CD11b + pulmonary DC subset. (E) Absolute cell number of total pulmonary cells in four groups. (F) Absolute cell number of conventional DCs and two pulmonary DC subsets in four groups. (G) Absolute cell number of IL-12-producing CD103 + CD11b - pulmonary DC subset. (H) Absolute cell number of IL-12-producing CD103 - CD11b + pulmonary DC subset. One representative experiment of three independent experiments with four mice in each group is shown. Data are shown as the mean±SD. *, p
    Figure Legend Snippet: Impaired function of conventional DCs in lung of SND1 -/- mice compared with that of control mice following Cm lung infection. Lung collected at day 7 p.i. were analyzed for population of conventional DCs by surface staining, as described in Materials and Methods . (A) MHC-II + CD11c + conventional DCs in lung. (B) Two pulmonary DC subsets of CD11b + CD103 - DCs and CD11b - CD103 + DCs gated at the population of CD11c + MHC-II + conventional DCs. (C) Population of IL-12-producing CD11c + DCs gated from CD103 + CD11b - pulmonary DC subset. (D) Population of IL-12-producing CD11c + DCs gated from CD103 - CD11b + pulmonary DC subset. (E) Absolute cell number of total pulmonary cells in four groups. (F) Absolute cell number of conventional DCs and two pulmonary DC subsets in four groups. (G) Absolute cell number of IL-12-producing CD103 + CD11b - pulmonary DC subset. (H) Absolute cell number of IL-12-producing CD103 - CD11b + pulmonary DC subset. One representative experiment of three independent experiments with four mice in each group is shown. Data are shown as the mean±SD. *, p

    Techniques Used: Mouse Assay, Infection, Staining

    Impaired function of conventional DCs in spleen of SND1 -/- mice compared with that of control mice following Cm lung infection. Spleen collected at day 7 p.i. were analyzed for population of conventional DCs by surface staining, as described in Materials and Methods . (A) MHC-II + CD11c + conventional DC in spleen. (B) Two splenic DC subsets of CD8 + CD11b - DCs and CD8 - CD11b + DCs gated at the population of CD11c + MHC-II + conventional DCs. (C) Population of IL-12-producing CD11c + DCs gated from CD8 + CD11b - splenic DC subset. (D) Population of IL-12-producing CD11c + DCs gated from CD8 - CD11b + splenic DC subset. (E) Absolute cell number of total splenic cells in four groups. (F) Absolute cell number of conventional DC and two splenic DC subsets in four groups. (G and I) Absolute cell number and percentage of IL-12-producing CD8 + CD11b - splenic DC subset. (H and J) Absolute cell number and percentage of IL-12-producing CD8 - CD11b + splenic DC subset. One representative experiment of three independent experiments with four mice in each group is shown. Data are shown as the mean±SD. *, p
    Figure Legend Snippet: Impaired function of conventional DCs in spleen of SND1 -/- mice compared with that of control mice following Cm lung infection. Spleen collected at day 7 p.i. were analyzed for population of conventional DCs by surface staining, as described in Materials and Methods . (A) MHC-II + CD11c + conventional DC in spleen. (B) Two splenic DC subsets of CD8 + CD11b - DCs and CD8 - CD11b + DCs gated at the population of CD11c + MHC-II + conventional DCs. (C) Population of IL-12-producing CD11c + DCs gated from CD8 + CD11b - splenic DC subset. (D) Population of IL-12-producing CD11c + DCs gated from CD8 - CD11b + splenic DC subset. (E) Absolute cell number of total splenic cells in four groups. (F) Absolute cell number of conventional DC and two splenic DC subsets in four groups. (G and I) Absolute cell number and percentage of IL-12-producing CD8 + CD11b - splenic DC subset. (H and J) Absolute cell number and percentage of IL-12-producing CD8 - CD11b + splenic DC subset. One representative experiment of three independent experiments with four mice in each group is shown. Data are shown as the mean±SD. *, p

    Techniques Used: Mouse Assay, Infection, Staining

    8) Product Images from "Dendritic cells and B cells maximize mucosal Th1 memory response to herpes simplex virus"

    Article Title: Dendritic cells and B cells maximize mucosal Th1 memory response to herpes simplex virus

    Journal: The Journal of Experimental Medicine

    doi: 10.1084/jem.20082039

    Conventional DCs are not required but are partially sufficient to elicit recall Th1 responses in the vagina. (A and B) TK − HSV-2-immunized CD11c-DTR→WT chimeras ( n = 5) were inoculated with DT or PBS i.p., as shown in Fig. S9 (available at http://www.jem.org/cgi/content/full/jem.20082039/DC1 ). Depletion of vaginal DCs (CD11c + GFP + and MHC II + GFP + ) in the epithelium and lamina propria after DT injection was examined by FACS analysis (A). Viral titers in vaginal secretion were measured at the indicated days after challenge (B). (C–F) To examine the sufficiency of MHC class II expression on CD11c + cells for protection against WT HSV-2 secondary challenge, Abb mice and MHC class II −/− mice were transplanted with naive polyclonal CD4 T cells (10 7 cells) 1 d before TK − HSV-2 immunization (C). MHC class II expression on vaginal DCs in TK − HSV-2–immunized mice (4 wk) was analyzed (D). Frozen sections were stained with MHC class II (green) and counterstained with DAPI (blue). Images were captured using a 10 (left) or 40× (right) objective. Bars, 100 μm. (E) Adoptively transferred naive CD4 T cells normally differentiate into Th1 cells in Abb mice immunized with TK − HSV-2. CD4 T cells isolated from the draining LNs of Abb, MHC class II −/− (MHCII −/− ), and C57BL6 mice (WT) mice immunized ivag with TK − HSV-2 (7 d after infection) were cocultured with syngeneic splenocytes in the presence of HSV-2 antigens (left) or mock antigen (right) and analyzed for IFN-γ secretion (in nanograms/milliliter). Each column represents the mean ± SD of triplicate samples. (F) 3 wk after TK − immunization, Abb and MHC class II −/− mice were challenged with 10 4 PFU WT HSV-2 virus. Viral titers in vaginal secretion were measured at the indicated days after challenge. Error bars represent the mean ± SD. These data are representative of two similar experiments.
    Figure Legend Snippet: Conventional DCs are not required but are partially sufficient to elicit recall Th1 responses in the vagina. (A and B) TK − HSV-2-immunized CD11c-DTR→WT chimeras ( n = 5) were inoculated with DT or PBS i.p., as shown in Fig. S9 (available at http://www.jem.org/cgi/content/full/jem.20082039/DC1 ). Depletion of vaginal DCs (CD11c + GFP + and MHC II + GFP + ) in the epithelium and lamina propria after DT injection was examined by FACS analysis (A). Viral titers in vaginal secretion were measured at the indicated days after challenge (B). (C–F) To examine the sufficiency of MHC class II expression on CD11c + cells for protection against WT HSV-2 secondary challenge, Abb mice and MHC class II −/− mice were transplanted with naive polyclonal CD4 T cells (10 7 cells) 1 d before TK − HSV-2 immunization (C). MHC class II expression on vaginal DCs in TK − HSV-2–immunized mice (4 wk) was analyzed (D). Frozen sections were stained with MHC class II (green) and counterstained with DAPI (blue). Images were captured using a 10 (left) or 40× (right) objective. Bars, 100 μm. (E) Adoptively transferred naive CD4 T cells normally differentiate into Th1 cells in Abb mice immunized with TK − HSV-2. CD4 T cells isolated from the draining LNs of Abb, MHC class II −/− (MHCII −/− ), and C57BL6 mice (WT) mice immunized ivag with TK − HSV-2 (7 d after infection) were cocultured with syngeneic splenocytes in the presence of HSV-2 antigens (left) or mock antigen (right) and analyzed for IFN-γ secretion (in nanograms/milliliter). Each column represents the mean ± SD of triplicate samples. (F) 3 wk after TK − immunization, Abb and MHC class II −/− mice were challenged with 10 4 PFU WT HSV-2 virus. Viral titers in vaginal secretion were measured at the indicated days after challenge. Error bars represent the mean ± SD. These data are representative of two similar experiments.

    Techniques Used: Injection, FACS, Expressing, Mouse Assay, Staining, Isolation, Infection

    Direct recognition of the infected vaginal epithelial cells by Th1 cells is not required for protection. C57BL/6 (WT; n = 7), MHC class II −/− ( n = 4), WT thymus-transplanted MHC class II −/− BM chimera, and WT→{MHC II −/− + WT thymus} ( n = 9) were immunized with TK − HSV-2 ivag and, 4 wk later, challenged with lethal WT HSV-2. As a control, nonimmunized C57/BL6 mice ( n = 6) were challenged with the virus. (A) FACS analyses of CD4 and CD8 T cells (live CD3 + cells) in the peripheral blood of C57BL/6 (WT)→WT, MHC class II −/− , and WT→{MHC II −/− + WT thymus} mice. (B–D) Viral titers in vaginal washes (B), survival (C), and genital mean pathology scores (D) after secondary challenge were examined. Error bars represent the mean ± SE of the number of mice per group. These data are representative of three similar experiments.
    Figure Legend Snippet: Direct recognition of the infected vaginal epithelial cells by Th1 cells is not required for protection. C57BL/6 (WT; n = 7), MHC class II −/− ( n = 4), WT thymus-transplanted MHC class II −/− BM chimera, and WT→{MHC II −/− + WT thymus} ( n = 9) were immunized with TK − HSV-2 ivag and, 4 wk later, challenged with lethal WT HSV-2. As a control, nonimmunized C57/BL6 mice ( n = 6) were challenged with the virus. (A) FACS analyses of CD4 and CD8 T cells (live CD3 + cells) in the peripheral blood of C57BL/6 (WT)→WT, MHC class II −/− , and WT→{MHC II −/− + WT thymus} mice. (B–D) Viral titers in vaginal washes (B), survival (C), and genital mean pathology scores (D) after secondary challenge were examined. Error bars represent the mean ± SE of the number of mice per group. These data are representative of three similar experiments.

    Techniques Used: Infection, Mouse Assay, FACS

    Memory CD4 T cells localize in the vagina of immune mice and provide protection against secondary HSV-2 challenge. (A and B) The localization of CD4 and CD8 T cells in the vagina of nonimmunized (A) or TK − HSV-2–immunized mice at 3 wk after immunization (B). (C and D) Congenic (CD45.1 + ) effector CD4, CD8, or naive CD4 T cells were adoptively transferred into naive recipient (CD45.2 + ) mice. At 3 d after ivag infection with HSV-2, viral antigen (anti–HSV-2, green; C and D), transferred T cells (anti-CD45.1, red; C and D), and MHC class II (blue; D) were visualized. Images were captured using a 10 (A–C) or 40× (D) objective lens. Arrowheads indicate the basement membrane. Bars, 100 μm. (E and F) CD4 or CD8 T cells were depleted from TK − HSV-2–primed mice and challenged ivag with WT HSV-2. (E) The dot plot represents CD4 + and CD8 + cells in vagina 3 d after challenge. (F) Virus titers in the vaginal fluids (nonimmune, n = 4; immune/control Ab, n = 6; immune/anti-CD4, n = 4; and immune/anti-CD8, n = 4) were measured at the indicated days after HSV-2 secondary challenge. Error bars represent the mean ± SD of the number of mice per group. These data are representative of three similar experiments.
    Figure Legend Snippet: Memory CD4 T cells localize in the vagina of immune mice and provide protection against secondary HSV-2 challenge. (A and B) The localization of CD4 and CD8 T cells in the vagina of nonimmunized (A) or TK − HSV-2–immunized mice at 3 wk after immunization (B). (C and D) Congenic (CD45.1 + ) effector CD4, CD8, or naive CD4 T cells were adoptively transferred into naive recipient (CD45.2 + ) mice. At 3 d after ivag infection with HSV-2, viral antigen (anti–HSV-2, green; C and D), transferred T cells (anti-CD45.1, red; C and D), and MHC class II (blue; D) were visualized. Images were captured using a 10 (A–C) or 40× (D) objective lens. Arrowheads indicate the basement membrane. Bars, 100 μm. (E and F) CD4 or CD8 T cells were depleted from TK − HSV-2–primed mice and challenged ivag with WT HSV-2. (E) The dot plot represents CD4 + and CD8 + cells in vagina 3 d after challenge. (F) Virus titers in the vaginal fluids (nonimmune, n = 4; immune/control Ab, n = 6; immune/anti-CD4, n = 4; and immune/anti-CD8, n = 4) were measured at the indicated days after HSV-2 secondary challenge. Error bars represent the mean ± SD of the number of mice per group. These data are representative of three similar experiments.

    Techniques Used: Mouse Assay, Infection

    9) Product Images from "Abrogation of self-tolerance by misfolded self-antigens complexed with MHC class II molecules"

    Article Title: Abrogation of self-tolerance by misfolded self-antigens complexed with MHC class II molecules

    Journal: Science Advances

    doi: 10.1126/sciadv.abj9867

    Mouse TSHR ECD is presented on the cell surface by mouse MHC class II molecules. ( A ) Mouse TSHR (mTSHR) ECD (shaded histograms) or mTSHR ECD with each mouse MHC class II allele (red lines) was transfected into HEK293T cells, respectively. The expression of MHC class II and TSHR is shown. ( B ) mTSHR ECD (shaded histograms), mTSHR ECD with I-A k (red lines), or mTSHR ECD with HEL peptide–attached I-A k (peptide-I-A k , blue lines) was transfected into HEK293T cells. The expression of I-A k and TSHR is shown. Mock: Empty pME18S plasmid. All data are representative of at least three independent experiments.
    Figure Legend Snippet: Mouse TSHR ECD is presented on the cell surface by mouse MHC class II molecules. ( A ) Mouse TSHR (mTSHR) ECD (shaded histograms) or mTSHR ECD with each mouse MHC class II allele (red lines) was transfected into HEK293T cells, respectively. The expression of MHC class II and TSHR is shown. ( B ) mTSHR ECD (shaded histograms), mTSHR ECD with I-A k (red lines), or mTSHR ECD with HEL peptide–attached I-A k (peptide-I-A k , blue lines) was transfected into HEK293T cells. The expression of I-A k and TSHR is shown. Mock: Empty pME18S plasmid. All data are representative of at least three independent experiments.

