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anti il 33 neutralizing antibody  (Bio-Techne corporation)


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    Bio-Techne corporation anti il 33 neutralizing antibody
    a , Activity of A-Eos-specific regulons across clusters. g, number of genes in regulon. b , Expression of NF-κB signalling components. c , Quantification of pNF-κB p65 + cells in colonic A-Eos and B-Eos ( n = 3, B6J). Data are mean ± s.e.m. Two-tailed unpaired Student’s t -test. d , e , A-Eos and B-Eos frequencies in antibiotic-treated ( d ) ( n = 16, B6J) and germ-free (GF) ( e ) ( n = 9, B6J) mice relative to controls. SPF, specific pathogen free. d , Data are pooled from two independent experiments. Medians are shown. Two-tailed unpaired Student’s t -test. f , Depleted gene sets in PD-L1 + CD80 + A-Eos (red) and PD-L1 − CD80 − eosinophils (grey), relative to BM stem cells. Kolmogorov–Smirnov test. Dot size indicates gene-set size. Dashed line indicates P = 0.05. g , A-Eos frequencies after conditioning of BM-Eos <t>with</t> <t>IL-33,</t> colon CM and anti-IL-33 ( n = 2, pooled B6J). Technical replicates and mean ± s.e.m. are shown. One-way ANOVA. h , Colonic A-Eos and B-Eos frequencies in B6J ( n = 21) and Myd88 −/− ( n = 15) mice treated with IL-33, relative to untreated controls. Medians are shown. Two-tailed unpaired Student’s t -test. WT, wild type.
    Anti Il 33 Neutralizing Antibody, supplied by Bio-Techne corporation, used in various techniques. Bioz Stars score: 99/100, based on 285 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti il 33 neutralizing antibody/product/Bio-Techne corporation
    Average 99 stars, based on 285 article reviews
    anti il 33 neutralizing antibody - by Bioz Stars, 2026-03
    99/100 stars

    Images

    1) Product Images from "Active eosinophils regulate host defence and immune responses in colitis"

    Article Title: Active eosinophils regulate host defence and immune responses in colitis

    Journal: Nature

    doi: 10.1038/s41586-022-05628-7

    a , Activity of A-Eos-specific regulons across clusters. g, number of genes in regulon. b , Expression of NF-κB signalling components. c , Quantification of pNF-κB p65 + cells in colonic A-Eos and B-Eos ( n = 3, B6J). Data are mean ± s.e.m. Two-tailed unpaired Student’s t -test. d , e , A-Eos and B-Eos frequencies in antibiotic-treated ( d ) ( n = 16, B6J) and germ-free (GF) ( e ) ( n = 9, B6J) mice relative to controls. SPF, specific pathogen free. d , Data are pooled from two independent experiments. Medians are shown. Two-tailed unpaired Student’s t -test. f , Depleted gene sets in PD-L1 + CD80 + A-Eos (red) and PD-L1 − CD80 − eosinophils (grey), relative to BM stem cells. Kolmogorov–Smirnov test. Dot size indicates gene-set size. Dashed line indicates P = 0.05. g , A-Eos frequencies after conditioning of BM-Eos with IL-33, colon CM and anti-IL-33 ( n = 2, pooled B6J). Technical replicates and mean ± s.e.m. are shown. One-way ANOVA. h , Colonic A-Eos and B-Eos frequencies in B6J ( n = 21) and Myd88 −/− ( n = 15) mice treated with IL-33, relative to untreated controls. Medians are shown. Two-tailed unpaired Student’s t -test. WT, wild type.
    Figure Legend Snippet: a , Activity of A-Eos-specific regulons across clusters. g, number of genes in regulon. b , Expression of NF-κB signalling components. c , Quantification of pNF-κB p65 + cells in colonic A-Eos and B-Eos ( n = 3, B6J). Data are mean ± s.e.m. Two-tailed unpaired Student’s t -test. d , e , A-Eos and B-Eos frequencies in antibiotic-treated ( d ) ( n = 16, B6J) and germ-free (GF) ( e ) ( n = 9, B6J) mice relative to controls. SPF, specific pathogen free. d , Data are pooled from two independent experiments. Medians are shown. Two-tailed unpaired Student’s t -test. f , Depleted gene sets in PD-L1 + CD80 + A-Eos (red) and PD-L1 − CD80 − eosinophils (grey), relative to BM stem cells. Kolmogorov–Smirnov test. Dot size indicates gene-set size. Dashed line indicates P = 0.05. g , A-Eos frequencies after conditioning of BM-Eos with IL-33, colon CM and anti-IL-33 ( n = 2, pooled B6J). Technical replicates and mean ± s.e.m. are shown. One-way ANOVA. h , Colonic A-Eos and B-Eos frequencies in B6J ( n = 21) and Myd88 −/− ( n = 15) mice treated with IL-33, relative to untreated controls. Medians are shown. Two-tailed unpaired Student’s t -test. WT, wild type.

