- 12-9883-42 - Invitrogen Antibodies IKZF2 antibody

Submitted references Adiponectin/AdipoR1 Axis Promotes IL-10 Release by Human Regulatory T Cells.

Ramos-Ramírez P, Malmhäll C, Tliba O, Rådinger M, Bossios A
Frontiers in immunology 2021;12:677550

Differences in Maturation Status and Immune Phenotypes of Circulating Helios(+) and Helios(-) Tregs and Their Disrupted Correlations With Monocyte Subsets in Autoantibody-Positive T1D Individuals.

Zhang Y, Zhang J, Shi Y, Shen M, Lv H, Chen S, Feng Y, Chen H, Xu X, Yang T, Xu K
Frontiers in immunology 2021;12:628504

Notch4 signaling limits regulatory T-cell-mediated tissue repair and promotes severe lung inflammation in viral infections.

Harb H, Benamar M, Lai PS, Contini P, Griffith JW, Crestani E, Schmitz-Abe K, Chen Q, Fong J, Marri L, Filaci G, Del Zotto G, Pishesha N, Kolifrath S, Broggi A, Ghosh S, Gelmez MY, Oktelik FB, Cetin EA, Kiykim A, Kose M, Wang Z, Cui Y, Yu XG, Li JZ, Berra L, Stephen-Victor E, Charbonnier LM, Zanoni I, Ploegh H, Deniz G, De Palma R, Chatila TA
Immunity 2021 Jun 8;54(6):1186-1199.e7

Potential therapeutic effects of interleukin-35 on the differentiation of naïve T cells into Helios(+)Foxp3(+) Tregs in clinical and experimental acute respiratory distress syndrome.

Wang C, Xie K, Li K, Lin S, Xu F
Molecular immunology 2021 Apr;132:236-249

Distinct actions of the fermented beverage kefir on host behaviour, immunity and microbiome gut-brain modules in the mouse.

van de Wouw M, Walsh AM, Crispie F, van Leuven L, Lyte JM, Boehme M, Clarke G, Dinan TG, Cotter PD, Cryan JF
Microbiome 2020 May 18;8(1):67

CD19-targeted CAR regulatory T cells suppress B cell pathology without GvHD.

Imura Y, Ando M, Kondo T, Ito M, Yoshimura A
JCI insight 2020 Jul 23;5(14)

B7-CD28 co-stimulation modulates central tolerance via thymic clonal deletion and Treg generation through distinct mechanisms.

Watanabe M, Lu Y, Breen M, Hodes RJ
Nature communications 2020 Dec 8;11(1):6264

Umbilical cord blood‑derived Helios‑positive regulatory T cells promote angiogenesis in acute lymphoblastic leukemia in mice via CCL22 and the VEGFA‑VEGFR2 pathway.

Li X, Li D, Shi Q, Huang X, Ju X
Molecular medicine reports 2019 May;19(5):4195-4204

CD150(high) Bone Marrow Tregs Maintain Hematopoietic Stem Cell Quiescence and Immune Privilege via Adenosine.

Hirata Y, Furuhashi K, Ishii H, Li HW, Pinho S, Ding L, Robson SC, Frenette PS, Fujisaki J
Cell stem cell 2018 Mar 1;22(3):445-453.e5

Decreased Helios Expression in Regulatory T Cells in Acute Coronary Syndrome.

Jiang L, Chen F, Hu X, Hu Y, Wang Y, Zhang W, Peng Y, Cheng L
Disease markers 2017;2017:7909407

Ablation of Transcription Factor IRF4 Promotes Transplant Acceptance by Driving Allogenic CD4(+) T Cell Dysfunction.

Wu J, Zhang H, Shi X, Xiao X, Fan Y, Minze LJ, Wang J, Ghobrial RM, Xia J, Sciammas R, Li XC, Chen W
Immunity 2017 Dec 19;47(6):1114-1128.e6

Foxp3 and Toll-like receptor signaling balance T(reg) cell anabolic metabolism for suppression.