    Techniques Used: Transfection, Expressing, Plasmid Preparation

    TSHR forms a complex with MHC class II molecules. ( A ) Flag-tagged full-length human TSHR (Flag-TSHR) and His-tagged HLA-DP5 (HLA-DP5-His) were transfected into HEK293T cells. HLA-DP5 or TSHR was immunoprecipitated from the cell lysates and immunoblotted. ( B ) Tissue sections from GD patients ( n = 2) and non–GD patients ( n = 2) were stained with anti-TSHR Ab (green) and anti–HLA-DP Ab (red). Scale bars, 50 μm. ( C ) In situ association of TSHR with HLA-DP in tissue sections from GD patients ( n = 2) and non–GD patients ( n = 2) was analyzed via PLA. Photographs were taken at the same locations on consecutive tissue sections to compare the difference between the secondary Abs only and anti-TSHR/anti–HLA-DP. PLA signals and nuclei are shown as red and blue colors, respectively. Scale bars, 50 μm. ( D ) Bar plots of (C); the PLA signals per follicle are shown with the mean ± SD, multiple t tests, n = 12. **** P
    Figure Legend Snippet: TSHR forms a complex with MHC class II molecules. ( A ) Flag-tagged full-length human TSHR (Flag-TSHR) and His-tagged HLA-DP5 (HLA-DP5-His) were transfected into HEK293T cells. HLA-DP5 or TSHR was immunoprecipitated from the cell lysates and immunoblotted. ( B ) Tissue sections from GD patients ( n = 2) and non–GD patients ( n = 2) were stained with anti-TSHR Ab (green) and anti–HLA-DP Ab (red). Scale bars, 50 μm. ( C ) In situ association of TSHR with HLA-DP in tissue sections from GD patients ( n = 2) and non–GD patients ( n = 2) was analyzed via PLA. Photographs were taken at the same locations on consecutive tissue sections to compare the difference between the secondary Abs only and anti-TSHR/anti–HLA-DP. PLA signals and nuclei are shown as red and blue colors, respectively. Scale bars, 50 μm. ( D ) Bar plots of (C); the PLA signals per follicle are shown with the mean ± SD, multiple t tests, n = 12. **** P

    Techniques Used: Transfection, Immunoprecipitation, Staining, In Situ, Proximity Ligation Assay

    Autoantibodies are induced by mTSHR complexed with MHC class II molecules in mice. ( A ) DAP.3 cells expressing mTSHR alone (first row) or mTSHR and I-A k (second row) were stained with anti–I-A k mAb or K1-18 mAb (red), respectively. The GFP level is also shown (red). Shaded histograms are DAP.3 parental cells. ( B ) Timeline for the administration of DAP.3 transfectants into mice. i.p., intraperitoneally. ( C and D ) HEK293T cells transfected with mTSHR or I-A k were mixed with sera from (C) C3H/HeN mice immunized with DAP.3 cells expressing mTSHR alone (black, n = 10) or mTSHR and I-A k (red, n = 10), or (D) AKR/N mice immunized by DAP.3 cells expressing mTSHR alone (black, n = 10) or mTSHR and I-A k (red, n = 9). IgG Ab binding to TSHR transfectants was analyzed. The MFIs of Ab binding to mTSHR are represented as dots (one per mouse) with the mean ± SD, unpaired two-tailed Student’s t test. ( E ) TRAb in the mice immunized with DAP.3 cells expressing mTSHR alone (green bars) or mTSHR and I-A k (yellow bars). Data are represented as the mean ± SD of triplicate experiments, two-way analysis of variance (ANOVA). ( F ) HEK293T cells transfected with porcine TSHR (pTSHR) were mixed with PBS (gray bar), sera IgGs from mice immunized by DAP.3 cells expressing mTSHR alone (green bars), or mTSHR and I-A k (yellow bars), followed by staining with M22 mAb. The MFIs for the binding of M22 mAb to pTSHR are bar plotted as the mean ± SD of triplicate experiments, unpaired two-tailed Student’s t test. All MFIs were calculated by subtracting the MFI of Ab binding to that of GFP-negative transfectants. All data are representative of at least three independent experiments. * P
    Figure Legend Snippet: Autoantibodies are induced by mTSHR complexed with MHC class II molecules in mice. ( A ) DAP.3 cells expressing mTSHR alone (first row) or mTSHR and I-A k (second row) were stained with anti–I-A k mAb or K1-18 mAb (red), respectively. The GFP level is also shown (red). Shaded histograms are DAP.3 parental cells. ( B ) Timeline for the administration of DAP.3 transfectants into mice. i.p., intraperitoneally. ( C and D ) HEK293T cells transfected with mTSHR or I-A k were mixed with sera from (C) C3H/HeN mice immunized with DAP.3 cells expressing mTSHR alone (black, n = 10) or mTSHR and I-A k (red, n = 10), or (D) AKR/N mice immunized by DAP.3 cells expressing mTSHR alone (black, n = 10) or mTSHR and I-A k (red, n = 9). IgG Ab binding to TSHR transfectants was analyzed. The MFIs of Ab binding to mTSHR are represented as dots (one per mouse) with the mean ± SD, unpaired two-tailed Student’s t test. ( E ) TRAb in the mice immunized with DAP.3 cells expressing mTSHR alone (green bars) or mTSHR and I-A k (yellow bars). Data are represented as the mean ± SD of triplicate experiments, two-way analysis of variance (ANOVA). ( F ) HEK293T cells transfected with porcine TSHR (pTSHR) were mixed with PBS (gray bar), sera IgGs from mice immunized by DAP.3 cells expressing mTSHR alone (green bars), or mTSHR and I-A k (yellow bars), followed by staining with M22 mAb. The MFIs for the binding of M22 mAb to pTSHR are bar plotted as the mean ± SD of triplicate experiments, unpaired two-tailed Student’s t test. All MFIs were calculated by subtracting the MFI of Ab binding to that of GFP-negative transfectants. All data are representative of at least three independent experiments. * P

    Techniques Used: Mouse Assay, Expressing, Staining, Transfection, Binding Assay, Two Tailed Test

    10) Product Images from "Natural Killer Cell-activating receptor NKG2D mediates innate immune targeting of allogeneic neural progenitor cell grafts"

    Article Title: Natural Killer Cell-activating receptor NKG2D mediates innate immune targeting of allogeneic neural progenitor cell grafts

    Journal: Stem cells (Dayton, Ohio)

    doi: 10.1002/stem.1422

    Expression of MHC class I and NK cell activating ligands on NPC. A , Expression of MHC class I on NPC or splenocytes (positive control). B , Rae-1, MULT-1, and H60 expression on NPC or YAC-1 cells (positive control). C , NPC or YAC-1 were incubated with
    Figure Legend Snippet: Expression of MHC class I and NK cell activating ligands on NPC. A , Expression of MHC class I on NPC or splenocytes (positive control). B , Rae-1, MULT-1, and H60 expression on NPC or YAC-1 cells (positive control). C , NPC or YAC-1 were incubated with

    Techniques Used: Expressing, Positive Control, Incubation

    11) Product Images from "Basophils as a primary inducer of the T helper type 2 immunity in ovalbumin-induced allergic airway inflammation"

    Article Title: Basophils as a primary inducer of the T helper type 2 immunity in ovalbumin-induced allergic airway inflammation

    Journal: Immunology

    doi: 10.1111/imm.12240

    Lung-derived basophils expressed co-stimulatory molecules, uptake of DQ-ovalbumin (OVA), secretion of interleukin-4 (IL-4) and promotion of T helper type 2 (Th2) differentiation. The expression of MHC II, CD40, CD80 and CD86 by basophils, and DQ-OVA taken
    Figure Legend Snippet: Lung-derived basophils expressed co-stimulatory molecules, uptake of DQ-ovalbumin (OVA), secretion of interleukin-4 (IL-4) and promotion of T helper type 2 (Th2) differentiation. The expression of MHC II, CD40, CD80 and CD86 by basophils, and DQ-OVA taken

    Techniques Used: Derivative Assay, Expressing

    12) Product Images from "Fucoidan from Macrocystis pyrifera Has Powerful Immune-Modulatory Effects Compared to Three Other Fucoidans"

    Article Title: Fucoidan from Macrocystis pyrifera Has Powerful Immune-Modulatory Effects Compared to Three Other Fucoidans

    Journal: Marine Drugs

    doi: 10.3390/md13031084

    Administration of fucoidans induces spleen DC activation. C57BL/6 mice were injected i.v. with 50 mg/kg fucoidans for 24 h. ( A ) Percentage of DCs, defined as lineage − CD11c + , was analyzed by flow cytometry; ( B ) Absolute number of lineage − CD11c + cells within live cells was shown; ( C ) Expression levels of CD40, CD80, CD86, MHC class I and MHC class II were measured by flow cytometry (upper panel). Mean fluorescence intensity (MFI) of these molecules is shown (lower panel). Data are representative of or the average of analyses of six independent samples (two mice per experiment, total of three independent experiments); ( D ) Expression levels of IL-6, IL-12p40 and TNF-α mRNA were measured from spleen 2 h after 50 mg/kg fucoidans injection; ( E ) IL-6, IL-12p70 and TNF-α concentrations in sera were shown from fucoidan-treated mice at 24 h after treatment. Data are representative of or the average of analyses of six independent samples (two mice per experiment, total of three independent experiments). Data shown are the mean ± SEM. * p
    Figure Legend Snippet: Administration of fucoidans induces spleen DC activation. C57BL/6 mice were injected i.v. with 50 mg/kg fucoidans for 24 h. ( A ) Percentage of DCs, defined as lineage − CD11c + , was analyzed by flow cytometry; ( B ) Absolute number of lineage − CD11c + cells within live cells was shown; ( C ) Expression levels of CD40, CD80, CD86, MHC class I and MHC class II were measured by flow cytometry (upper panel). Mean fluorescence intensity (MFI) of these molecules is shown (lower panel). Data are representative of or the average of analyses of six independent samples (two mice per experiment, total of three independent experiments); ( D ) Expression levels of IL-6, IL-12p40 and TNF-α mRNA were measured from spleen 2 h after 50 mg/kg fucoidans injection; ( E ) IL-6, IL-12p70 and TNF-α concentrations in sera were shown from fucoidan-treated mice at 24 h after treatment. Data are representative of or the average of analyses of six independent samples (two mice per experiment, total of three independent experiments). Data shown are the mean ± SEM. * p

    Techniques Used: Activation Assay, Mouse Assay, Injection, Flow Cytometry, Cytometry, Expressing, Fluorescence

    13) Product Images from "Rehmannia glutinosa polysaccharide induces toll-like receptor 4 dependent spleen dendritic cell maturation and anti-cancer immunity"

    Article Title: Rehmannia glutinosa polysaccharide induces toll-like receptor 4 dependent spleen dendritic cell maturation and anti-cancer immunity

    Journal: Oncoimmunology

    doi: 10.1080/2162402X.2017.1325981

    RGP-induced activation spleen DCs were dependent on TLR4. C56BL/6 wild-type, TLR2-KO, TLR4-KO, and SR-A-KO mice were treated i.v. with PBS and 50 mg/kg RGP for 24 h. (A) Co-stimulatory molecules and MHC class I and II were measured in the spleen DCs. (B) The serum concentrations of IL-6, IL-12p40, and TNF-α after RGP treatment are shown. (C) The mRNA levels of IFNγ and T-bet in the spleen are shown. All data are the average of analyses of six independent samples (two mice per experiment, total three independent experiments). ** p
    Figure Legend Snippet: RGP-induced activation spleen DCs were dependent on TLR4. C56BL/6 wild-type, TLR2-KO, TLR4-KO, and SR-A-KO mice were treated i.v. with PBS and 50 mg/kg RGP for 24 h. (A) Co-stimulatory molecules and MHC class I and II were measured in the spleen DCs. (B) The serum concentrations of IL-6, IL-12p40, and TNF-α after RGP treatment are shown. (C) The mRNA levels of IFNγ and T-bet in the spleen are shown. All data are the average of analyses of six independent samples (two mice per experiment, total three independent experiments). ** p

    Techniques Used: Activation Assay, Mouse Assay

    RGP-promoted DC maturation in the tumor microenvironment. C57BL/6 mice were injected subcutaneously ( s.c. ) with 1 × 10 6 B16 melanoma cells or B16-OVA cells. Fifteen days after tumor injection, the mice were treated with PBS, 50 mg/kg RGP and 1 mg/kg LPS for 24 h. (A) MFI of CD40, CD80, CD86, and MHC class I and II levels was measured in the spleen and tumor drLN DCs. (B) Concentrations of IL-6, IL-12p40, and TNF-α in serum. (C) Surface OVA peptide (257–264) presentation was measured in the tumor drLN DCs after treatment of PBS, 50 mg/mL RGP, 1 mg/kg LPS with or without 2.5 mg/kg OVA (left panel). Mean positive cells of OVA peptide presenting DCs are shown (right panel). (D) CFSE-labeled OT-I and OT-II T-cell proliferation in B16 tumor-bearing CD45.1 congenic mice were analyzed with flow cytometry. (E) Percentage of IFNγ + and TNF-α + cells in B16-OVA tumor-infiltrated OT-I and OT-II cells. (F) OT-I and OT-II T-cell proliferation in wild-type and TLR4-KO mice. (G) Intracellular IFNγ- and TNF-α-producing OT-I and OT-II cells in B16-OVA tumor in the wild-type and TLR4-KO mice. All data are representative of the average of analyses of six independent samples (three mice per experiment, total two independent experiments). ** p
    Figure Legend Snippet: RGP-promoted DC maturation in the tumor microenvironment. C57BL/6 mice were injected subcutaneously ( s.c. ) with 1 × 10 6 B16 melanoma cells or B16-OVA cells. Fifteen days after tumor injection, the mice were treated with PBS, 50 mg/kg RGP and 1 mg/kg LPS for 24 h. (A) MFI of CD40, CD80, CD86, and MHC class I and II levels was measured in the spleen and tumor drLN DCs. (B) Concentrations of IL-6, IL-12p40, and TNF-α in serum. (C) Surface OVA peptide (257–264) presentation was measured in the tumor drLN DCs after treatment of PBS, 50 mg/mL RGP, 1 mg/kg LPS with or without 2.5 mg/kg OVA (left panel). Mean positive cells of OVA peptide presenting DCs are shown (right panel). (D) CFSE-labeled OT-I and OT-II T-cell proliferation in B16 tumor-bearing CD45.1 congenic mice were analyzed with flow cytometry. (E) Percentage of IFNγ + and TNF-α + cells in B16-OVA tumor-infiltrated OT-I and OT-II cells. (F) OT-I and OT-II T-cell proliferation in wild-type and TLR4-KO mice. (G) Intracellular IFNγ- and TNF-α-producing OT-I and OT-II cells in B16-OVA tumor in the wild-type and TLR4-KO mice. All data are representative of the average of analyses of six independent samples (three mice per experiment, total two independent experiments). ** p