    Techniques Used: Activity Assay, Expressing, Two Tailed Test

    a , Experimental workflow of the CRISPR inhibition screen. b , Log10 negative score per gene, as calculated by MAGeCK. Cd80 and Cd274 evidenced in orange. Genes involved in TNF signalling pathway via NF-κB in red, and MAPK signalling pathway in darkred. c , IL-33 concentrations measured by ELISA in colon of DSS-treated mice (n = 12, B6J) and colon and blood of C. rodentium -infected mice (n = 7, B6J), compared to untreated controls (n = 7, B6J). Medians are shown. Two-tailed unpaired Student’s t -test. d , IFNγ, TNF and IL-22 concentrations measured by LEGENDplex in colon (left) and blood (right) of C. rod entium-infected mice (n = 7, B6J), compared to untreated controls (n = 7, B6J). Medians are shown. Two-tailed unpaired Student’s t -test. e , A-Eos (PD-L1 + CD80 + ) frequencies upon conditioning of BM-Eos with colon CM, IL-22, IL-25, TNF or IL-33 (n = 4, B6J). Data are pooled from two independent experiments. Medians are shown. One-way ANOVA. f , A-Eos frequencies after conditioning with increasing doses of IL-33. Input: BM-derived (n = 5, B6J), blood (n = 5, Il5 -tg) and splenic (n = 5, Il5 -tg) eosinophils. Medians are shown. One-way ANOVA. g , Western blot of phospho-p38 and phospho-p65 upon conditioning of BM-Eos with colon CM or IL-33 (n = 3, B6J). h , Gene expression normalized to Hprt measured by qRT–PCR of BM-Eos upon conditioning with IL-33 (n = 4, B6J). Data represents mean ± SEM. Two-tailed unpaired Student’s t -test. i , ST2 expression in BM-Eos upon IL-33 treatment (n = 4, B6J). Data represents mean ± SEM. Two-tailed unpaired Student’s t -test. j , ST2 expression in colonic A-Eos and B-Eos (n = 5, B6J). Data represents mean ± SEM. Two-tailed unpaired Student’s t -test. k , ST2 expression across organs (n = 5, B6J). Data represents mean ± SEM. One-way ANOVA. l , A-Eos frequencies upon conditioning of WT (n = 2, pooled B6J) or ST2 −/− (n = 2, pooled) BM-Eos with colon CM or IL-33. Technical replicates and mean ± SEM are shown. Two-tailed unpaired Student’s t -test. m , Left: Representative FACS plots of A-Eos (PD-L1 + CD80 + ) and PD-L1 − CD80 − eosinophils in the blood (top) and spleen (bottom). Numbers indicate % of eosinophils. Right: A-Eos frequencies in mice treated with IL-33 (n = 6-7, B6J). Medians are shown. Two-tailed unpaired Student’s t -test. n , A-Eos and B-Eos frequencies in the indicated organs of B6J (n = 7) and Il33 −/− (n = 5) mice at steady state. Medians are shown. Two-tailed unpaired Student’s t -test.
    Figure Legend Snippet: a , Experimental workflow of the CRISPR inhibition screen. b , Log10 negative score per gene, as calculated by MAGeCK. Cd80 and Cd274 evidenced in orange. Genes involved in TNF signalling pathway via NF-κB in red, and MAPK signalling pathway in darkred. c , IL-33 concentrations measured by ELISA in colon of DSS-treated mice (n = 12, B6J) and colon and blood of C. rodentium -infected mice (n = 7, B6J), compared to untreated controls (n = 7, B6J). Medians are shown. Two-tailed unpaired Student’s t -test. d , IFNγ, TNF and IL-22 concentrations measured by LEGENDplex in colon (left) and blood (right) of C. rod entium-infected mice (n = 7, B6J), compared to untreated controls (n = 7, B6J). Medians are shown. Two-tailed unpaired Student’s t -test. e , A-Eos (PD-L1 + CD80 + ) frequencies upon conditioning of BM-Eos with colon CM, IL-22, IL-25, TNF or IL-33 (n = 4, B6J). Data are pooled from two independent experiments. Medians are shown. One-way ANOVA. f , A-Eos frequencies after conditioning with increasing doses of IL-33. Input: BM-derived (n = 5, B6J), blood (n = 5, Il5 -tg) and splenic (n = 5, Il5 -tg) eosinophils. Medians are shown. One-way ANOVA. g , Western blot of phospho-p38 and phospho-p65 upon conditioning of BM-Eos with colon CM or IL-33 (n = 3, B6J). h , Gene expression normalized to Hprt measured by qRT–PCR of BM-Eos upon conditioning with IL-33 (n = 4, B6J). Data represents mean ± SEM. Two-tailed unpaired Student’s t -test. i , ST2 expression in BM-Eos upon IL-33 treatment (n = 4, B6J). Data represents mean ± SEM. Two-tailed unpaired Student’s t -test. j , ST2 expression in colonic A-Eos and B-Eos (n = 5, B6J). Data represents mean ± SEM. Two-tailed unpaired Student’s t -test. k , ST2 expression across organs (n = 5, B6J). Data represents mean ± SEM. One-way ANOVA. l , A-Eos frequencies upon conditioning of WT (n = 2, pooled B6J) or ST2 −/− (n = 2, pooled) BM-Eos with colon CM or IL-33. Technical replicates and mean ± SEM are shown. Two-tailed unpaired Student’s t -test. m , Left: Representative FACS plots of A-Eos (PD-L1 + CD80 + ) and PD-L1 − CD80 − eosinophils in the blood (top) and spleen (bottom). Numbers indicate % of eosinophils. Right: A-Eos frequencies in mice treated with IL-33 (n = 6-7, B6J). Medians are shown. Two-tailed unpaired Student’s t -test. n , A-Eos and B-Eos frequencies in the indicated organs of B6J (n = 7) and Il33 −/− (n = 5) mice at steady state. Medians are shown. Two-tailed unpaired Student’s t -test.

    Techniques Used: CRISPR, Inhibition, Enzyme-linked Immunosorbent Assay, Infection, Two Tailed Test, Derivative Assay, Western Blot, Expressing, Quantitative RT-PCR