Gerriets VA, Kishton RJ, Johnson MO, Cohen S, Siska PJ, Nichols AG, Warmoes MO, de Cubas AA, MacIver NJ, Locasale JW, Turka LA, Wells AD, Rathmell JC
Nature immunology 2016 Dec;17(12):1459-1466

Human adipose-tissue derived mesenchymal stem cells induce functional de-novo regulatory T cells with methylated FOXP3 gene DNA.

Engela AU, Hoogduijn MJ, Boer K, Litjens NH, Betjes MG, Weimar W, Baan CC
Clinical and experimental immunology 2013 Aug;173(2):343-54

Helios is associated with CD4 T cells differentiating to T helper 2 and follicular helper T cells in vivo independently of Foxp3 expression.

Serre K, Bénézech C, Desanti G, Bobat S, Toellner KM, Bird R, Chan S, Kastner P, Cunningham AF, Maclennan IC, Mohr E
PloS one 2011;6(6):e20731

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Intracellular staining of C57Bl/6 mouse lymph node cells with Anti-Mouse/Rat Foxp3 eFluor® 450 (Product # 48-5773-82) and Armenian Hamster IgG Isotype Control PE (Product # 12-4888-81) (left) or Anti-Helios PE (right) using the Foxp3/Transcription Factor Buffer Set (Product # 00-5523-00) and protocol. CD4+ (Product # 17-0041-82) cells in the lymphocyte gate were used for analysis.
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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 5 Notch4 deficiency reprograms the lung Treg cell transcriptome of poly(I:C)-treated mice (A-C) Volcano plot (A), heatmap (B), and pathway analysis (C) of gene transcripts of lung Treg cells isolated from Foxp3 YFPCre and Foxp3 YFPCre Notch4 Delta/Delta mice treated with poly(I:C) (n = 4 and n = 5, respectively). (D) Flow cytometry histograms and graphical representation of lung tissue Treg cell expression of CD25, Helios, CTLA4, ICOS, and OX40 in Foxp3 YFPCre and Foxp3 YFPCre Notch4 Delta/Delta mice sampled on day 7 after poly(I:C) treatment (n = 5 for each time point). (E-H) Flow cytometry analysis (E and G) and graphical representation (F and H) of Helios expression in Foxp3 + Notch4 + and Foxp3 + Notch4 - lung tissue Treg cells in Foxp3 YFPCre mice sampled at the indicated dates after poly(I:C) treatment (n = 5 for each time point). Each symbol represents one mouse. Numbers in flow plots indicate percentages. Error bars indicate SEM. Statistical tests: pairwise comparisons of differential gene expression were computed using DESeq2 (A-C); Student''s unpaired two-tailed t test (D) and two-way ANOVA with Sidak''s post hoc analysis (F and H). * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.
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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Fig. 3 UK4 increases Treg cells levels, while Fr1 decreases neutrophil levels. T regulatory cells (CD4+, CD25+, FoxP3+) were assessed in mesenteric lymph nodes (MLNs) ( a ) and were subsequently analysed for Helios expression ( b ), to investigate their origin as Helios+ cells exclusively originate in the thymus. Blood was also assessed for Treg cell levels ( c ), and plasma interleukin 10 (IL-10) levels ( d ). Similarly, neutrophil (CD11b+, LY6C mid , SSC high and CXCL1 levels were also investigated in peripheral blood ( e , f ). All data were normally distributed and analysed using a one-way ANOVA, followed by a Dunnett's post hoc test. Significant differences are depicted as * p < 0.05, ** p < 0.01; milk control compared to Kefir supplementation, $ p < 0.05 and $$ p < 0.01; undisturbed control compared to milk control. All data are expressed as mean +- SEM ( n = 11-12). Dots on each graph represent individual animals
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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Fig. 6 C57BL/5 mice were assigned to the untreated group, CLP (6 h) group, CLP (24 h) group, CLP (48 h) group, CLP + IgG group and CLP + IL-35 group. Each group had n = 5, the indicators of each mouse were repeatedly tested 3 times, and the best data were selected as the representative data. (A, B) T cells were isolated from mouse spleens or peripheral blood. The frequency of Foxp3+Helios + Treg cells was subsequently determined by flow cytometry, and the results reflect five mice per time point. Foxp3+Helios + Treg cells gradually decreased in the blood and spleen with time in CLP-induced ARDS. (C, D) Treatment with recombinant IL-35 after CLP challenge restored Foxp3+Helios + Treg cells in the blood and spleens. (E) Naive CD4 + T lymphocytes induce the differentiation of CD4 + Foxp3+ Helios + T cells (Tregs) via the IL-35/NF-kappaB signalling pathway. The comparison between the two groups was performed by one-way ANOVA followed by LSD multiple comparisons test. Fig. 6
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Supportive validation