    Techniques Used: Mouse Assay, Injection, Labeling, Flow Cytometry, Cytometry

    RGP-induced activation of BMDCs and spleen DCs. BMDCs were treated with 10, 50, and 100 μg/mL RGP and with 2 μg/mL LPS for 24 h. (A) Morphological changes of BMDCs. (B) Mean fluorescence intensity levels of CD40, CD80, CD86, MHC class I, and MHC class II in the BMDCs. (C) C57BL/6 mice were injected intravenously ( i.v. ) with 12.5, 25, and 50 mg/kg RGP and with 1 mg/kg LPS. The flow cytometric analyses of co-stimulatory molecules and MHC class I and II in gated lineage-CD11c+ cells from the spleen are shown. (D) Concentrations of IL-6, IL-12p40, and TNF-α in BMDC-cultured medium. (E) The serum concentration of IL-6, IL-12p40, and TNF-α in RGP-treated mouse is shown. (F) C57BL/6 mice were injected i.v. with 50 mg/kg RGP. Three days later, the mice were injected again with the same amount of RGP, and then 3 d later, intracellular IFNγ, IL-4, and IL-17 in CD4 + and CD8 + T cells in the spleen were analyzed with flow cytometry. All data are representative of the average of analyses of six independent samples (two mice per experiment, total three independent experiments). * p
    Figure Legend Snippet: RGP-induced activation of BMDCs and spleen DCs. BMDCs were treated with 10, 50, and 100 μg/mL RGP and with 2 μg/mL LPS for 24 h. (A) Morphological changes of BMDCs. (B) Mean fluorescence intensity levels of CD40, CD80, CD86, MHC class I, and MHC class II in the BMDCs. (C) C57BL/6 mice were injected intravenously ( i.v. ) with 12.5, 25, and 50 mg/kg RGP and with 1 mg/kg LPS. The flow cytometric analyses of co-stimulatory molecules and MHC class I and II in gated lineage-CD11c+ cells from the spleen are shown. (D) Concentrations of IL-6, IL-12p40, and TNF-α in BMDC-cultured medium. (E) The serum concentration of IL-6, IL-12p40, and TNF-α in RGP-treated mouse is shown. (F) C57BL/6 mice were injected i.v. with 50 mg/kg RGP. Three days later, the mice were injected again with the same amount of RGP, and then 3 d later, intracellular IFNγ, IL-4, and IL-17 in CD4 + and CD8 + T cells in the spleen were analyzed with flow cytometry. All data are representative of the average of analyses of six independent samples (two mice per experiment, total three independent experiments). * p

    Techniques Used: Activation Assay, Fluorescence, Mouse Assay, Injection, Flow Cytometry, Cell Culture, Concentration Assay, Cytometry

    14) Product Images from "Access of protective antiviral antibody to neuronal tissues requires CD4 T cell help"

    Article Title: Access of protective antiviral antibody to neuronal tissues requires CD4 T cell help

    Journal: Nature

    doi: 10.1038/nature17979

    In the absence of T RM , B cells are required for protection of the host against genital HSV-2 challenge (a) C57/BL6 mice and µMT mice were immunized ivag or i.n. with TK − HSV-2. Five weeks later, vaginal tissue sections were stained for CD4 + cells (red) and MHC class II + cells (green). Blue labeling depicts nuclear staining with DAPI (blue). Images were captured using a 10× or 40× objective lens. Scale bars indicate 100 µm. These data are representative of three similar experiments. (b–d) Balb/c mice and J H D mice were immunized with TK − HSV-2 (10 5 pfu) via i.n. or ivag route. Six weeks later, the number of total CD4 + T cells and HSV-2-specific IFN-γ + CD4 + T cells in the vagina (b), spleen (c), and peripheral blood (d) were analyzed by flow cytometry. Percentages and total number of IFN-γ + cells among CD4 + CD90.2 + cells are shown. Data are means ± s.e.m. *, p
    Figure Legend Snippet: In the absence of T RM , B cells are required for protection of the host against genital HSV-2 challenge (a) C57/BL6 mice and µMT mice were immunized ivag or i.n. with TK − HSV-2. Five weeks later, vaginal tissue sections were stained for CD4 + cells (red) and MHC class II + cells (green). Blue labeling depicts nuclear staining with DAPI (blue). Images were captured using a 10× or 40× objective lens. Scale bars indicate 100 µm. These data are representative of three similar experiments. (b–d) Balb/c mice and J H D mice were immunized with TK − HSV-2 (10 5 pfu) via i.n. or ivag route. Six weeks later, the number of total CD4 + T cells and HSV-2-specific IFN-γ + CD4 + T cells in the vagina (b), spleen (c), and peripheral blood (d) were analyzed by flow cytometry. Percentages and total number of IFN-γ + cells among CD4 + CD90.2 + cells are shown. Data are means ± s.e.m. *, p

    Techniques Used: Mouse Assay, Staining, Labeling, Flow Cytometry, Cytometry

    Majority of CD4 T cells recruited to the DRG and spinal cord of immunized mice is localized in the parenchyma of neuronal tissues (a) C57/BL6 mice were immunized i.n. with TK − HSV-2. Six days after challenge of immunized mice six weeks prior, neuronal tissue sections (DRG and spinal cord) were stained for CD4 + cells and VCAM-1 + cells or CD31 + cells (red or green). Blue labeling depicts nuclear staining with DAPI (blue). Images were captured using a 10× or 40× objective lens. Scale bars indicate 100 µm. (b) C57/BL6 mice were immunized i.n. with TK − HSV-2. Six weeks later, mice were challenged with WT HSV-2 ivag and neuronal tissues were collected 6 days later. DRG and spinal cord were stained for CD4 + cells (red) and MHC class II + cells, CD11b + cells or Ly6G + cells (green). Blue labeling depicts nuclear staining with DAPI (blue). Images were captured using a 10× or 40× objective lens. Scale bars indicate 100 µm. These data are representative of at least three similar experiments.
    Figure Legend Snippet: Majority of CD4 T cells recruited to the DRG and spinal cord of immunized mice is localized in the parenchyma of neuronal tissues (a) C57/BL6 mice were immunized i.n. with TK − HSV-2. Six days after challenge of immunized mice six weeks prior, neuronal tissue sections (DRG and spinal cord) were stained for CD4 + cells and VCAM-1 + cells or CD31 + cells (red or green). Blue labeling depicts nuclear staining with DAPI (blue). Images were captured using a 10× or 40× objective lens. Scale bars indicate 100 µm. (b) C57/BL6 mice were immunized i.n. with TK − HSV-2. Six weeks later, mice were challenged with WT HSV-2 ivag and neuronal tissues were collected 6 days later. DRG and spinal cord were stained for CD4 + cells (red) and MHC class II + cells, CD11b + cells or Ly6G + cells (green). Blue labeling depicts nuclear staining with DAPI (blue). Images were captured using a 10× or 40× objective lens. Scale bars indicate 100 µm. These data are representative of at least three similar experiments.

    Techniques Used: Mouse Assay, Staining, Labeling

    15) Product Images from "Age-associated alterations in CD8?+ dendritic cells impair CD8 T-cell expansion in response to an intracellular bacterium"

    Article Title: Age-associated alterations in CD8?+ dendritic cells impair CD8 T-cell expansion in response to an intracellular bacterium

    Journal: Aging Cell

    doi: 10.1111/j.1474-9726.2012.00867.x

    CD8α+ DC exhibit maturation defects in vivo following infection, yet function normally in vitro . (A–D) DCs were isolated from spleens of naïve and Lm-OVA-infected adult (closed circles) and old mice (open squares) at various time points, and the expression levels of (A) MHC-II, (B) CD40, and (C) CD86 were determined. Representative histograms of expression by adult (gray) and old (open-white) CD8α+ DC are shown. (D–F) CD11c+ DC from naïve adult (gray) or old (open/white histograms) spleens were purified and stimulated in vitro overnight with either Lm-OVA or LPS, and the upregulation within the CD8α+ DC subset of (D) MHC-II, (E) CD40, and (F) CD86 was determined 18 h later. (G) Adult (gray histograms) and old (white histograms) CD11c+ DC were stimulated overnight with LPS plus (upper row) 10 −8 m or (lower row) 10 −10 m SIINFEKL peptide, and then co-cultured with CFSE-labeled adult OT-I T cells at various T:DC ratios, as indicated, for 3 days. Data represent two independent experiments with 3 mice group −1 time point −1 . * P
    Figure Legend Snippet: CD8α+ DC exhibit maturation defects in vivo following infection, yet function normally in vitro . (A–D) DCs were isolated from spleens of naïve and Lm-OVA-infected adult (closed circles) and old mice (open squares) at various time points, and the expression levels of (A) MHC-II, (B) CD40, and (C) CD86 were determined. Representative histograms of expression by adult (gray) and old (open-white) CD8α+ DC are shown. (D–F) CD11c+ DC from naïve adult (gray) or old (open/white histograms) spleens were purified and stimulated in vitro overnight with either Lm-OVA or LPS, and the upregulation within the CD8α+ DC subset of (D) MHC-II, (E) CD40, and (F) CD86 was determined 18 h later. (G) Adult (gray histograms) and old (white histograms) CD11c+ DC were stimulated overnight with LPS plus (upper row) 10 −8 m or (lower row) 10 −10 m SIINFEKL peptide, and then co-cultured with CFSE-labeled adult OT-I T cells at various T:DC ratios, as indicated, for 3 days. Data represent two independent experiments with 3 mice group −1 time point −1 . * P

    Techniques Used: In Vivo, Infection, In Vitro, Isolation, Mouse Assay, Expressing, Purification, Cell Culture, Labeling

    16) Product Images from "Identification of Immune and Viral Correlates of Norovirus Protective Immunity through Comparative Study of Intra-Cluster Norovirus Strains"

    Article Title: Identification of Immune and Viral Correlates of Norovirus Protective Immunity through Comparative Study of Intra-Cluster Norovirus Strains

    Journal: PLoS Pathogens

    doi: 10.1371/journal.ppat.1003592

    VP2 regulates antigen presenting cell maturation in vitro. 5.0×10 5 RAW 264.7 cells were plated overnight in 24-well plates and then exposed to mock inoculum or recombinant MNV-1, MNV-3, MNV-1.3 VP2, or MNV-3.1 VP2 at MOI 5 for 24 h. In certain experiments, a well was also co-infected with equivalent titers of parental MNV-1 and MNV-3 such that cells were infected at MOI 5. Cells were stained with antibodies to MHC class I and MHC class II molecules, co-stimulatory molecules CD40 and CD80, or CD103. Flow cytometry was carried out as described in the Methods . Six independent experiments were performed for single infections, and three experiments included co-infections. A ) Representative data from one experiment are presented in the histograms. In these graphs, surface expression of the indicated marker is shown for mock-inoculated cells (light grey histogram filled with dashed lines); cells infected with parental MNV-1 or MNV-3 (black histogram); and cells infected with chimeric MNV-1.3VP2 or MNV-3.1VP2 (dark grey histogram). The parental and chimeric virus pair is shown to the left of the histograms. The dashed vertical lines indicate the mean basal expression levels in mock-inoculated cells. B ) In the bar graphs, the percentage of cells displaying upregulated surface expression of the indicated marker is presented. Data from all experimental replicates are averaged. Cells inoculated with UV-inactivated MNV-1 or MNV-3 displayed statistically similar levels of each marker compared to mock-inoculated cells (data not shown). Statistical comparisons were made between mock and each parental MNV; MNV-1 and MNV-3; each parental and chimeric pair; and each parental MNV and the co-infected cells.
    Figure Legend Snippet: VP2 regulates antigen presenting cell maturation in vitro. 5.0×10 5 RAW 264.7 cells were plated overnight in 24-well plates and then exposed to mock inoculum or recombinant MNV-1, MNV-3, MNV-1.3 VP2, or MNV-3.1 VP2 at MOI 5 for 24 h. In certain experiments, a well was also co-infected with equivalent titers of parental MNV-1 and MNV-3 such that cells were infected at MOI 5. Cells were stained with antibodies to MHC class I and MHC class II molecules, co-stimulatory molecules CD40 and CD80, or CD103. Flow cytometry was carried out as described in the Methods . Six independent experiments were performed for single infections, and three experiments included co-infections. A ) Representative data from one experiment are presented in the histograms. In these graphs, surface expression of the indicated marker is shown for mock-inoculated cells (light grey histogram filled with dashed lines); cells infected with parental MNV-1 or MNV-3 (black histogram); and cells infected with chimeric MNV-1.3VP2 or MNV-3.1VP2 (dark grey histogram). The parental and chimeric virus pair is shown to the left of the histograms. The dashed vertical lines indicate the mean basal expression levels in mock-inoculated cells. B ) In the bar graphs, the percentage of cells displaying upregulated surface expression of the indicated marker is presented. Data from all experimental replicates are averaged. Cells inoculated with UV-inactivated MNV-1 or MNV-3 displayed statistically similar levels of each marker compared to mock-inoculated cells (data not shown). Statistical comparisons were made between mock and each parental MNV; MNV-1 and MNV-3; each parental and chimeric pair; and each parental MNV and the co-infected cells.