    a , Regulon activity in A-Eos across conditions (n = 4, Il5 -tg). b , Multidimensional scaling (MDS) plot of bulk RNA-seq samples shown in Fig. . c , Heat map of signature gene expression across conditions of samples shown in Fig. . d , A-Eos (PD-L1 + CD80 + ) frequencies upon treatment of BM-Eos with IL-33 and/or IFNγ. (n = 4, B6J). Data represents mean ± SEM. One-way ANOVA. e , EPX immunofluorescence of A-Eos upon exposure to IFNγ for 90 min. Splenic eosinophils were magnetically enriched (n = 2, Il5 -tg), treated overnight with colon CM and A-Eos sorted by flow cytometry. Scale bar, 10 µm. f , Frequencies of PD-L1 + and CD80 + in colonic eosinophils of WT (n = 6, B6J) and Eo- Cre;Ifngr fl/fl mice (n = 4) upon C. rodentium infection, relative to uninfected controls (n = 2, B6J). Medians are shown. Two-tailed unpaired Student’s t -test. g , Left: UMAP of single-cell eosinophil transcriptomes isolated from the colon of anti-IFNγR-treated, C. rodentium -infected or control Il5 -tg mice (n = 3). Middle: expression of IFNγ target genes. Right: Expression of granule and antimicrobial signatures. Data represents mean ± SD. Two-sided Wilcoxon test. h , Observed vs. expected number of contacts between clusters of SIGLEC8 and CD4 molecules shown per slide. P Values are computed based on a two-sided permutation test (see ). i , Proportions of segmented cells expressing SIGLEC8 only (blue) or co-expressing both SIGLEC8 and CD4 (red) across slides. Dotted horizontal line shows mean. j , Mean count per slide of molecules of a given transcript in the proximity (<10 µm) of SIGLEC8 RNA molecules spatially associated with CD4 molecules vs SIGLEC8 molecules not associated with CD4 molecules. The central line in the box plot represents the median count per slide, the lower and upper hinge corresponds to the first quartiles and the whisker extends from the hinge to the smallest or largest value no further than 1.5 x IQR from the hinge. Two-sided paired Wilcoxon test (17 ROIs, n = 4 patients).
    Figure Legend Snippet: a , Regulon activity in A-Eos across conditions (n = 4, Il5 -tg). b , Multidimensional scaling (MDS) plot of bulk RNA-seq samples shown in Fig. . c , Heat map of signature gene expression across conditions of samples shown in Fig. . d , A-Eos (PD-L1 + CD80 + ) frequencies upon treatment of BM-Eos with IL-33 and/or IFNγ. (n = 4, B6J). Data represents mean ± SEM. One-way ANOVA. e , EPX immunofluorescence of A-Eos upon exposure to IFNγ for 90 min. Splenic eosinophils were magnetically enriched (n = 2, Il5 -tg), treated overnight with colon CM and A-Eos sorted by flow cytometry. Scale bar, 10 µm. f , Frequencies of PD-L1 + and CD80 + in colonic eosinophils of WT (n = 6, B6J) and Eo- Cre;Ifngr fl/fl mice (n = 4) upon C. rodentium infection, relative to uninfected controls (n = 2, B6J). Medians are shown. Two-tailed unpaired Student’s t -test. g , Left: UMAP of single-cell eosinophil transcriptomes isolated from the colon of anti-IFNγR-treated, C. rodentium -infected or control Il5 -tg mice (n = 3). Middle: expression of IFNγ target genes. Right: Expression of granule and antimicrobial signatures. Data represents mean ± SD. Two-sided Wilcoxon test. h , Observed vs. expected number of contacts between clusters of SIGLEC8 and CD4 molecules shown per slide. P Values are computed based on a two-sided permutation test (see ). i , Proportions of segmented cells expressing SIGLEC8 only (blue) or co-expressing both SIGLEC8 and CD4 (red) across slides. Dotted horizontal line shows mean. j , Mean count per slide of molecules of a given transcript in the proximity (<10 µm) of SIGLEC8 RNA molecules spatially associated with CD4 molecules vs SIGLEC8 molecules not associated with CD4 molecules. The central line in the box plot represents the median count per slide, the lower and upper hinge corresponds to the first quartiles and the whisker extends from the hinge to the smallest or largest value no further than 1.5 x IQR from the hinge. Two-sided paired Wilcoxon test (17 ROIs, n = 4 patients).

    Techniques Used: Activity Assay, RNA Sequencing Assay, Expressing, Immunofluorescence, Flow Cytometry, Infection, Two Tailed Test, Isolation, Whisker Assay

    a , Top, Venn diagram of significant differentially expressed genes (DEGs) (false discovery rate (FDR) < 0.05, logFC > |2|) in BM-Eos treated with IL-33 and/or IFNγ ( n = 4, B6J). All DEGs are listed in Supplementary Table . Bottom, expression of subset markers across conditions. Columns are clustered, rows are scaled. CPM, counts per million. b , Proliferation of anti-CD3/CD28-activated, CFSE-labelled naive CD4 + T cells co-cultured with BM-Eos conditioned with IL-33 and/or IFNγ ( n = 4, B6J). Data are pooled from two independent experiments. Medians are shown. One-way ANOVA. c , A-Eos frequencies in mice treated with IL-33 and/or IFNγ ( n = 5, B6J). Medians are shown. One-way ANOVA. d , A-Eos and B-Eos frequencies in DSS-treated B6J ( n = 5) and Il33 −/ − (n = 4) mice. Medians are shown. Two-tailed unpaired Student’s t -test. e , Frequencies of IFNγ-, IL-17- and TNF-expressing colonic CD4 + T cells from mice in d . Medians are shown. Two-tailed unpaired Student’s t -test. f , Left, representative haematoxylin and eosin (H&E)-stained colonic sections of mice in d . Scale bar, 100 µm. Right, colitis score assessed by histopathological examination. Medians are shown. Two-tailed unpaired Student’s t -test. g , Representative molecular cartography images of human ulcerative colitis samples. Nuclei are stained with DAPI; CD4 , SIGLEC8 and CD80 RNA molecules are shown in blue, red and yellow, respectively. Scale bar, 200 µm. h , Pairwise proximity score of transcripts across slides. The score indicates the fraction of slides in which the proximity of a pair of transcripts is significantly higher than expected by chance. P values are computed using a permutation test . T reg cells, regulatory T cells. i , Mean counts per slide of CD80 and NFKB1 transcripts in the proximity (<10 µm) of SIGLEC8 transcripts spatially associated with CD4 molecules versus SIGLEC8 molecules not associated with CD4 molecules. The central line in the box plot represents the median count per slide, the lower and upper hinge correspond to the first quartiles and the whisker extends from the hinge to the smallest or largest value no further than 1.5 times the interquartile range (IQR) from the hinge. Two-sided paired Wilcoxon test (17 regions of interest (ROIs), n = 4 patients).
    Figure Legend Snippet: a , Top, Venn diagram of significant differentially expressed genes (DEGs) (false discovery rate (FDR) < 0.05, logFC > |2|) in BM-Eos treated with IL-33 and/or IFNγ ( n = 4, B6J). All DEGs are listed in Supplementary Table . Bottom, expression of subset markers across conditions. Columns are clustered, rows are scaled. CPM, counts per million. b , Proliferation of anti-CD3/CD28-activated, CFSE-labelled naive CD4 + T cells co-cultured with BM-Eos conditioned with IL-33 and/or IFNγ ( n = 4, B6J). Data are pooled from two independent experiments. Medians are shown. One-way ANOVA. c , A-Eos frequencies in mice treated with IL-33 and/or IFNγ ( n = 5, B6J). Medians are shown. One-way ANOVA. d , A-Eos and B-Eos frequencies in DSS-treated B6J ( n = 5) and Il33 −/ − (n = 4) mice. Medians are shown. Two-tailed unpaired Student’s t -test. e , Frequencies of IFNγ-, IL-17- and TNF-expressing colonic CD4 + T cells from mice in d . Medians are shown. Two-tailed unpaired Student’s t -test. f , Left, representative haematoxylin and eosin (H&E)-stained colonic sections of mice in d . Scale bar, 100 µm. Right, colitis score assessed by histopathological examination. Medians are shown. Two-tailed unpaired Student’s t -test. g , Representative molecular cartography images of human ulcerative colitis samples. Nuclei are stained with DAPI; CD4 , SIGLEC8 and CD80 RNA molecules are shown in blue, red and yellow, respectively. Scale bar, 200 µm. h , Pairwise proximity score of transcripts across slides. The score indicates the fraction of slides in which the proximity of a pair of transcripts is significantly higher than expected by chance. P values are computed using a permutation test . T reg cells, regulatory T cells. i , Mean counts per slide of CD80 and NFKB1 transcripts in the proximity (<10 µm) of SIGLEC8 transcripts spatially associated with CD4 molecules versus SIGLEC8 molecules not associated with CD4 molecules. The central line in the box plot represents the median count per slide, the lower and upper hinge correspond to the first quartiles and the whisker extends from the hinge to the smallest or largest value no further than 1.5 times the interquartile range (IQR) from the hinge. Two-sided paired Wilcoxon test (17 regions of interest (ROIs), n = 4 patients).