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Experimental details: Fig. 2 In the absence of B7-CD28 co-stimulation, the frequency of cells undergoing clonal deletion at DP and CD4SP stages is decreased. a - d WT and B7 DKO thymocytes were gated on developmental stages as defined by TCR and CD69 expression in Fig. 1 . a Frequency of clonally deleting cells (Helios + PD-1 high ) was decreased in Stage III (CD4 + CD8 low ) thymocytes in B7 DKO mice. Each group n = 3. Data are representative of three independent experiments. p = 0.0365. b Frequency of clonally deleting cells (Helios + ) was decreased in Stage IV and V SP4 Tconv (CD25/Foxp3 nega ) in B7 DKO mice. Each group n = 3. Data are representative of three independent experiments. Stage IV p = 0.0004, Stage V p = 0.0003. c Frequency of clonally deleting cells (Active Caspase3 + ) was decreased in TCR-signaled DP and SP cells in B7 DKO mice. Each group n = 5. Data are pooled results of two independent experiments. Signaled DP p < 0.0001, Signaled SP4 p = 0.0038. d Frequency of clonally deleting cells (Helios + ) was decreased in Stage IV and V SP4 Tconv (CD25/Foxp3 nega ) in Bcl2-Tg/B7 DKO mice. WT n = 9, B7 DKO n = 3, Bcl2-Tg n = 5, Bcl2-Tg/B7 DKO n = 3. Data are pooled results of three independent experiments. Stage IV (WT and B7DKO) p = 0.0003, Stage IV (Bcl2-Tg and Bcl2-Tg/B7DKO) p = 0.0037, Stage V (WT and B7DKO) p = 0.0008, Stage V (Bcl2-Tg and Bcl2-Tg/B7DKO) p = 0.020. e Nur77-GFP high SP4 cells were increased in B7 DKO mice. Left panel, representative FACS plot. Right panel, quantif
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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 1 Human Circulating FOXP3 + Tregs Express AdipoR1. Flow cytometry analysis of AdipoR1 expression in Treg subsets was performed in peripheral whole blood from healthy donors. (A) Gating strategy of human circulating CD4 + CD25 + T cells. Dead cells, positive for LD Aqua dye, were excluded and lymphocytes were gated. CD4 + T cells were gated from single cells (excluding of doublets, FSC-A vs FSC-H) and analyzed for CD25 expression. (B) Helios + and Helios - FOXP3 + Tregs were gated on CD4 + CD25 + cells and then analyzed for AdipoR1 expression. FACS plots are from a representative experiment. (C) Frequency of AdipoR1 + cells and (D) MFI (mean fluorescence intensity) AdipoR1 in Helios + and Helios - FOXP3 + Tregs. Data were obtained from four independent experiments and are shown as the mean +- SEM of n=5. Student's t-test, *P< 0.05 and **P< 0.01 .
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12-9883-42

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