    Techniques Used: In Vitro, Recombinant, Infection, Staining, Flow Cytometry, Cytometry, Expressing, Marker

    17) Product Images from "Small intestinal eosinophils regulate Th17 cells by producing IL-1 receptor antagonist"

    Article Title: Small intestinal eosinophils regulate Th17 cells by producing IL-1 receptor antagonist

    Journal: The Journal of Experimental Medicine

    doi: 10.1084/jem.20141388

    Small intestinal Th17 and Th1 cells are increased in eosinophil - deficient mice. (A) Flow cytometry analysis of the frequency of MHC class II neg/low Siglec-F high eosinophils in various tissues from WT BALB/c or eosinophil-deficient (ΔdblGATA-1) mice. (B–D) Flow cytometry analysis of IFN-γ + and IL-17A + (B), CD25 + Foxp3 + (C), and GATA3 + (D) CD4 + T cells in the small intestine (SI), spleen (SP), and MLN from ΔdblGATA-1 or littermate control mice (BALB/c). FACS plots show cells gated on TCRβ + CD4 + cells. Data in B and C are representative of at least 10 independent experiments ( n = 3–5 mice per group per experiment), and data in D are representative of four independent experiments ( n = 3–5 mice per group per experiment). (E and F) Flow cytometry analysis of RORγt + and GATA3 + (E) and IL-22 + (F) innate lymphoid cells in the small intestine of ΔdblGATA-1 or littermate control mice (BALB/c). FACS plots show cells gated on Thy1.2 + lineage − (CD3 − CD4 − CD8α − CD11b − CD11c − CD19 − B220 − TER119 − Gr-1 − NK1.1 − ) cells and are representative of four independent experiments ( n = 2–4 mice per group per experiment). (G–I) Frequency of IFN-γ + and IL-17A + (G), CD25 + Foxp3 + (H), and GATA3 + (I) CD4 + T cells in the small intestine of ΔdblGATA-1 or littermate control mice (C57BL/6). ΔdblGATA-1 mice backcrossed with C57BL/6 for at least 10 generations were used. FACS plots show cells gated on TCRβ + CD4 + cells and are representative of at least three independent experiments ( n = 3–4 mice per group per experiment). (J) Flow cytometry analysis of the frequency of c-kit − FcεR1 + and IgE + CD49b + basophils in peripheral blood and the small intestine. (K–M) Frequency of IFN-γ + and IL-17A + (K), Foxp3 + (L), and GATA3 + (M) CD4 + T cells in the small intestine of C57BL/6 mice administered every second day with Siglec-F–specific antibodies or isotype control (Rat IgG2a) for 2 wk. FACS plots show cells gated on TCRβ + CD4 + cells and are representative of two independent experiments ( n = 4 mice per group per experiment). Bar graphs show the mean ± SEM. *, P
    Figure Legend Snippet: Small intestinal Th17 and Th1 cells are increased in eosinophil - deficient mice. (A) Flow cytometry analysis of the frequency of MHC class II neg/low Siglec-F high eosinophils in various tissues from WT BALB/c or eosinophil-deficient (ΔdblGATA-1) mice. (B–D) Flow cytometry analysis of IFN-γ + and IL-17A + (B), CD25 + Foxp3 + (C), and GATA3 + (D) CD4 + T cells in the small intestine (SI), spleen (SP), and MLN from ΔdblGATA-1 or littermate control mice (BALB/c). FACS plots show cells gated on TCRβ + CD4 + cells. Data in B and C are representative of at least 10 independent experiments ( n = 3–5 mice per group per experiment), and data in D are representative of four independent experiments ( n = 3–5 mice per group per experiment). (E and F) Flow cytometry analysis of RORγt + and GATA3 + (E) and IL-22 + (F) innate lymphoid cells in the small intestine of ΔdblGATA-1 or littermate control mice (BALB/c). FACS plots show cells gated on Thy1.2 + lineage − (CD3 − CD4 − CD8α − CD11b − CD11c − CD19 − B220 − TER119 − Gr-1 − NK1.1 − ) cells and are representative of four independent experiments ( n = 2–4 mice per group per experiment). (G–I) Frequency of IFN-γ + and IL-17A + (G), CD25 + Foxp3 + (H), and GATA3 + (I) CD4 + T cells in the small intestine of ΔdblGATA-1 or littermate control mice (C57BL/6). ΔdblGATA-1 mice backcrossed with C57BL/6 for at least 10 generations were used. FACS plots show cells gated on TCRβ + CD4 + cells and are representative of at least three independent experiments ( n = 3–4 mice per group per experiment). (J) Flow cytometry analysis of the frequency of c-kit − FcεR1 + and IgE + CD49b + basophils in peripheral blood and the small intestine. (K–M) Frequency of IFN-γ + and IL-17A + (K), Foxp3 + (L), and GATA3 + (M) CD4 + T cells in the small intestine of C57BL/6 mice administered every second day with Siglec-F–specific antibodies or isotype control (Rat IgG2a) for 2 wk. FACS plots show cells gated on TCRβ + CD4 + cells and are representative of two independent experiments ( n = 4 mice per group per experiment). Bar graphs show the mean ± SEM. *, P

    Techniques Used: Mouse Assay, Flow Cytometry, Cytometry, FACS

    18) Product Images from "Dendritic Cell Editing by Activated Natural Killer Cells Results in a More Protective Cancer-Specific Immune Response"

    Article Title: Dendritic Cell Editing by Activated Natural Killer Cells Results in a More Protective Cancer-Specific Immune Response

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0039170

    Subcutaneous administration of MHC-negative cells results in an NK cell-dependent decrease of CD11c bright DCs in the draining lymph nodes. A: representative analyses of mononuclear cells isolated from either draining or controlateral (Control LN) lymph nodes of mice depleted or not of NK cells (Draining LN + anti-asialo GM1). B: NK cells are efficiently depleted in mice upon administration of anti-asialo GM1 mAbs. C: the percentage (left) and the absolute number (right) of DCs among mononuclear cells isolated from lymph nodes. Bars represents mean values and SD of five independent experiments (three mice per group). ** = p
    Figure Legend Snippet: Subcutaneous administration of MHC-negative cells results in an NK cell-dependent decrease of CD11c bright DCs in the draining lymph nodes. A: representative analyses of mononuclear cells isolated from either draining or controlateral (Control LN) lymph nodes of mice depleted or not of NK cells (Draining LN + anti-asialo GM1). B: NK cells are efficiently depleted in mice upon administration of anti-asialo GM1 mAbs. C: the percentage (left) and the absolute number (right) of DCs among mononuclear cells isolated from lymph nodes. Bars represents mean values and SD of five independent experiments (three mice per group). ** = p

    Techniques Used: Isolation, Mouse Assay

    19) Product Images from "Naive tumor-specific CD4+ T cells differentiated in vivo eradicate established melanoma"

    Article Title: Naive tumor-specific CD4+ T cells differentiated in vivo eradicate established melanoma

    Journal: The Journal of Experimental Medicine

    doi: 10.1084/jem.20091921

    γ c signaling on host DC is required for survival and differentiation of TRP-1 T cells in vivo. (A) Differential activation of TRP-1 CD4 + T cells in RAG −/− and RAG −/− γ c −/− hosts. Spleens from RAG −/− or RAG −/− γ c −/− mice were isolated and stained for TRP-1 CD4 + T cells. Shown are CD62L, CD122, ICOS, and CD25 expression on gated TRP-1 CD4 + T cells from indicated host. (B) IFN-γ and CXCL9 are differentially expressed in the serum at 1 wk in RAG −/− and RAG −/− γ c −/− hosts after TRP-1 CD4 + T cell transfer. Horizontal bars represent mean. (C) TRP-1 CD4 + T cells expand in RAG −/− γ c −/− hosts initially but fail to survive after 4 wk. Error bars indicate SEM. (D) Flow cytometry of IFN-γ and IL-17 expression in TRP-1 CD4 + T cells isolated from tumor bearing RAG −/− and RAG −/− γ c −/− mice 4 wk after transfer. T cells were activated with PMA and ionomycin for 4 h and then fixed and permeabilized and stained with anti–IFN-γ and IL-17 antibodies. (E) Tbet expression in TRP-1 CD4 + T cells 4 wk after transfer. (F) MHC class II, CD80, and CD40 expression in RAG −/− γ c −/− hosts. Top flow diagram indicates CD11c high MHC class II + cells; bottom flow histograms show CD80 and CD40 expression on gated CD11c high MHC class II + cells. Solid line represents RAG −/− mice treated with TRP-1 CD4 + T cells. Shaded histogram represents RAG −/− γ c −/− mice treated with TRP-1 CD4 + T cells. Data are representative of three independent experiments ( n = 5 mice/group).
    Figure Legend Snippet: γ c signaling on host DC is required for survival and differentiation of TRP-1 T cells in vivo. (A) Differential activation of TRP-1 CD4 + T cells in RAG −/− and RAG −/− γ c −/− hosts. Spleens from RAG −/− or RAG −/− γ c −/− mice were isolated and stained for TRP-1 CD4 + T cells. Shown are CD62L, CD122, ICOS, and CD25 expression on gated TRP-1 CD4 + T cells from indicated host. (B) IFN-γ and CXCL9 are differentially expressed in the serum at 1 wk in RAG −/− and RAG −/− γ c −/− hosts after TRP-1 CD4 + T cell transfer. Horizontal bars represent mean. (C) TRP-1 CD4 + T cells expand in RAG −/− γ c −/− hosts initially but fail to survive after 4 wk. Error bars indicate SEM. (D) Flow cytometry of IFN-γ and IL-17 expression in TRP-1 CD4 + T cells isolated from tumor bearing RAG −/− and RAG −/− γ c −/− mice 4 wk after transfer. T cells were activated with PMA and ionomycin for 4 h and then fixed and permeabilized and stained with anti–IFN-γ and IL-17 antibodies. (E) Tbet expression in TRP-1 CD4 + T cells 4 wk after transfer. (F) MHC class II, CD80, and CD40 expression in RAG −/− γ c −/− hosts. Top flow diagram indicates CD11c high MHC class II + cells; bottom flow histograms show CD80 and CD40 expression on gated CD11c high MHC class II + cells. Solid line represents RAG −/− mice treated with TRP-1 CD4 + T cells. Shaded histogram represents RAG −/− γ c −/− mice treated with TRP-1 CD4 + T cells. Data are representative of three independent experiments ( n = 5 mice/group).

    Techniques Used: In Vivo, Activation Assay, Mouse Assay, Isolation, Staining, Expressing, Flow Cytometry, Cytometry

    20) Product Images from "Laminarin promotes anti-cancer immunity by the maturation of dendritic cells"

    Article Title: Laminarin promotes anti-cancer immunity by the maturation of dendritic cells

    Journal: Oncotarget

    doi: 10.18632/oncotarget.16170

    Laminarin-promoted maturation of drLNs and DCs in the tumor microenvironment C57BL/6 mice were inoculated s.c . with 1 × 10 6 B16 melanoma cells or B16-OVA cells. Fifteen days after tumor injection, the mice were treated with PBS and 25 mg/kg of laminarin for 24 hours. (A) MFI of CD40, CD80, CD86, and MHC classes I and II levels were measured in the spleen and tumor drLNs and DCs. (B) Concentrations of IL-6, IL-12p40, and TNF-α in the mice sera. (C) Surface OVA peptide (257-264) presentation was measured in the tumor drLNs and DCs. (D) CFSE-labeled OT-I and OT-II T cell proliferation in B16-OVA tumor-bearing CD45.1 congenic mice were analyzed using flow cytometry. (E) The means of the absolute numbers of OT-I (left panel) and OT-II (right panel) cells in the tumor. (F) Percentage of IFN-γ + and TNF-α + cells in tumor-infiltrated OT-I and OT-II cells. All data are representative of or the average of analyses of six independent samples (two mice per experiment, for a total of three independent experiments). ** p
    Figure Legend Snippet: Laminarin-promoted maturation of drLNs and DCs in the tumor microenvironment C57BL/6 mice were inoculated s.c . with 1 × 10 6 B16 melanoma cells or B16-OVA cells. Fifteen days after tumor injection, the mice were treated with PBS and 25 mg/kg of laminarin for 24 hours. (A) MFI of CD40, CD80, CD86, and MHC classes I and II levels were measured in the spleen and tumor drLNs and DCs. (B) Concentrations of IL-6, IL-12p40, and TNF-α in the mice sera. (C) Surface OVA peptide (257-264) presentation was measured in the tumor drLNs and DCs. (D) CFSE-labeled OT-I and OT-II T cell proliferation in B16-OVA tumor-bearing CD45.1 congenic mice were analyzed using flow cytometry. (E) The means of the absolute numbers of OT-I (left panel) and OT-II (right panel) cells in the tumor. (F) Percentage of IFN-γ + and TNF-α + cells in tumor-infiltrated OT-I and OT-II cells. All data are representative of or the average of analyses of six independent samples (two mice per experiment, for a total of three independent experiments). ** p

    Techniques Used: Mouse Assay, Injection, Labeling, Flow Cytometry, Cytometry

    Laminarin-induced activation of spleen DCs in vivo C57BL/6 mice were injected intravenously ( i.v .) with 12.5, 25, and 50 mg/kg of laminarin and 10 mg/kg of fucoidan for 24 hours before the spleens were harvested. (A) Definition of spleen DCs. Lineage markers included CD3, Thy1.1, B220, Gr-1, CD49b, and TER-119. (B) Percentages of lineage − CD11c + DCs in the spleens. (C) Mean of the absolute numbers of lineage − CD11c + spleen DCs within live cells. (D) Mean fluorescence intensity (MFI) of co-stimulatory molecules and MHC classes I and II in gated lineage − CD11c + cells from the spleens were analyzed using flow cytometry. (E) mRNA levels. (F) Sera concentration of IL-6, IL-12p70, and TNF-α. (G) Intracellular IL-6, IL-12, and TNF-α production in spleen DCs. All data are representative of or the average of analyses of six independent samples (two mice per experiment, totaling three independent experiments). * p
    Figure Legend Snippet: Laminarin-induced activation of spleen DCs in vivo C57BL/6 mice were injected intravenously ( i.v .) with 12.5, 25, and 50 mg/kg of laminarin and 10 mg/kg of fucoidan for 24 hours before the spleens were harvested. (A) Definition of spleen DCs. Lineage markers included CD3, Thy1.1, B220, Gr-1, CD49b, and TER-119. (B) Percentages of lineage − CD11c + DCs in the spleens. (C) Mean of the absolute numbers of lineage − CD11c + spleen DCs within live cells. (D) Mean fluorescence intensity (MFI) of co-stimulatory molecules and MHC classes I and II in gated lineage − CD11c + cells from the spleens were analyzed using flow cytometry. (E) mRNA levels. (F) Sera concentration of IL-6, IL-12p70, and TNF-α. (G) Intracellular IL-6, IL-12, and TNF-α production in spleen DCs. All data are representative of or the average of analyses of six independent samples (two mice per experiment, totaling three independent experiments). * p