    Techniques Used: Expressing, Cell Culture, Two Tailed Test, Staining, Whisker Assay



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    a , Activity of A-Eos-specific regulons across clusters. g, number of genes in regulon. b , Expression of NF-κB signalling components. c , Quantification of pNF-κB p65 + cells in colonic A-Eos and B-Eos ( n = 3, B6J). Data are mean ± s.e.m. Two-tailed unpaired Student’s t -test. d , e , A-Eos and B-Eos frequencies in antibiotic-treated ( d ) ( n = 16, B6J) and germ-free (GF) ( e ) ( n = 9, B6J) mice relative to controls. SPF, specific pathogen free. d , Data are pooled from two independent experiments. Medians are shown. Two-tailed unpaired Student’s t -test. f , Depleted gene sets in PD-L1 + CD80 + A-Eos (red) and PD-L1 − CD80 − eosinophils (grey), relative to BM stem cells. Kolmogorov–Smirnov test. Dot size indicates gene-set size. Dashed line indicates P = 0.05. g , A-Eos frequencies after conditioning of BM-Eos <t>with</t> <t>IL-33,</t> colon CM and anti-IL-33 ( n = 2, pooled B6J). Technical replicates and mean ± s.e.m. are shown. One-way ANOVA. h , Colonic A-Eos and B-Eos frequencies in B6J ( n = 21) and Myd88 −/− ( n = 15) mice treated with IL-33, relative to untreated controls. Medians are shown. Two-tailed unpaired Student’s t -test. WT, wild type.
    Neutralizing Monoclonal Antibody Against Mouse Il 33 Bondy 1 1, supplied by Adipogen, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    anti-il-33 neutralizing antibody  (R&D Systems)
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    PA protected against intestinal I/R injury via <t>an</t> <t>IL-33/ST2</t> signal. (A) IL-33 immunohistochemical staining in the ileum from sham, I/R and I/R + PA mice, scale bar is 100 μm (n = 8). (B, C) Relative mRNA levels of IL-33 and IL-33 receptor (ST2) (n = 8). (D) HE staining, ZO-1, Occludin and Ki67 immunofluorescent staining and IL-33 immunohistochemical staining in WT mice and IL-33 -/- mice, scale bar is 100 μm (n = 8). (E) Pathological damage score in the ileum. (F–H) The relative fluorescence intensity quantification analysis of ZO-1, Occludin and Ki67 in the ileum. (I) The relative mRNA level of Lgr5 in the ileum was measured by quantitative PCR (n = 8). (J) The relative IL-33 intensity quantification analysis in the ileum. (K, L) Intestinal lamina propria cells were analyzed by flow cytometry for the ratio of ILC2/ILCs and IL-13 + ILC2/ILC2 in WT mice (n = 3-4). (M, N) IL-13 immunohistochemical staining and intensity quantification analysis in the ileum, scale bar is 100 μm (n = 8). The results are expressed as the mean ± SEM (B, C, E–J, N) . * p < 0.05, ** p < 0.01, *** p < 0.001, NS means No statistically significant difference by one-way ANOVA (Tukey’s test). PA, pravastatin; I/R, ischemia/reperfusion; ILC2, type II innate lymphoid cells.
    Anti Il 33 Neutralizing Antibody, supplied by R&D Systems, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti-il-33 neutralizing antibody/product/R&D Systems
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    a , Activity of A-Eos-specific regulons across clusters. g, number of genes in regulon. b , Expression of NF-κB signalling components. c , Quantification of pNF-κB p65 + cells in colonic A-Eos and B-Eos ( n = 3, B6J). Data are mean ± s.e.m. Two-tailed unpaired Student’s t -test. d , e , A-Eos and B-Eos frequencies in antibiotic-treated ( d ) ( n = 16, B6J) and germ-free (GF) ( e ) ( n = 9, B6J) mice relative to controls. SPF, specific pathogen free. d , Data are pooled from two independent experiments. Medians are shown. Two-tailed unpaired Student’s t -test. f , Depleted gene sets in PD-L1 + CD80 + A-Eos (red) and PD-L1 − CD80 − eosinophils (grey), relative to BM stem cells. Kolmogorov–Smirnov test. Dot size indicates gene-set size. Dashed line indicates P = 0.05. g , A-Eos frequencies after conditioning of BM-Eos with IL-33, colon CM and anti-IL-33 ( n = 2, pooled B6J). Technical replicates and mean ± s.e.m. are shown. One-way ANOVA. h , Colonic A-Eos and B-Eos frequencies in B6J ( n = 21) and Myd88 −/− ( n = 15) mice treated with IL-33, relative to untreated controls. Medians are shown. Two-tailed unpaired Student’s t -test. WT, wild type.

    Journal: Nature

    Article Title: Active eosinophils regulate host defence and immune responses in colitis

    doi: 10.1038/s41586-022-05628-7

    Figure Lengend Snippet: a , Activity of A-Eos-specific regulons across clusters. g, number of genes in regulon. b , Expression of NF-κB signalling components. c , Quantification of pNF-κB p65 + cells in colonic A-Eos and B-Eos ( n = 3, B6J). Data are mean ± s.e.m. Two-tailed unpaired Student’s t -test. d , e , A-Eos and B-Eos frequencies in antibiotic-treated ( d ) ( n = 16, B6J) and germ-free (GF) ( e ) ( n = 9, B6J) mice relative to controls. SPF, specific pathogen free. d , Data are pooled from two independent experiments. Medians are shown. Two-tailed unpaired Student’s t -test. f , Depleted gene sets in PD-L1 + CD80 + A-Eos (red) and PD-L1 − CD80 − eosinophils (grey), relative to BM stem cells. Kolmogorov–Smirnov test. Dot size indicates gene-set size. Dashed line indicates P = 0.05. g , A-Eos frequencies after conditioning of BM-Eos with IL-33, colon CM and anti-IL-33 ( n = 2, pooled B6J). Technical replicates and mean ± s.e.m. are shown. One-way ANOVA. h , Colonic A-Eos and B-Eos frequencies in B6J ( n = 21) and Myd88 −/− ( n = 15) mice treated with IL-33, relative to untreated controls. Medians are shown. Two-tailed unpaired Student’s t -test. WT, wild type.