    Techniques Used: Activation Assay, In Vivo, Mouse Assay, Injection, Fluorescence, Flow Cytometry, Cytometry, Concentration Assay

    21) Product Images from "Ginseng Berry Extract Promotes Maturation of Mouse Dendritic Cells"

    Article Title: Ginseng Berry Extract Promotes Maturation of Mouse Dendritic Cells

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0130926

    In vivo administration of GB induces spleen DC activation. C57BL/6 mice were injected i . p . with 10 or 50 mg/kg GB for 24 hours. (A) Percentage of lineage - CD11c + spleen DCs was analyzed on a flow cytometry (left panels). Absolute cell number of lineage - CD11c + cells within live cells were shown (right panel). (B) Expression levels of CD40, CD80, CD86, MHC class I and MHC class II were measured by flow cytometry. Data are representative of analyses of 8 independent samples. (C) C57BL/6 mice were treated orally, i . p . or i . v . with 50 mg/kg GB for 24 hours and measured Expression levels of CD40, CD80, CD86, MHC class I and MHC class II by flow cytometry. (D) Expression levels of IL-6, IL-12p40 and TNF-α mRNA were measured from spleen 2 hours after 50 mg/kg GB injection. (E) IL-6, IL-12p70 and TNF-α levels in sera were shown from spleen 24 hours after 50 mg/kg GB injection. (F) Intracellular cytokine production levels were measured spleen DCs. All data are from analyses of 6 individual mice each group (2 mice per experiment, total 3 independent experiments). * p
    Figure Legend Snippet: In vivo administration of GB induces spleen DC activation. C57BL/6 mice were injected i . p . with 10 or 50 mg/kg GB for 24 hours. (A) Percentage of lineage - CD11c + spleen DCs was analyzed on a flow cytometry (left panels). Absolute cell number of lineage - CD11c + cells within live cells were shown (right panel). (B) Expression levels of CD40, CD80, CD86, MHC class I and MHC class II were measured by flow cytometry. Data are representative of analyses of 8 independent samples. (C) C57BL/6 mice were treated orally, i . p . or i . v . with 50 mg/kg GB for 24 hours and measured Expression levels of CD40, CD80, CD86, MHC class I and MHC class II by flow cytometry. (D) Expression levels of IL-6, IL-12p40 and TNF-α mRNA were measured from spleen 2 hours after 50 mg/kg GB injection. (E) IL-6, IL-12p70 and TNF-α levels in sera were shown from spleen 24 hours after 50 mg/kg GB injection. (F) Intracellular cytokine production levels were measured spleen DCs. All data are from analyses of 6 individual mice each group (2 mice per experiment, total 3 independent experiments). * p

    Techniques Used: In Vivo, Activation Assay, Mouse Assay, Injection, Flow Cytometry, Cytometry, Expressing

    TLR4 and MyD88 singling pathway is essential for GB-induced DC activation. C57BL/6, TLR2 -/- , TLR4 -/- or MyD88 -/- mice were injected i . p . with PBS or 50 mg/kg GB for 24 hours. (A) Percentage of lineage - CD11c + spleen DCs was analyzed on a flow cytometry (left panels). Absolute cell number of spleen DCs within live cells were shown (right panel). (B) Expression level of CD86, MHC class I and MHC class II were measured by flow cytometry. (C) IL-6, IL-12p70 and TNF-α levels in sera were shown. All data are representative or the average of analyses of 6 samples from 3 independent experiments. * p
    Figure Legend Snippet: TLR4 and MyD88 singling pathway is essential for GB-induced DC activation. C57BL/6, TLR2 -/- , TLR4 -/- or MyD88 -/- mice were injected i . p . with PBS or 50 mg/kg GB for 24 hours. (A) Percentage of lineage - CD11c + spleen DCs was analyzed on a flow cytometry (left panels). Absolute cell number of spleen DCs within live cells were shown (right panel). (B) Expression level of CD86, MHC class I and MHC class II were measured by flow cytometry. (C) IL-6, IL-12p70 and TNF-α levels in sera were shown. All data are representative or the average of analyses of 6 samples from 3 independent experiments. * p

    Techniques Used: Activation Assay, Mouse Assay, Injection, Flow Cytometry, Cytometry, Expressing

    22) Product Images from "Thy1+IL-7+ lymphatic endothelial cells in iBALT provide a survival niche for memory T-helper cells in allergic airway inflammation"

    Article Title: Thy1+IL-7+ lymphatic endothelial cells in iBALT provide a survival niche for memory T-helper cells in allergic airway inflammation

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    doi: 10.1073/pnas.1512600113

    Distribution and phenotypic characterization of antigen-specific memory Th2 cells after i.n. antigen administration. ( A ) Effector Th2 cells from DO11.10 OVA-specific αβTCR Tg mice were transferred i.v. into BALB/c mice, which were subsequently challenged i.n. with OVA or PBS solution (control) or i.p. with OVA (control) at days 1 and 3. Indicated examinations were performed at day 42. ( B ) Typical staining patterns of CD44 and KJ1 among CD4 + cells in the indicated organs are shown. ( C ) Kinetics of the ratio of KJ1 + cells among CD4 + cells in the indicated organs are shown with SD. Three independent experiments were performed with similar results ( B and C ). ( D ) Representative cell surface staining profiles on KJ1 + CD4 + memory Th2 cells (red lines) and KJ1 − CD4 + recipient CD4 T cells (black lines) at day 42; gray shaded curves indicate staining with isotype-matched control antibody. Two independent experiments were performed with similar results. ( E ) For cytokine production profiles of memory Th2 cells, CD4 + cells recovered from spleen and lung were stimulated in vitro with immobilized anti-TCRβ for 6 h. Intracellular staining profiles of IL-4, IL-5, and IL-13 are shown gated on KJ1 + memory Th2 cells. ( F ) For detection of DNA synthesis, mice were injected i.p. with BrdU during days 42–47. Cells from the indicated organs were harvested at day 43 (24 h) or day 48 (6 d). Representative staining profiles of KJ1 and BrdU among CD4 + cells are shown. ( G ) For in vivo proliferation, cells from the indicated organs were harvested at day 42 and analyzed by intracellular staining of Ki67. Representative staining profiles of Ki67 gated on KJ1 + CD4 + cells in the indicated organs are shown. Two independent experiments were performed with similar results ( F and G ). ( H ) At day 42, frozen sections of lungs obtained from mice transferred with Th2 cells stained with anti-KJ1 (blue), anti-MHC class II (red), and anti-PNAd (green) are shown. Two independent experiments were performed with similar results. ( I ) At day 42, frozen sections of lungs obtained from mice transferred with the indicated type of cells stained for anti-KJ1 (blue), anti-CD3ε (red), and anti-B220 (green) are shown. (Scale bars, 20 μm.) Three independent experiments were performed with similar results.
    Figure Legend Snippet: Distribution and phenotypic characterization of antigen-specific memory Th2 cells after i.n. antigen administration. ( A ) Effector Th2 cells from DO11.10 OVA-specific αβTCR Tg mice were transferred i.v. into BALB/c mice, which were subsequently challenged i.n. with OVA or PBS solution (control) or i.p. with OVA (control) at days 1 and 3. Indicated examinations were performed at day 42. ( B ) Typical staining patterns of CD44 and KJ1 among CD4 + cells in the indicated organs are shown. ( C ) Kinetics of the ratio of KJ1 + cells among CD4 + cells in the indicated organs are shown with SD. Three independent experiments were performed with similar results ( B and C ). ( D ) Representative cell surface staining profiles on KJ1 + CD4 + memory Th2 cells (red lines) and KJ1 − CD4 + recipient CD4 T cells (black lines) at day 42; gray shaded curves indicate staining with isotype-matched control antibody. Two independent experiments were performed with similar results. ( E ) For cytokine production profiles of memory Th2 cells, CD4 + cells recovered from spleen and lung were stimulated in vitro with immobilized anti-TCRβ for 6 h. Intracellular staining profiles of IL-4, IL-5, and IL-13 are shown gated on KJ1 + memory Th2 cells. ( F ) For detection of DNA synthesis, mice were injected i.p. with BrdU during days 42–47. Cells from the indicated organs were harvested at day 43 (24 h) or day 48 (6 d). Representative staining profiles of KJ1 and BrdU among CD4 + cells are shown. ( G ) For in vivo proliferation, cells from the indicated organs were harvested at day 42 and analyzed by intracellular staining of Ki67. Representative staining profiles of Ki67 gated on KJ1 + CD4 + cells in the indicated organs are shown. Two independent experiments were performed with similar results ( F and G ). ( H ) At day 42, frozen sections of lungs obtained from mice transferred with Th2 cells stained with anti-KJ1 (blue), anti-MHC class II (red), and anti-PNAd (green) are shown. Two independent experiments were performed with similar results. ( I ) At day 42, frozen sections of lungs obtained from mice transferred with the indicated type of cells stained for anti-KJ1 (blue), anti-CD3ε (red), and anti-B220 (green) are shown. (Scale bars, 20 μm.) Three independent experiments were performed with similar results.

    Techniques Used: Mouse Assay, Staining, In Vitro, DNA Synthesis, Injection, In Vivo

    Memory Th2 cells preferentially localized in iBALT. Effector Th2 cells from DO11.10 OVA-specific αβTCR Tg mice were transferred i.v. into BALB/c mice, which were subsequently challenged i.n. with OVA or PBS solution (control) or i.p. with OVA (control) at days 1 and 3, and the indicated assays were performed at day 42. ( A ) Representative cryosections of the lungs stained with H E are depicted ( Left ), as is the absolute number of iBALT structures in the lungs ( Right ). (Scale bars, 1 mm.) Mean values with SD from at least three mice per group are shown. Two independent experiments were performed with similar results. ( B ) Representative staining profiles of CD44 and KJ1 expression on CD4 + cells from the indicated organs ( Left ) and absolute cell numbers of KJ1 + CD44 hi CD4 + T cells in the indicated organs ( Right ). Mean values with SD from five mice per group are shown. More than five independent experiments were performed with similar results. ( C ) Representative confocal micrograph of lung tissue stained with anti-KJ1 (blue), anti-MHC class II (red), and anti-CD4 (green) ( Upper Left ); anti-KJ1 (blue), anti-MHC class II (red), and anti-B220 (green) ( Upper Middle ); anti-KJ1 (blue), anti-CD3ε (red), and anti-B220 (green) ( Upper Right ); anti-KJ1 (blue), anti-MHC class II (red), and anti-CD11c (green) ( Lower Left ); anti-KJ1 (blue), anti-MHC class II (Red), and anti-VCAM1 (green) ( Lower Middle ); or anti-KJ1 (blue), anti-MHC class II (Red), and anti-CD21 (green) ( Lower Right ) are shown. (Scale bars, 40 μm.) More than three independent experiments were performed with similar results. ( D ) Morphometric analysis of KJ1 + cells localized in lymphoid areas of the lungs is shown. Means and SD calculated from analysis of three slides per mouse from four mice. Two independent experiments were performed with similar results (** P
    Figure Legend Snippet: Memory Th2 cells preferentially localized in iBALT. Effector Th2 cells from DO11.10 OVA-specific αβTCR Tg mice were transferred i.v. into BALB/c mice, which were subsequently challenged i.n. with OVA or PBS solution (control) or i.p. with OVA (control) at days 1 and 3, and the indicated assays were performed at day 42. ( A ) Representative cryosections of the lungs stained with H E are depicted ( Left ), as is the absolute number of iBALT structures in the lungs ( Right ). (Scale bars, 1 mm.) Mean values with SD from at least three mice per group are shown. Two independent experiments were performed with similar results. ( B ) Representative staining profiles of CD44 and KJ1 expression on CD4 + cells from the indicated organs ( Left ) and absolute cell numbers of KJ1 + CD44 hi CD4 + T cells in the indicated organs ( Right ). Mean values with SD from five mice per group are shown. More than five independent experiments were performed with similar results. ( C ) Representative confocal micrograph of lung tissue stained with anti-KJ1 (blue), anti-MHC class II (red), and anti-CD4 (green) ( Upper Left ); anti-KJ1 (blue), anti-MHC class II (red), and anti-B220 (green) ( Upper Middle ); anti-KJ1 (blue), anti-CD3ε (red), and anti-B220 (green) ( Upper Right ); anti-KJ1 (blue), anti-MHC class II (red), and anti-CD11c (green) ( Lower Left ); anti-KJ1 (blue), anti-MHC class II (Red), and anti-VCAM1 (green) ( Lower Middle ); or anti-KJ1 (blue), anti-MHC class II (Red), and anti-CD21 (green) ( Lower Right ) are shown. (Scale bars, 40 μm.) More than three independent experiments were performed with similar results. ( D ) Morphometric analysis of KJ1 + cells localized in lymphoid areas of the lungs is shown. Means and SD calculated from analysis of three slides per mouse from four mice. Two independent experiments were performed with similar results (** P