    Article Snippet: The NF-κB inhibitor BAY11-7082 (B5556, Sigma) was added at a concentration of 5 μM and anti-IL-33 neutralizing antibody (AF3626, Biotechne) at 30 ng ml −1 .

    Techniques: Activity Assay, Expressing, Two Tailed Test

    a , Experimental workflow of the CRISPR inhibition screen. b , Log10 negative score per gene, as calculated by MAGeCK. Cd80 and Cd274 evidenced in orange. Genes involved in TNF signalling pathway via NF-κB in red, and MAPK signalling pathway in darkred. c , IL-33 concentrations measured by ELISA in colon of DSS-treated mice (n = 12, B6J) and colon and blood of C. rodentium -infected mice (n = 7, B6J), compared to untreated controls (n = 7, B6J). Medians are shown. Two-tailed unpaired Student’s t -test. d , IFNγ, TNF and IL-22 concentrations measured by LEGENDplex in colon (left) and blood (right) of C. rod entium-infected mice (n = 7, B6J), compared to untreated controls (n = 7, B6J). Medians are shown. Two-tailed unpaired Student’s t -test. e , A-Eos (PD-L1 + CD80 + ) frequencies upon conditioning of BM-Eos with colon CM, IL-22, IL-25, TNF or IL-33 (n = 4, B6J). Data are pooled from two independent experiments. Medians are shown. One-way ANOVA. f , A-Eos frequencies after conditioning with increasing doses of IL-33. Input: BM-derived (n = 5, B6J), blood (n = 5, Il5 -tg) and splenic (n = 5, Il5 -tg) eosinophils. Medians are shown. One-way ANOVA. g , Western blot of phospho-p38 and phospho-p65 upon conditioning of BM-Eos with colon CM or IL-33 (n = 3, B6J). h , Gene expression normalized to Hprt measured by qRT–PCR of BM-Eos upon conditioning with IL-33 (n = 4, B6J). Data represents mean ± SEM. Two-tailed unpaired Student’s t -test. i , ST2 expression in BM-Eos upon IL-33 treatment (n = 4, B6J). Data represents mean ± SEM. Two-tailed unpaired Student’s t -test. j , ST2 expression in colonic A-Eos and B-Eos (n = 5, B6J). Data represents mean ± SEM. Two-tailed unpaired Student’s t -test. k , ST2 expression across organs (n = 5, B6J). Data represents mean ± SEM. One-way ANOVA. l , A-Eos frequencies upon conditioning of WT (n = 2, pooled B6J) or ST2 −/− (n = 2, pooled) BM-Eos with colon CM or IL-33. Technical replicates and mean ± SEM are shown. Two-tailed unpaired Student’s t -test. m , Left: Representative FACS plots of A-Eos (PD-L1 + CD80 + ) and PD-L1 − CD80 − eosinophils in the blood (top) and spleen (bottom). Numbers indicate % of eosinophils. Right: A-Eos frequencies in mice treated with IL-33 (n = 6-7, B6J). Medians are shown. Two-tailed unpaired Student’s t -test. n , A-Eos and B-Eos frequencies in the indicated organs of B6J (n = 7) and Il33 −/− (n = 5) mice at steady state. Medians are shown. Two-tailed unpaired Student’s t -test.

    Journal: Nature

    Article Title: Active eosinophils regulate host defence and immune responses in colitis

    doi: 10.1038/s41586-022-05628-7

    Figure Lengend Snippet: a , Experimental workflow of the CRISPR inhibition screen. b , Log10 negative score per gene, as calculated by MAGeCK. Cd80 and Cd274 evidenced in orange. Genes involved in TNF signalling pathway via NF-κB in red, and MAPK signalling pathway in darkred. c , IL-33 concentrations measured by ELISA in colon of DSS-treated mice (n = 12, B6J) and colon and blood of C. rodentium -infected mice (n = 7, B6J), compared to untreated controls (n = 7, B6J). Medians are shown. Two-tailed unpaired Student’s t -test. d , IFNγ, TNF and IL-22 concentrations measured by LEGENDplex in colon (left) and blood (right) of C. rod entium-infected mice (n = 7, B6J), compared to untreated controls (n = 7, B6J). Medians are shown. Two-tailed unpaired Student’s t -test. e , A-Eos (PD-L1 + CD80 + ) frequencies upon conditioning of BM-Eos with colon CM, IL-22, IL-25, TNF or IL-33 (n = 4, B6J). Data are pooled from two independent experiments. Medians are shown. One-way ANOVA. f , A-Eos frequencies after conditioning with increasing doses of IL-33. Input: BM-derived (n = 5, B6J), blood (n = 5, Il5 -tg) and splenic (n = 5, Il5 -tg) eosinophils. Medians are shown. One-way ANOVA. g , Western blot of phospho-p38 and phospho-p65 upon conditioning of BM-Eos with colon CM or IL-33 (n = 3, B6J). h , Gene expression normalized to Hprt measured by qRT–PCR of BM-Eos upon conditioning with IL-33 (n = 4, B6J). Data represents mean ± SEM. Two-tailed unpaired Student’s t -test. i , ST2 expression in BM-Eos upon IL-33 treatment (n = 4, B6J). Data represents mean ± SEM. Two-tailed unpaired Student’s t -test. j , ST2 expression in colonic A-Eos and B-Eos (n = 5, B6J). Data represents mean ± SEM. Two-tailed unpaired Student’s t -test. k , ST2 expression across organs (n = 5, B6J). Data represents mean ± SEM. One-way ANOVA. l , A-Eos frequencies upon conditioning of WT (n = 2, pooled B6J) or ST2 −/− (n = 2, pooled) BM-Eos with colon CM or IL-33. Technical replicates and mean ± SEM are shown. Two-tailed unpaired Student’s t -test. m , Left: Representative FACS plots of A-Eos (PD-L1 + CD80 + ) and PD-L1 − CD80 − eosinophils in the blood (top) and spleen (bottom). Numbers indicate % of eosinophils. Right: A-Eos frequencies in mice treated with IL-33 (n = 6-7, B6J). Medians are shown. Two-tailed unpaired Student’s t -test. n , A-Eos and B-Eos frequencies in the indicated organs of B6J (n = 7) and Il33 −/− (n = 5) mice at steady state. Medians are shown. Two-tailed unpaired Student’s t -test.