    Techniques Used: Mouse Assay, Staining, Expressing

    IL-7 controls persistence of iBALT-residing memory Th2 cells. ( A ) Effector Th2 cells from Ly5.1 × OT-II Tg mice were transferred into IL-7:GFP knock-in mice ( Il-7 gfp/+ ). Mice were challenged i.n. with OVA on days 1 and 3 and assessed on day 42. Representative confocal micrograph of lung tissue stained with anti-GFP:IL-7 (green), anti-MHC class II (blue), and anti-Ly5.1 (red) are shown. (Scale bars: Left , 100 μm; Right , 20 μm.) ( B ) Quantification analysis of memory Th2 cells (Ly5.1 + ) proximal to IL-7 + , CD11c + , or B220 + cells in the host lung is shown. ( C ) Representative FACS plots of Annexin V and PI staining ( Upper ) and Bcl2 expression (open histogram) ( Lower ) by KJ1 + CD4 + cells are shown. Percentages of Annexin V + PI + cells ( Upper Right ) and Bcl2 hi cells ( Lower Right ) in the KJ1 + CD4 + population are shown. ( D ) IL-7Rα–specific antibody or isotype control antibody was administered i.n. every second day, starting from day 42 after Th2 cell transfer, and mice were assessed at day 49. Representative confocal micrograph of iBALT in the lung tissue from mice treated with anti–IL-7Rα antibody and stained with anti-B220 (red), anti-CD3ε (blue), and anti-KJ1 (green) are shown ( Left ). Quantification analysis of KJ1 + cells localized in lymphoid areas of the lungs is shown ( Right ). (Scale bars, 100 μm.) ( E – H ) Effector Th2 cells from Ly5.2 Il-7Rα fl/fl × OT-II Tg ( Il-7Rα fl/fl ) mice or Ly5.2 Il-7Rα fl/fl × Cre-ERT Tg × OT-II Tg ( Il-7Rα fl/fl Cre-ERT Tg) mice were transferred into Ly5.1 mice. Mice were challenged i.n. with OVA on days 1 and 3 and assessed on day 42. Tamoxifen was injected i.p. on days 42–46 and 49–53, and mice were analyzed on day 56 ( E and F ) or challenged i.n. with OVA on days 56 and 58, and BAL fluid and airway hyperresponsiveness were assessed on day 59 ( G and H ). ( E ) Representative confocal micrographs of lung tissue stained with anti-B220 (blue), anti-CD3ε (green), and anti-Ly5.2 (red) are shown. (Scale bars, 100 μm.) ( F ) Representative staining profiles of Ly5.2 and CD44 expression on CD4 + cells are shown ( Left ) with absolute cell numbers of memory Th2 cells (Ly5.2 + ) in the lungs ( Right ). ( G ) Absolute cell numbers of eosinophils (Eos.), neutrophils (Neut.), lymphocytes (Lymph.), and macrophages (Mac.) in the BAL fluid are shown. ( H ) Airway hyperresponsiveness was assessed by measuring methacholine-induced changes in lung resistance. The mean values from five ( A , B , E , and F ), four ( G and H ), or three ( D ) mice per group or technical triplicates ( C ) are shown with SD or SEM. More than two independent experiments were performed with similar results ( A – F ; ** P
    Figure Legend Snippet: IL-7 controls persistence of iBALT-residing memory Th2 cells. ( A ) Effector Th2 cells from Ly5.1 × OT-II Tg mice were transferred into IL-7:GFP knock-in mice ( Il-7 gfp/+ ). Mice were challenged i.n. with OVA on days 1 and 3 and assessed on day 42. Representative confocal micrograph of lung tissue stained with anti-GFP:IL-7 (green), anti-MHC class II (blue), and anti-Ly5.1 (red) are shown. (Scale bars: Left , 100 μm; Right , 20 μm.) ( B ) Quantification analysis of memory Th2 cells (Ly5.1 + ) proximal to IL-7 + , CD11c + , or B220 + cells in the host lung is shown. ( C ) Representative FACS plots of Annexin V and PI staining ( Upper ) and Bcl2 expression (open histogram) ( Lower ) by KJ1 + CD4 + cells are shown. Percentages of Annexin V + PI + cells ( Upper Right ) and Bcl2 hi cells ( Lower Right ) in the KJ1 + CD4 + population are shown. ( D ) IL-7Rα–specific antibody or isotype control antibody was administered i.n. every second day, starting from day 42 after Th2 cell transfer, and mice were assessed at day 49. Representative confocal micrograph of iBALT in the lung tissue from mice treated with anti–IL-7Rα antibody and stained with anti-B220 (red), anti-CD3ε (blue), and anti-KJ1 (green) are shown ( Left ). Quantification analysis of KJ1 + cells localized in lymphoid areas of the lungs is shown ( Right ). (Scale bars, 100 μm.) ( E – H ) Effector Th2 cells from Ly5.2 Il-7Rα fl/fl × OT-II Tg ( Il-7Rα fl/fl ) mice or Ly5.2 Il-7Rα fl/fl × Cre-ERT Tg × OT-II Tg ( Il-7Rα fl/fl Cre-ERT Tg) mice were transferred into Ly5.1 mice. Mice were challenged i.n. with OVA on days 1 and 3 and assessed on day 42. Tamoxifen was injected i.p. on days 42–46 and 49–53, and mice were analyzed on day 56 ( E and F ) or challenged i.n. with OVA on days 56 and 58, and BAL fluid and airway hyperresponsiveness were assessed on day 59 ( G and H ). ( E ) Representative confocal micrographs of lung tissue stained with anti-B220 (blue), anti-CD3ε (green), and anti-Ly5.2 (red) are shown. (Scale bars, 100 μm.) ( F ) Representative staining profiles of Ly5.2 and CD44 expression on CD4 + cells are shown ( Left ) with absolute cell numbers of memory Th2 cells (Ly5.2 + ) in the lungs ( Right ). ( G ) Absolute cell numbers of eosinophils (Eos.), neutrophils (Neut.), lymphocytes (Lymph.), and macrophages (Mac.) in the BAL fluid are shown. ( H ) Airway hyperresponsiveness was assessed by measuring methacholine-induced changes in lung resistance. The mean values from five ( A , B , E , and F ), four ( G and H ), or three ( D ) mice per group or technical triplicates ( C ) are shown with SD or SEM. More than two independent experiments were performed with similar results ( A – F ; ** P

    Techniques Used: Mouse Assay, Knock-In, Staining, FACS, Expressing, Injection

    23) Product Images from "Dual-Sized Microparticle System for Generating Suppressive Dendritic Cells Prevents and Reverses Type 1 Diabetes in the Nonobese Diabetic Mouse Model"

    Article Title: Dual-Sized Microparticle System for Generating Suppressive Dendritic Cells Prevents and Reverses Type 1 Diabetes in the Nonobese Diabetic Mouse Model

    Journal: ACS Biomaterials Science & Engineering

    doi: 10.1021/acsbiomaterials.9b00332

    Co-incubation of vitamin D 3 (VD 3 ) MPs and TGF-β1 MPs induce DCs with suppressive phenotypes in vitro. Dendritic cells were incubated with 10 mg of nonphagocytosable TGF-β1 MPs, and phagocytosable VD 3 MPs were added at a 10:1 MP to DC ratio. Microparticles were incubated with bone marrow-derived DCs for 48 h and subsequently washed with PBS to remove MPs. Untreated, immature DCs (iDC), DCs stimulated with LPS (1 μg/mL), and DCs incubated with unloaded MPs were included as controls. (A) Maturation markers CD80, CD86, and MHC-II were characterized by flow cytometry on MP-treated DCs and controls ( n = 3). Surface expression is normalized to iDCs. (B) Maturation resistance in response to LPS was quantified ( n = 3). Dendritic cells were stimulated with LPS (1 μg/mL) for 24 h following MP treatment. Flow cytometric analysis quantified expression of CD80, CD86, and MHC-II. Surface expression is normalized to LPS stimulated DCs. (C) Dendritic cell expression of the immunosuppressive enzyme IDO was quantified in response to MP treatment ( n = 3). P -values (∗ = ≤0.05, ∗∗ = ≤0.01, ∗∗∗ = ≤0.001) were obtained by one-way ANOVA with Dunnett’s multiple comparisons test against the iDC control (A, C) or the LPS-stimulated control (B). Data are represented by the mean ± SEM.
    Figure Legend Snippet: Co-incubation of vitamin D 3 (VD 3 ) MPs and TGF-β1 MPs induce DCs with suppressive phenotypes in vitro. Dendritic cells were incubated with 10 mg of nonphagocytosable TGF-β1 MPs, and phagocytosable VD 3 MPs were added at a 10:1 MP to DC ratio. Microparticles were incubated with bone marrow-derived DCs for 48 h and subsequently washed with PBS to remove MPs. Untreated, immature DCs (iDC), DCs stimulated with LPS (1 μg/mL), and DCs incubated with unloaded MPs were included as controls. (A) Maturation markers CD80, CD86, and MHC-II were characterized by flow cytometry on MP-treated DCs and controls ( n = 3). Surface expression is normalized to iDCs. (B) Maturation resistance in response to LPS was quantified ( n = 3). Dendritic cells were stimulated with LPS (1 μg/mL) for 24 h following MP treatment. Flow cytometric analysis quantified expression of CD80, CD86, and MHC-II. Surface expression is normalized to LPS stimulated DCs. (C) Dendritic cell expression of the immunosuppressive enzyme IDO was quantified in response to MP treatment ( n = 3). P -values (∗ = ≤0.05, ∗∗ = ≤0.01, ∗∗∗ = ≤0.001) were obtained by one-way ANOVA with Dunnett’s multiple comparisons test against the iDC control (A, C) or the LPS-stimulated control (B). Data are represented by the mean ± SEM.

    Techniques Used: Incubation, In Vitro, Derivative Assay, Flow Cytometry, Cytometry, Expressing

    24) Product Images from "CD8 T cell-mediated killing of orexinergic neurons induces a narcolepsy-like phenotype in mice"

    Article Title: CD8 T cell-mediated killing of orexinergic neurons induces a narcolepsy-like phenotype in mice

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    doi: 10.1073/pnas.1603325113

    CTL-mediated cytotoxicity contributes to orexin + neuron loss in Orex-HA mice. ( A and B ) Representative confocal micrographs of CTLs (anti-CD8; red) and orexin + neurons (anti–orexin-A; green) ( A ) or MHC class I expression (anti–β2-microglobulin;
    Figure Legend Snippet: CTL-mediated cytotoxicity contributes to orexin + neuron loss in Orex-HA mice. ( A and B ) Representative confocal micrographs of CTLs (anti-CD8; red) and orexin + neurons (anti–orexin-A; green) ( A ) or MHC class I expression (anti–β2-microglobulin;

    Techniques Used: CTL Assay, Mouse Assay, Expressing

    25) Product Images from "Hapten sensitization to vaginal mucosa induces less recruitment of dendritic cells accompanying TGF‐β‐expressing CD206+ cells compared with skin. Hapten sensitization to vaginal mucosa induces less recruitment of dendritic cells accompanying TGF‐β‐expressing CD206+ cells compared with skin"

    Article Title: Hapten sensitization to vaginal mucosa induces less recruitment of dendritic cells accompanying TGF‐β‐expressing CD206+ cells compared with skin. Hapten sensitization to vaginal mucosa induces less recruitment of dendritic cells accompanying TGF‐β‐expressing CD206+ cells compared with skin

    Journal: Immunity, Inflammation and Disease

    doi: 10.1002/iid3.605

    Changes in DC migration to dLNs after VM sensitization with FITC application compared with skin sensitization. (A) Changes in the number of total cells 6, 24, and 48 h after sensitization and without sensitization (0 h). (B) Representative flow cytometry data showing the percentage of FITC + cells among 7‐ADD − MHC class II high CD11c + cells in dLNs from naïve and sensitized mice (see Figure S1 for gating strategy). Changes in the proportion of FITC + cells among 7‐ADD − MHC class II high CD11c + cells (C) and the number of FITC + MHC class II high CD11c + migratory DCs (D) 6, 24, 48 h after sensitization and without sensitization (0 h). Data show the mean values ± SD ( n = 3–9). * p
    Figure Legend Snippet: Changes in DC migration to dLNs after VM sensitization with FITC application compared with skin sensitization. (A) Changes in the number of total cells 6, 24, and 48 h after sensitization and without sensitization (0 h). (B) Representative flow cytometry data showing the percentage of FITC + cells among 7‐ADD − MHC class II high CD11c + cells in dLNs from naïve and sensitized mice (see Figure S1 for gating strategy). Changes in the proportion of FITC + cells among 7‐ADD − MHC class II high CD11c + cells (C) and the number of FITC + MHC class II high CD11c + migratory DCs (D) 6, 24, 48 h after sensitization and without sensitization (0 h). Data show the mean values ± SD ( n = 3–9). * p

    Techniques Used: Migration, Flow Cytometry, Mouse Assay

    Changes in DC distribution and number in the skin and VM after DNFB sensitization. Representative images of DCs immunostained using anti‐MHC class II (green) (A) and anti‐CD11c (green) (B), and counterstained with DAPI (blue) in sections of intact and DNFB applied tissues at 6 and 24 h. The dotted areas indicate the epidermis and epithelium. Scale bar, 50 µm. (C) Quantification of the number of MHC class II + LCs per unit length of basement membrane in the epidermis and epithelium. (D) Quantification of the number of MHC class II + dermal and submucosal DCs per unit length of basement membrane in the dermis and lamina propria. Data show the mean values ± SD ( n = 3). * p
    Figure Legend Snippet: Changes in DC distribution and number in the skin and VM after DNFB sensitization. Representative images of DCs immunostained using anti‐MHC class II (green) (A) and anti‐CD11c (green) (B), and counterstained with DAPI (blue) in sections of intact and DNFB applied tissues at 6 and 24 h. The dotted areas indicate the epidermis and epithelium. Scale bar, 50 µm. (C) Quantification of the number of MHC class II + LCs per unit length of basement membrane in the epidermis and epithelium. (D) Quantification of the number of MHC class II + dermal and submucosal DCs per unit length of basement membrane in the dermis and lamina propria. Data show the mean values ± SD ( n = 3). * p

    Techniques Used:

    26) Product Images from "CD40Ig treatment results in allograft acceptance mediated by CD8+CD45RClow T cells, IFN-?, and indoleamine 2,3-dioxygenase"

    Article Title: CD40Ig treatment results in allograft acceptance mediated by CD8+CD45RClow T cells, IFN-?, and indoleamine 2,3-dioxygenase