    Article Snippet: The NF-κB inhibitor BAY11-7082 (B5556, Sigma) was added at a concentration of 5 μM and anti-IL-33 neutralizing antibody (AF3626, Biotechne) at 30 ng ml −1 .

    Techniques: CRISPR, Inhibition, Enzyme-linked Immunosorbent Assay, Infection, Two Tailed Test, Derivative Assay, Western Blot, Expressing, Quantitative RT-PCR

    a , Regulon activity in A-Eos across conditions (n = 4, Il5 -tg). b , Multidimensional scaling (MDS) plot of bulk RNA-seq samples shown in Fig. . c , Heat map of signature gene expression across conditions of samples shown in Fig. . d , A-Eos (PD-L1 + CD80 + ) frequencies upon treatment of BM-Eos with IL-33 and/or IFNγ. (n = 4, B6J). Data represents mean ± SEM. One-way ANOVA. e , EPX immunofluorescence of A-Eos upon exposure to IFNγ for 90 min. Splenic eosinophils were magnetically enriched (n = 2, Il5 -tg), treated overnight with colon CM and A-Eos sorted by flow cytometry. Scale bar, 10 µm. f , Frequencies of PD-L1 + and CD80 + in colonic eosinophils of WT (n = 6, B6J) and Eo- Cre;Ifngr fl/fl mice (n = 4) upon C. rodentium infection, relative to uninfected controls (n = 2, B6J). Medians are shown. Two-tailed unpaired Student’s t -test. g , Left: UMAP of single-cell eosinophil transcriptomes isolated from the colon of anti-IFNγR-treated, C. rodentium -infected or control Il5 -tg mice (n = 3). Middle: expression of IFNγ target genes. Right: Expression of granule and antimicrobial signatures. Data represents mean ± SD. Two-sided Wilcoxon test. h , Observed vs. expected number of contacts between clusters of SIGLEC8 and CD4 molecules shown per slide. P Values are computed based on a two-sided permutation test (see ). i , Proportions of segmented cells expressing SIGLEC8 only (blue) or co-expressing both SIGLEC8 and CD4 (red) across slides. Dotted horizontal line shows mean. j , Mean count per slide of molecules of a given transcript in the proximity (<10 µm) of SIGLEC8 RNA molecules spatially associated with CD4 molecules vs SIGLEC8 molecules not associated with CD4 molecules. The central line in the box plot represents the median count per slide, the lower and upper hinge corresponds to the first quartiles and the whisker extends from the hinge to the smallest or largest value no further than 1.5 x IQR from the hinge. Two-sided paired Wilcoxon test (17 ROIs, n = 4 patients).

    Journal: Nature

    Article Title: Active eosinophils regulate host defence and immune responses in colitis

    doi: 10.1038/s41586-022-05628-7

    Figure Lengend Snippet: a , Regulon activity in A-Eos across conditions (n = 4, Il5 -tg). b , Multidimensional scaling (MDS) plot of bulk RNA-seq samples shown in Fig. . c , Heat map of signature gene expression across conditions of samples shown in Fig. . d , A-Eos (PD-L1 + CD80 + ) frequencies upon treatment of BM-Eos with IL-33 and/or IFNγ. (n = 4, B6J). Data represents mean ± SEM. One-way ANOVA. e , EPX immunofluorescence of A-Eos upon exposure to IFNγ for 90 min. Splenic eosinophils were magnetically enriched (n = 2, Il5 -tg), treated overnight with colon CM and A-Eos sorted by flow cytometry. Scale bar, 10 µm. f , Frequencies of PD-L1 + and CD80 + in colonic eosinophils of WT (n = 6, B6J) and Eo- Cre;Ifngr fl/fl mice (n = 4) upon C. rodentium infection, relative to uninfected controls (n = 2, B6J). Medians are shown. Two-tailed unpaired Student’s t -test. g , Left: UMAP of single-cell eosinophil transcriptomes isolated from the colon of anti-IFNγR-treated, C. rodentium -infected or control Il5 -tg mice (n = 3). Middle: expression of IFNγ target genes. Right: Expression of granule and antimicrobial signatures. Data represents mean ± SD. Two-sided Wilcoxon test. h , Observed vs. expected number of contacts between clusters of SIGLEC8 and CD4 molecules shown per slide. P Values are computed based on a two-sided permutation test (see ). i , Proportions of segmented cells expressing SIGLEC8 only (blue) or co-expressing both SIGLEC8 and CD4 (red) across slides. Dotted horizontal line shows mean. j , Mean count per slide of molecules of a given transcript in the proximity (<10 µm) of SIGLEC8 RNA molecules spatially associated with CD4 molecules vs SIGLEC8 molecules not associated with CD4 molecules. The central line in the box plot represents the median count per slide, the lower and upper hinge corresponds to the first quartiles and the whisker extends from the hinge to the smallest or largest value no further than 1.5 x IQR from the hinge. Two-sided paired Wilcoxon test (17 ROIs, n = 4 patients).

    Article Snippet: The NF-κB inhibitor BAY11-7082 (B5556, Sigma) was added at a concentration of 5 μM and anti-IL-33 neutralizing antibody (AF3626, Biotechne) at 30 ng ml −1 .

    Techniques: Activity Assay, RNA Sequencing Assay, Expressing, Immunofluorescence, Flow Cytometry, Infection, Two Tailed Test, Isolation, Whisker Assay

    a , Top, Venn diagram of significant differentially expressed genes (DEGs) (false discovery rate (FDR) < 0.05, logFC > |2|) in BM-Eos treated with IL-33 and/or IFNγ ( n = 4, B6J). All DEGs are listed in Supplementary Table . Bottom, expression of subset markers across conditions. Columns are clustered, rows are scaled. CPM, counts per million. b , Proliferation of anti-CD3/CD28-activated, CFSE-labelled naive CD4 + T cells co-cultured with BM-Eos conditioned with IL-33 and/or IFNγ ( n = 4, B6J). Data are pooled from two independent experiments. Medians are shown. One-way ANOVA. c , A-Eos frequencies in mice treated with IL-33 and/or IFNγ ( n = 5, B6J). Medians are shown. One-way ANOVA. d , A-Eos and B-Eos frequencies in DSS-treated B6J ( n = 5) and Il33 −/ − (n = 4) mice. Medians are shown. Two-tailed unpaired Student’s t -test. e , Frequencies of IFNγ-, IL-17- and TNF-expressing colonic CD4 + T cells from mice in d . Medians are shown. Two-tailed unpaired Student’s t -test. f , Left, representative haematoxylin and eosin (H&E)-stained colonic sections of mice in d . Scale bar, 100 µm. Right, colitis score assessed by histopathological examination. Medians are shown. Two-tailed unpaired Student’s t -test. g , Representative molecular cartography images of human ulcerative colitis samples. Nuclei are stained with DAPI; CD4 , SIGLEC8 and CD80 RNA molecules are shown in blue, red and yellow, respectively. Scale bar, 200 µm. h , Pairwise proximity score of transcripts across slides. The score indicates the fraction of slides in which the proximity of a pair of transcripts is significantly higher than expected by chance. P values are computed using a permutation test . T reg cells, regulatory T cells. i , Mean counts per slide of CD80 and NFKB1 transcripts in the proximity (<10 µm) of SIGLEC8 transcripts spatially associated with CD4 molecules versus SIGLEC8 molecules not associated with CD4 molecules. The central line in the box plot represents the median count per slide, the lower and upper hinge correspond to the first quartiles and the whisker extends from the hinge to the smallest or largest value no further than 1.5 times the interquartile range (IQR) from the hinge. Two-sided paired Wilcoxon test (17 regions of interest (ROIs), n = 4 patients).