    Journal: Journal of Clinical Investigation

    doi: 10.1172/JCI28801

    CD8 + T cells mediate transfer of transplantation tolerance after CD40Ig treatment. Cells from control rats that had rejected their grafts or from CD40Ig-treated recipients were injected i.v. the day of transplantation into LEW.1A recipients that received LEW.1W heart transplants (day 0) and that were sublethally irradiated (4.5 Gy, at day –1). ( A ) Total splenocytes (50 × 10 6 ; n = 6) or splenocytes depleted of T cells ( n = 2), CD4 + cells ( n = 2), CD8 + cells ( n = 4), or NK cells ( n = 2) were injected. ( B ) Splenocytes (50 × 10 6 ; n = 6), purified T cells ( n = 3), CD8 + T cells ( n = 4), or CD4 + T cells ( n = 2) were injected. Graft survival was assessed by abdominal palpation of cardiac beating. ( C ) Grafts were either untreated ( n = 9; thick black line), treated with a control mAb (3G8) ( n = 6, gray line), transduced with Addl324 (5 × 10 10 IP; n = 9; dotted line), transduced with AdCD40Ig ( n = 27, black line with open circles), or transduced with AdCD40Ig and injected with a depleting anti-CD8 mAb (OX8) ( n = 12; black line with open triangles) or an anti-MHC class I mAb (OX18) ( n = 3; black line with filled diamonds). A group of control animals received the control mAb (3G8) in addition to CD40Ig ( n = 6; black line with open diamonds). ** P
    Figure Legend Snippet: CD8 + T cells mediate transfer of transplantation tolerance after CD40Ig treatment. Cells from control rats that had rejected their grafts or from CD40Ig-treated recipients were injected i.v. the day of transplantation into LEW.1A recipients that received LEW.1W heart transplants (day 0) and that were sublethally irradiated (4.5 Gy, at day –1). ( A ) Total splenocytes (50 × 10 6 ; n = 6) or splenocytes depleted of T cells ( n = 2), CD4 + cells ( n = 2), CD8 + cells ( n = 4), or NK cells ( n = 2) were injected. ( B ) Splenocytes (50 × 10 6 ; n = 6), purified T cells ( n = 3), CD8 + T cells ( n = 4), or CD4 + T cells ( n = 2) were injected. Graft survival was assessed by abdominal palpation of cardiac beating. ( C ) Grafts were either untreated ( n = 9; thick black line), treated with a control mAb (3G8) ( n = 6, gray line), transduced with Addl324 (5 × 10 10 IP; n = 9; dotted line), transduced with AdCD40Ig ( n = 27, black line with open circles), or transduced with AdCD40Ig and injected with a depleting anti-CD8 mAb (OX8) ( n = 12; black line with open triangles) or an anti-MHC class I mAb (OX18) ( n = 3; black line with filled diamonds). A group of control animals received the control mAb (3G8) in addition to CD40Ig ( n = 6; black line with open diamonds). ** P

    Techniques Used: Transplantation Assay, Injection, Irradiation, Purification, Transduction

    27) Product Images from "Ascophyllan functions as an adjuvant to promote anti-cancer effect by dendritic cell activation"

    Article Title: Ascophyllan functions as an adjuvant to promote anti-cancer effect by dendritic cell activation

    Journal: Oncotarget

    doi: 10.18632/oncotarget.8200

    Ascophyllan promotes Ag-specific CD4 and CD8 T cell proliferation in the tumor-bearing mice B16 tumor-bearing C57BL/6 mice were injected with PBS, OVA or combination of ascophyllan and OVA for 24 hours. A. Percentage of lineage − CD11c + CD8α + and lineage − CD11c + CD8α − cDCs in spleen was analyzed on a flow cytometry (upper panel). Expression levels of MHC class I and II in the CD11c + CD8α + and CD11c + CD8α − cDCs are shown (lower panel). B. Mean of absolute number of CD11c + CD8α + and CD11c + CD8α − cDCs within live cells in spleen. C. Presentation of OVA peptide in the CD11c + CD8α + and CD11c + CD8α − cDCs was analyzed by Flow cytometry. D. Purified CD8 T cells from OT-I or CD4 T cells from OT-II mice were labeled with CFSE and transferred into B16 tumor-bearing CD45.1 congenic mice, and 24 hours later, mice were injected with PBS, OVA or combination of ascophyllan and OVA. On day 3 after ascophyllan and OVA injection, splenocytes from these mice were stained for CD45.2 to identify the donor OT-I or OT-II cells, and the proliferation of these cells was determined by CFSE dilution. E. Mean of absolute numbers of OT-I (left panel) and OT-II (right panel) cells in the tumor are shown. F. Percentage of IFN-γ + and TNF-α + cells in tumor-infiltrated OT-I and OT-II cells. All data are representative of or the average of analyses of 6 independent samples (2 mice per experiment, total 3 independent experiments). ** p
    Figure Legend Snippet: Ascophyllan promotes Ag-specific CD4 and CD8 T cell proliferation in the tumor-bearing mice B16 tumor-bearing C57BL/6 mice were injected with PBS, OVA or combination of ascophyllan and OVA for 24 hours. A. Percentage of lineage − CD11c + CD8α + and lineage − CD11c + CD8α − cDCs in spleen was analyzed on a flow cytometry (upper panel). Expression levels of MHC class I and II in the CD11c + CD8α + and CD11c + CD8α − cDCs are shown (lower panel). B. Mean of absolute number of CD11c + CD8α + and CD11c + CD8α − cDCs within live cells in spleen. C. Presentation of OVA peptide in the CD11c + CD8α + and CD11c + CD8α − cDCs was analyzed by Flow cytometry. D. Purified CD8 T cells from OT-I or CD4 T cells from OT-II mice were labeled with CFSE and transferred into B16 tumor-bearing CD45.1 congenic mice, and 24 hours later, mice were injected with PBS, OVA or combination of ascophyllan and OVA. On day 3 after ascophyllan and OVA injection, splenocytes from these mice were stained for CD45.2 to identify the donor OT-I or OT-II cells, and the proliferation of these cells was determined by CFSE dilution. E. Mean of absolute numbers of OT-I (left panel) and OT-II (right panel) cells in the tumor are shown. F. Percentage of IFN-γ + and TNF-α + cells in tumor-infiltrated OT-I and OT-II cells. All data are representative of or the average of analyses of 6 independent samples (2 mice per experiment, total 3 independent experiments). ** p

    Techniques Used: Mouse Assay, Injection, Flow Cytometry, Cytometry, Expressing, Purification, Labeling, Staining

    In vivo administration of ascophyllan induces maturation of DCs in the tumor-bearing mice C57BL/6 mice were injected subcutaneously ( s.c. ) with 1 × 10 6 B16 melanoma cells. On day 10 after tumor cell injection, once tumors were well established, the mice were treated with 50 mg/kg ascophyllan for 24 hours and spleen and tumor draining lymph node (drLN) were harvested. A. Percentages of lineage − CD11c + DCs in spleen and tumor drLN were analyzed on a flow cytometry. Lineage markers included CD3, Thy1.1, B220, Gr-1, CD49b and TER-119. B. Mean of absolute numbers of lineage − CD11c + cells within live cells in spleen (left panel) and tumor drLN (right panel) are shown. C. Expression levels of CCR7 in gated lineage − CD11c + cells from spleen and tumor drLN. D. Real-time PCR analysis of gene expression, presented relative to that of β-actin, in splenocytes. E. Isolated DCs from naïve mice were labeled with CellVeu Maroon and transferred into tumor-bearing mice. One hour after the DC transfer, the mice were treated with ascophyllan for 24 hours. Migration of CellVeu Maroon-labeled DCs was determined by flow cytometry. F. Flow cytometric analysis of co-stimulatory molecules and MHC class I and II in gated lineage − CD11c + cells from spleen and tumor drLN. G. IL-6, IL-12p70 and TNF-α levels in sera are shown. H. Intracellular IL-6, IL-12 and TNF-α production in spleen DCs. All data are representative of or the average of analyses of 6 independent samples (2 mice per experiment, total 3 independent experiments). * p
    Figure Legend Snippet: In vivo administration of ascophyllan induces maturation of DCs in the tumor-bearing mice C57BL/6 mice were injected subcutaneously ( s.c. ) with 1 × 10 6 B16 melanoma cells. On day 10 after tumor cell injection, once tumors were well established, the mice were treated with 50 mg/kg ascophyllan for 24 hours and spleen and tumor draining lymph node (drLN) were harvested. A. Percentages of lineage − CD11c + DCs in spleen and tumor drLN were analyzed on a flow cytometry. Lineage markers included CD3, Thy1.1, B220, Gr-1, CD49b and TER-119. B. Mean of absolute numbers of lineage − CD11c + cells within live cells in spleen (left panel) and tumor drLN (right panel) are shown. C. Expression levels of CCR7 in gated lineage − CD11c + cells from spleen and tumor drLN. D. Real-time PCR analysis of gene expression, presented relative to that of β-actin, in splenocytes. E. Isolated DCs from naïve mice were labeled with CellVeu Maroon and transferred into tumor-bearing mice. One hour after the DC transfer, the mice were treated with ascophyllan for 24 hours. Migration of CellVeu Maroon-labeled DCs was determined by flow cytometry. F. Flow cytometric analysis of co-stimulatory molecules and MHC class I and II in gated lineage − CD11c + cells from spleen and tumor drLN. G. IL-6, IL-12p70 and TNF-α levels in sera are shown. H. Intracellular IL-6, IL-12 and TNF-α production in spleen DCs. All data are representative of or the average of analyses of 6 independent samples (2 mice per experiment, total 3 independent experiments). * p

    Techniques Used: In Vivo, Mouse Assay, Injection, Flow Cytometry, Cytometry, Expressing, Real-time Polymerase Chain Reaction, Isolation, Labeling, Migration

    28) Product Images from "Langerhans Cell Homeostasis and Activation Is Altered in Hyperplastic Human Papillomavirus Type 16 E7 Expressing Epidermis"

    Article Title: Langerhans Cell Homeostasis and Activation Is Altered in Hyperplastic Human Papillomavirus Type 16 E7 Expressing Epidermis

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0127155

    LC migration from the epidermis is reduced the in K14E7 mice. C57Bl/6 and K14E7 mice were painted with FITC on a 15mm 2 area of abdomen and inguinal lymph nodes harvested 48 h later. Single cell suspensions were stained with CD11c, MHC class II and CD207 (A). The absolute number of FITC positive cells was determined (B). The total migrated LC in the lymph nodes as a percentage of the mean total number of LC in the painted skin is shown (C). * P
    Figure Legend Snippet: LC migration from the epidermis is reduced the in K14E7 mice. C57Bl/6 and K14E7 mice were painted with FITC on a 15mm 2 area of abdomen and inguinal lymph nodes harvested 48 h later. Single cell suspensions were stained with CD11c, MHC class II and CD207 (A). The absolute number of FITC positive cells was determined (B). The total migrated LC in the lymph nodes as a percentage of the mean total number of LC in the painted skin is shown (C). * P

    Techniques Used: Migration, Mouse Assay, Staining

    Antigen uptake is reduced in K14E7 mice. Epidermal cell suspensions from C57BL/6 and K14E7 mice were cultured with Alexa-555-labelled OVA overnight at 37°C, stained and analysed by flow cytometry (A). The Alexa-555-OVA+ LC (CD45.2+, MHC Class II+, CD207+) as a percentage of total LC (B) and the level of expression of Alexa-555-OVA on the positive stained LC is shown (C). ** P
    Figure Legend Snippet: Antigen uptake is reduced in K14E7 mice. Epidermal cell suspensions from C57BL/6 and K14E7 mice were cultured with Alexa-555-labelled OVA overnight at 37°C, stained and analysed by flow cytometry (A). The Alexa-555-OVA+ LC (CD45.2+, MHC Class II+, CD207+) as a percentage of total LC (B) and the level of expression of Alexa-555-OVA on the positive stained LC is shown (C). ** P

    Techniques Used: Mouse Assay, Cell Culture, Staining, Flow Cytometry, Cytometry, Expressing

    29) Product Images from "Mycobacterium tuberculosis Rv3628 isan effective adjuvant via activationof dendritic cells for cancer immunotherapy"

    Article Title: Mycobacterium tuberculosis Rv3628 isan effective adjuvant via activationof dendritic cells for cancer immunotherapy

    Journal: Molecular Therapy Oncolytics

    doi: 10.1016/j.omto.2021.10.003

    Rv3628-promoted DCs maturation in the tumor microenvironment (A) C57BL/6 CD45-2 mice were injected s.c. with 1 × 10 6 B16-OVA melanoma cells. Fifteen days after tumor injection, the mice were injected s.c. with PBS, 2.5 mg/kg Rv3628, and 1 mg/kg LPS for 24 h, and the mLN were harvested. The expression levels of co-stimulatory molecules and MHC classes I and II in CD8α + DCs (upper) and CD8α − DCs (lower) are shown ( n = 6 mice, one-way ANOVA, mean ± SEM). (B) CD45-1 mice bearing B16-OVA tumors were injected with fluorescent-labeled OVA-specific CD8 + OT-I and CD4 + OT-II cells and 24 h later injected with Ag. The proliferation of the cells in tumor-draining mLNs was measured 72 h later. (C–E) The division index (C), the mean percentages of the proliferating cells (D), and the absolute numbers of OT-I and OT-II cells (E) ( n = 6 mice, two-way ANOVA, mean ± SEM). (F) The mean percentages of IFN-γ- and e TNF-α-producing OT-I and OT-II cells.
    Figure Legend Snippet: Rv3628-promoted DCs maturation in the tumor microenvironment (A) C57BL/6 CD45-2 mice were injected s.c. with 1 × 10 6 B16-OVA melanoma cells. Fifteen days after tumor injection, the mice were injected s.c. with PBS, 2.5 mg/kg Rv3628, and 1 mg/kg LPS for 24 h, and the mLN were harvested. The expression levels of co-stimulatory molecules and MHC classes I and II in CD8α + DCs (upper) and CD8α − DCs (lower) are shown ( n = 6 mice, one-way ANOVA, mean ± SEM). (B) CD45-1 mice bearing B16-OVA tumors were injected with fluorescent-labeled OVA-specific CD8 + OT-I and CD4 + OT-II cells and 24 h later injected with Ag. The proliferation of the cells in tumor-draining mLNs was measured 72 h later. (C–E) The division index (C), the mean percentages of the proliferating cells (D), and the absolute numbers of OT-I and OT-II cells (E) ( n = 6 mice, two-way ANOVA, mean ± SEM). (F) The mean percentages of IFN-γ- and e TNF-α-producing OT-I and OT-II cells.

    Techniques Used: Mouse Assay, Injection, Expressing, Labeling

    Rv3628 promotes the activation of BMDCs and splenic DCs (A) BMDCs were treated with 1, 5, and 10 μg/mL Rv3628 and with 100 ng/mL LPS as a positive control analyzed after 24 h. Mean fluorescence intensity levels of CD80, CD86, MHC class I, and MHC class II in BMDCs are shown ( n = 6, one-way ANOVA, mean ± SEM). (B) C57BL/6 mice were injected subcutaneously with 2.5 mg/kg Rv3628 and with 1 mg/kg LPS as a positive control and splenocytes harvested after 24 h. Lineage markers included CD3, Thy1.1, B220, Gr-1, CD49b, and TER-119. Mean fluorescence intensity (MFI) of co-stimulatory molecules and MHC classes I and II in CD8α + Lin − CD11c + cells (upper) and CD8α − Lin − CD11c + cells (lower) are shown ( n = 6 mice, one-way ANOVA, mean ± SEM). (C) Concentrations of TNF-α, IL-6, and IL-12p70 in BMDC-cultured medium from (A) ( n = 6, two-way ANOVA, mean ± SEM). (D) The serum concentration of TNF-α, IL-6, and IL-12p70 from (B) ( n = 6 mice, two-way ANOVA, mean ± SEM). (E) Mean values of the mRNA expression levels in the spleen from (B) ( n = 4 mice, two-way ANOVA, mean ± SEM).
    Figure Legend Snippet: Rv3628 promotes the activation of BMDCs and splenic DCs (A) BMDCs were treated with 1, 5, and 10 μg/mL Rv3628 and with 100 ng/mL LPS as a positive control analyzed after 24 h. Mean fluorescence intensity levels of CD80, CD86, MHC class I, and MHC class II in BMDCs are shown ( n = 6, one-way ANOVA, mean ± SEM). (B) C57BL/6 mice were injected subcutaneously with 2.5 mg/kg Rv3628 and with 1 mg/kg LPS as a positive control and splenocytes harvested after 24 h. Lineage markers included CD3, Thy1.1, B220, Gr-1, CD49b, and TER-119. Mean fluorescence intensity (MFI) of co-stimulatory molecules and MHC classes I and II in CD8α + Lin − CD11c + cells (upper) and CD8α − Lin − CD11c + cells (lower) are shown ( n = 6 mice, one-way ANOVA, mean ± SEM). (C) Concentrations of TNF-α, IL-6, and IL-12p70 in BMDC-cultured medium from (A) ( n = 6, two-way ANOVA, mean ± SEM). (D) The serum concentration of TNF-α, IL-6, and IL-12p70 from (B) ( n = 6 mice, two-way ANOVA, mean ± SEM). (E) Mean values of the mRNA expression levels in the spleen from (B) ( n = 4 mice, two-way ANOVA, mean ± SEM).

    Techniques Used: Activation Assay, Positive Control, Fluorescence, Mouse Assay, Injection, Cell Culture, Concentration Assay, Expressing

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    Thermo Fisher mouse anti rat mhc ii
    Immunofluorescence staining of differentiation, proliferation, and immune antigen marker expression of transplanted MSCs. Rats were euthanized at 7, 14 and 28 days post-transplantation and their lungs and pancreas were obtained in the TVT and PST groups, respectively. Representative micrographs illustrate the expression of GFP (green, transplanted cells), immune antigens (red, CD86 and <t>MHC</t> II), a proliferation marker (red, <t>PCNA)</t> and a differentiation marker (red, insulin) in the lung and pancreas tissues of recipients. GFP + MSCs were visible in all samples. By day 7, the expression of CD86, MHC II, PCNA and insulin was negative ( a ). However, expression of MHC II ( b ) and insulin ( c ) was observed at 14 and 28 days post-transplantation. Blue indicates nuclear staining with DAPI. MSC, mesenchymal stem cell; PST, pancreas subcapsular transplantation; TVT, tail vein transplantation.
    Mouse Anti Rat Mhc Ii, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher major histocompatibility complex class ii mhcii
    Prostate-specific Gr-1 + <t>CD11b</t> + cell expansion is associated with suppression of dendritic cell and macrophage maturation. (A, left) Mature CD11c + <t>MHCII</t> + dendritic cells are present in both wild-type and mutant prostates, but their levels decrease precipitously
    Major Histocompatibility Complex Class Ii Mhcii, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher anti canine mhc ii
    Flow cytometry results for expression of CD45, CD3, CD21, <t>CD79a,</t> CD14, CD172a, CD11c, <t>MHC</t> I, and MHC II. Expression of MHC II was also evaluated on cells treated with either LPS or IFNγ for 24 and 48 h. (BD cell line shown in red, isotype control in blue)
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    Thermo Fisher anti cd11c
    Type I and II IFNs restrict L . pneumophila infection through an alveolar macrophage-intrinsic mechanism. (A) Overview of generation of <t>CD11c-DTR-GFP</t> / Ifnar / Ifngr -/- mixed bone marrow chimeric mice followed by DTX mediated depletion of CD11c-DTR-GFP + cells. (B) Frequency of remaining CD11c + GFP + wild-type cells was correlated to bacterial load in the lungs of CD11c-DTR-GFP / Ifnar / Ifngr -/- + DTX chimeras including all DTX-treated mice (13 mice) at 6 d p.i.. (C) Only mice with
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    Immunofluorescence staining of differentiation, proliferation, and immune antigen marker expression of transplanted MSCs. Rats were euthanized at 7, 14 and 28 days post-transplantation and their lungs and pancreas were obtained in the TVT and PST groups, respectively. Representative micrographs illustrate the expression of GFP (green, transplanted cells), immune antigens (red, CD86 and MHC II), a proliferation marker (red, PCNA) and a differentiation marker (red, insulin) in the lung and pancreas tissues of recipients. GFP + MSCs were visible in all samples. By day 7, the expression of CD86, MHC II, PCNA and insulin was negative ( a ). However, expression of MHC II ( b ) and insulin ( c ) was observed at 14 and 28 days post-transplantation. Blue indicates nuclear staining with DAPI. MSC, mesenchymal stem cell; PST, pancreas subcapsular transplantation; TVT, tail vein transplantation.

    Journal: Cellular and Molecular Immunology

    Article Title: Immunogenicity of allogeneic mesenchymal stem cells transplanted via different routes in diabetic rats

    doi: 10.1038/cmi.2014.70

    Figure Lengend Snippet: Immunofluorescence staining of differentiation, proliferation, and immune antigen marker expression of transplanted MSCs. Rats were euthanized at 7, 14 and 28 days post-transplantation and their lungs and pancreas were obtained in the TVT and PST groups, respectively. Representative micrographs illustrate the expression of GFP (green, transplanted cells), immune antigens (red, CD86 and MHC II), a proliferation marker (red, PCNA) and a differentiation marker (red, insulin) in the lung and pancreas tissues of recipients. GFP + MSCs were visible in all samples. By day 7, the expression of CD86, MHC II, PCNA and insulin was negative ( a ). However, expression of MHC II ( b ) and insulin ( c ) was observed at 14 and 28 days post-transplantation. Blue indicates nuclear staining with DAPI. MSC, mesenchymal stem cell; PST, pancreas subcapsular transplantation; TVT, tail vein transplantation.

    Article Snippet: The sections were then incubated with a rabbit anti-GFP antibody (1∶800; Abcam, Cambridge, UK) either alone or with mouse anti-rat PCNA (GeneTex, Alton PkwyIrvine, CA, USA), mouse anti-rat MHC II (1∶200; eBioscience), mouse anti-rat CD86 (1∶200; eBioscience) or mouse anti-rat insulin (1∶400; Abcam) antibodies at 4 °C overnight.

    Techniques: Immunofluorescence, Staining, Marker, Expressing, Transplantation Assay

    Adipogenesis and osteogenesis of MSCs and immunogenicity testing in vitro . Cells were spindle-shaped after three passages and transduction with a GFP expression vector ( aI and II ). MSCs were then cultured in appropriate induction media for 3 weeks. Adipogenesis was confirmed by Oil Red O staining and osteogenesis was confirmed by Alizarin red staining ( aIII and IV , respectively). FACS showed that P7 MSCs expressed a low level of MHC I and did not express MHC II, CD40, CD80 or CD86 ( b ). PBMCs from normal and diabetic rats were cocultured with MSCs for 72 h. No proliferation was observed in the MSC group compared with the autoproliferation and ConA (positive) group ( c ). ** P

    Journal: Cellular and Molecular Immunology

    Article Title: Immunogenicity of allogeneic mesenchymal stem cells transplanted via different routes in diabetic rats

    doi: 10.1038/cmi.2014.70

    Figure Lengend Snippet: Adipogenesis and osteogenesis of MSCs and immunogenicity testing in vitro . Cells were spindle-shaped after three passages and transduction with a GFP expression vector ( aI and II ). MSCs were then cultured in appropriate induction media for 3 weeks. Adipogenesis was confirmed by Oil Red O staining and osteogenesis was confirmed by Alizarin red staining ( aIII and IV , respectively). FACS showed that P7 MSCs expressed a low level of MHC I and did not express MHC II, CD40, CD80 or CD86 ( b ). PBMCs from normal and diabetic rats were cocultured with MSCs for 72 h. No proliferation was observed in the MSC group compared with the autoproliferation and ConA (positive) group ( c ). ** P

    Article Snippet: The sections were then incubated with a rabbit anti-GFP antibody (1∶800; Abcam, Cambridge, UK) either alone or with mouse anti-rat PCNA (GeneTex, Alton PkwyIrvine, CA, USA), mouse anti-rat MHC II (1∶200; eBioscience), mouse anti-rat CD86 (1∶200; eBioscience) or mouse anti-rat insulin (1∶400; Abcam) antibodies at 4 °C overnight.

    Techniques: In Vitro, Transduction, Expressing, Plasmid Preparation, Cell Culture, Staining, FACS

    Prostate-specific Gr-1 + CD11b + cell expansion is associated with suppression of dendritic cell and macrophage maturation. (A, left) Mature CD11c + MHCII + dendritic cells are present in both wild-type and mutant prostates, but their levels decrease precipitously

    Journal: Molecular and Cellular Biology

    Article Title: Pten Null Prostate Epithelium Promotes Localized Myeloid-Derived Suppressor Cell Expansion and Immune Suppression during Tumor Initiation and Progression

    doi: 10.1128/MCB.00090-14

    Figure Lengend Snippet: Prostate-specific Gr-1 + CD11b + cell expansion is associated with suppression of dendritic cell and macrophage maturation. (A, left) Mature CD11c + MHCII + dendritic cells are present in both wild-type and mutant prostates, but their levels decrease precipitously

    Article Snippet: Single-cell suspensions were stained with directly conjugated antibodies against CD45, Gr-1, CD11b, CD4, CD8, CD69, B220 (BD Biosciences), F4/80, CD19, CD11c, major histocompatibility complex class II (MHCII) (eBioscience), and Ly6C (BioLegend), according to the manufacturers' instructions.

    Techniques: Mutagenesis

    Flow cytometry results for expression of CD45, CD3, CD21, CD79a, CD14, CD172a, CD11c, MHC I, and MHC II. Expression of MHC II was also evaluated on cells treated with either LPS or IFNγ for 24 and 48 h. (BD cell line shown in red, isotype control in blue)

    Journal: BMC Cancer

    Article Title: A novel canine histiocytic sarcoma cell line: initial characterization and utilization for drug screening studies

    doi: 10.1186/s12885-018-4132-0

    Figure Lengend Snippet: Flow cytometry results for expression of CD45, CD3, CD21, CD79a, CD14, CD172a, CD11c, MHC I, and MHC II. Expression of MHC II was also evaluated on cells treated with either LPS or IFNγ for 24 and 48 h. (BD cell line shown in red, isotype control in blue)

    Article Snippet: BD cells harvested from cell culture were labeled with the following monoclonal antibodies: anti-canine CD3 (CA17.2A12, Serotec), anti-canine CD11c (CA11.6A1, UC Davis/NIH NeuroMab Facility), anti-bovine CD14 (MM61A, WSU MAC), anti-canine CD21 (CA2.1D6, Serotec), anti-canine CD45 (YKIX716.13, Serotec), anti-canine CD79a (HM57, LS Bio), anti-bovine CD172a (DH59B, WSU MAC), anti-canine MHC II (YKIX334.2, eBioscience), and anti-bovine MHC I (MHC CL I, WSU MAC).

    Techniques: Flow Cytometry, Cytometry, Expressing

    Type I and II IFNs restrict L . pneumophila infection through an alveolar macrophage-intrinsic mechanism. (A) Overview of generation of CD11c-DTR-GFP / Ifnar / Ifngr -/- mixed bone marrow chimeric mice followed by DTX mediated depletion of CD11c-DTR-GFP + cells. (B) Frequency of remaining CD11c + GFP + wild-type cells was correlated to bacterial load in the lungs of CD11c-DTR-GFP / Ifnar / Ifngr -/- + DTX chimeras including all DTX-treated mice (13 mice) at 6 d p.i.. (C) Only mice with

    Journal: PLoS Pathogens

    Article Title: IFNs Modify the Proteome of Legionella-Containing Vacuoles and Restrict Infection Via IRG1-Derived Itaconic Acid

    doi: 10.1371/journal.ppat.1005408

    Figure Lengend Snippet: Type I and II IFNs restrict L . pneumophila infection through an alveolar macrophage-intrinsic mechanism. (A) Overview of generation of CD11c-DTR-GFP / Ifnar / Ifngr -/- mixed bone marrow chimeric mice followed by DTX mediated depletion of CD11c-DTR-GFP + cells. (B) Frequency of remaining CD11c + GFP + wild-type cells was correlated to bacterial load in the lungs of CD11c-DTR-GFP / Ifnar / Ifngr -/- + DTX chimeras including all DTX-treated mice (13 mice) at 6 d p.i.. (C) Only mice with

    Article Snippet: The lung cell suspension was labelled with anti-panCD45 (30-F11, eBioscience), anti-CD45.1 (A20, BD Pharmingen), anti-CD45.2 (104, BD Pharmingen), anti-Ly6G (1A8, BD Pharmingen), anti-CD11c (N418, eBioscience) anti-MHC-II (M5/114.15.2, eBioscience), anti-Siglec-F, (E50-2440, BD Pharmingen) and anti-CD64 (X54-5/7.1, BD Pharmingen).

    Techniques: Infection, Mouse Assay