    Journal: Nature

    Article Title: Active eosinophils regulate host defence and immune responses in colitis

    doi: 10.1038/s41586-022-05628-7

    Figure Lengend Snippet: a , Top, Venn diagram of significant differentially expressed genes (DEGs) (false discovery rate (FDR) < 0.05, logFC > |2|) in BM-Eos treated with IL-33 and/or IFNγ ( n = 4, B6J). All DEGs are listed in Supplementary Table . Bottom, expression of subset markers across conditions. Columns are clustered, rows are scaled. CPM, counts per million. b , Proliferation of anti-CD3/CD28-activated, CFSE-labelled naive CD4 + T cells co-cultured with BM-Eos conditioned with IL-33 and/or IFNγ ( n = 4, B6J). Data are pooled from two independent experiments. Medians are shown. One-way ANOVA. c , A-Eos frequencies in mice treated with IL-33 and/or IFNγ ( n = 5, B6J). Medians are shown. One-way ANOVA. d , A-Eos and B-Eos frequencies in DSS-treated B6J ( n = 5) and Il33 −/ − (n = 4) mice. Medians are shown. Two-tailed unpaired Student’s t -test. e , Frequencies of IFNγ-, IL-17- and TNF-expressing colonic CD4 + T cells from mice in d . Medians are shown. Two-tailed unpaired Student’s t -test. f , Left, representative haematoxylin and eosin (H&E)-stained colonic sections of mice in d . Scale bar, 100 µm. Right, colitis score assessed by histopathological examination. Medians are shown. Two-tailed unpaired Student’s t -test. g , Representative molecular cartography images of human ulcerative colitis samples. Nuclei are stained with DAPI; CD4 , SIGLEC8 and CD80 RNA molecules are shown in blue, red and yellow, respectively. Scale bar, 200 µm. h , Pairwise proximity score of transcripts across slides. The score indicates the fraction of slides in which the proximity of a pair of transcripts is significantly higher than expected by chance. P values are computed using a permutation test . T reg cells, regulatory T cells. i , Mean counts per slide of CD80 and NFKB1 transcripts in the proximity (<10 µm) of SIGLEC8 transcripts spatially associated with CD4 molecules versus SIGLEC8 molecules not associated with CD4 molecules. The central line in the box plot represents the median count per slide, the lower and upper hinge correspond to the first quartiles and the whisker extends from the hinge to the smallest or largest value no further than 1.5 times the interquartile range (IQR) from the hinge. Two-sided paired Wilcoxon test (17 regions of interest (ROIs), n = 4 patients).

    Article Snippet: The NF-κB inhibitor BAY11-7082 (B5556, Sigma) was added at a concentration of 5 μM and anti-IL-33 neutralizing antibody (AF3626, Biotechne) at 30 ng ml −1 .

    Techniques: Expressing, Cell Culture, Two Tailed Test, Staining, Whisker Assay

    PA protected against intestinal I/R injury via an IL-33/ST2 signal. (A) IL-33 immunohistochemical staining in the ileum from sham, I/R and I/R + PA mice, scale bar is 100 μm (n = 8). (B, C) Relative mRNA levels of IL-33 and IL-33 receptor (ST2) (n = 8). (D) HE staining, ZO-1, Occludin and Ki67 immunofluorescent staining and IL-33 immunohistochemical staining in WT mice and IL-33 -/- mice, scale bar is 100 μm (n = 8). (E) Pathological damage score in the ileum. (F–H) The relative fluorescence intensity quantification analysis of ZO-1, Occludin and Ki67 in the ileum. (I) The relative mRNA level of Lgr5 in the ileum was measured by quantitative PCR (n = 8). (J) The relative IL-33 intensity quantification analysis in the ileum. (K, L) Intestinal lamina propria cells were analyzed by flow cytometry for the ratio of ILC2/ILCs and IL-13 + ILC2/ILC2 in WT mice (n = 3-4). (M, N) IL-13 immunohistochemical staining and intensity quantification analysis in the ileum, scale bar is 100 μm (n = 8). The results are expressed as the mean ± SEM (B, C, E–J, N) . * p < 0.05, ** p < 0.01, *** p < 0.001, NS means No statistically significant difference by one-way ANOVA (Tukey’s test). PA, pravastatin; I/R, ischemia/reperfusion; ILC2, type II innate lymphoid cells.

    Journal: Frontiers in Immunology

    Article Title: Gut Microbial Metabolite Pravastatin Attenuates Intestinal Ischemia/Reperfusion Injury Through Promoting IL-13 Release From Type II Innate Lymphoid Cells via IL−33/ST2 Signaling

    doi: 10.3389/fimmu.2021.704836

    Figure Lengend Snippet: PA protected against intestinal I/R injury via an IL-33/ST2 signal. (A) IL-33 immunohistochemical staining in the ileum from sham, I/R and I/R + PA mice, scale bar is 100 μm (n = 8). (B, C) Relative mRNA levels of IL-33 and IL-33 receptor (ST2) (n = 8). (D) HE staining, ZO-1, Occludin and Ki67 immunofluorescent staining and IL-33 immunohistochemical staining in WT mice and IL-33 -/- mice, scale bar is 100 μm (n = 8). (E) Pathological damage score in the ileum. (F–H) The relative fluorescence intensity quantification analysis of ZO-1, Occludin and Ki67 in the ileum. (I) The relative mRNA level of Lgr5 in the ileum was measured by quantitative PCR (n = 8). (J) The relative IL-33 intensity quantification analysis in the ileum. (K, L) Intestinal lamina propria cells were analyzed by flow cytometry for the ratio of ILC2/ILCs and IL-13 + ILC2/ILC2 in WT mice (n = 3-4). (M, N) IL-13 immunohistochemical staining and intensity quantification analysis in the ileum, scale bar is 100 μm (n = 8). The results are expressed as the mean ± SEM (B, C, E–J, N) . * p < 0.05, ** p < 0.01, *** p < 0.001, NS means No statistically significant difference by one-way ANOVA (Tukey’s test). PA, pravastatin; I/R, ischemia/reperfusion; ILC2, type II innate lymphoid cells.

    Article Snippet: To explore the role of IL-33/ST2 signaling in the protective effect of PA against intestinal I/R injury, wild type (WT) mice were randomly divided into the following groups ( ): I/R; ( ) I/R + PA; ( ) I/R + Anti-IL-33, in which WT mice were injected i.p. with 60 μg/kg Anti-IL-33 neutralizing antibody (R&D Systems, Inc., Minneapolis, USA) 2 h before establishing the intestinal I/R model; ( ) I/R + PA + Anti-IL-33, in which WT mice were injected i.p. with 60 μg/kg Anti-IL-33 neutralizing antibody 2 h before intestinal I/R and 2 mg/kg PA 1 h before intestinal I/R in mice; ( ) I/R + PA + Anti-ST2, in which WT mice were injected i.p. with 2 mg/kg PA and 1.5 mg/kg Anti-ST2 neutralizing antibody (R&D Systems, Inc.) 1 and 2 h, respectively, before establishing the intestinal I/R model in mice.

    Techniques: Immunohistochemical staining, Staining, Fluorescence, Real-time Polymerase Chain Reaction, Flow Cytometry

    PA protected organoids H/R injury via an IL-33/ST2 signal. (A) The relative mRNA and protein level of IL-33 in the organoid from WT-Organoids + ILC2. (B) HE staining, ZO-1, Occludin and Ki67 immunofluorescent staining in the organoid from WT-Organoids + ILC2, scale bar is 20 μm (n = 6). (C, D) Relative organoids viability and LDH levels were detected in a co-culture system of organoids and ILC2 after H/R (n = 6). (E, F) The relative fluorescence intensity quantification analysis of ZO-1, Occludin and Ki67 in the organoids (n = 6). (G) Relative mRNA level of Lgr5 in the organoids was measured by quantitative PCR (n = 6). (H) The relative protein level of IL-13 in the organoid supernatant determined by ELISA (n = 6). The results are expressed as the mean ± SEM (A, C–H) . * p < 0.05, ** p < 0.01, *** p < 0.001, NS means No statistically significant difference by one-way ANOVA (Tukey’s test). PA, pravastatin; H/R, hypoxia/reoxygenation; ILC2, type II innate lymphoid cells; LDH, lactate dehydrogenase; WT-Organoids + ILC2, co-cultured WT-organoids and WT-ILC2; IL-33 -/– Organoids + ILC2, co-cultured IL-33 -/- organoids and WT-ILC2.

    Journal: Frontiers in Immunology

    Article Title: Gut Microbial Metabolite Pravastatin Attenuates Intestinal Ischemia/Reperfusion Injury Through Promoting IL-13 Release From Type II Innate Lymphoid Cells via IL−33/ST2 Signaling

    doi: 10.3389/fimmu.2021.704836

    Figure Lengend Snippet: PA protected organoids H/R injury via an IL-33/ST2 signal. (A) The relative mRNA and protein level of IL-33 in the organoid from WT-Organoids + ILC2. (B) HE staining, ZO-1, Occludin and Ki67 immunofluorescent staining in the organoid from WT-Organoids + ILC2, scale bar is 20 μm (n = 6). (C, D) Relative organoids viability and LDH levels were detected in a co-culture system of organoids and ILC2 after H/R (n = 6). (E, F) The relative fluorescence intensity quantification analysis of ZO-1, Occludin and Ki67 in the organoids (n = 6). (G) Relative mRNA level of Lgr5 in the organoids was measured by quantitative PCR (n = 6). (H) The relative protein level of IL-13 in the organoid supernatant determined by ELISA (n = 6). The results are expressed as the mean ± SEM (A, C–H) . * p < 0.05, ** p < 0.01, *** p < 0.001, NS means No statistically significant difference by one-way ANOVA (Tukey’s test). PA, pravastatin; H/R, hypoxia/reoxygenation; ILC2, type II innate lymphoid cells; LDH, lactate dehydrogenase; WT-Organoids + ILC2, co-cultured WT-organoids and WT-ILC2; IL-33 -/– Organoids + ILC2, co-cultured IL-33 -/- organoids and WT-ILC2.

    Article Snippet: To explore the role of IL-33/ST2 signaling in the protective effect of PA against intestinal I/R injury, wild type (WT) mice were randomly divided into the following groups ( ): I/R; ( ) I/R + PA; ( ) I/R + Anti-IL-33, in which WT mice were injected i.p. with 60 μg/kg Anti-IL-33 neutralizing antibody (R&D Systems, Inc., Minneapolis, USA) 2 h before establishing the intestinal I/R model; ( ) I/R + PA + Anti-IL-33, in which WT mice were injected i.p. with 60 μg/kg Anti-IL-33 neutralizing antibody 2 h before intestinal I/R and 2 mg/kg PA 1 h before intestinal I/R in mice; ( ) I/R + PA + Anti-ST2, in which WT mice were injected i.p. with 2 mg/kg PA and 1.5 mg/kg Anti-ST2 neutralizing antibody (R&D Systems, Inc.) 1 and 2 h, respectively, before establishing the intestinal I/R model in mice.

    Techniques: Staining, Co-Culture Assay, Fluorescence, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, Cell Culture

    Depletion of ILC2 abolished the protective effect of IL-33 on intestinal I/R injury and organoids H/R injury. (A–C) Intestinal lamina propria cells were analyzed by flow cytometry for the ratio of ILC2/ILCs and IL-13 + ILC2/ILC2 in Rag1 -/- mice (n = 3-4). (D) HE staining, ZO-1, Occludin and Ki67 immunofluorescent staining in Rag1 -/- mice, scale bar is 100 μm (n = 8). (E–G) Pathological damage score and the relative fluorescence intensity quantification analysis of ZO-1, Occludin and Ki67 in the ileum (n = 8). (H) The relative mRNA level of Lgr5 in the ileum was measured by quantitative PCR (n = 8). (I) HE staining, ZO-1, Occludin and Ki67 immunofluorescent staining in WT-organoid cultured with and without ILC2, scale bar is 20 μm (n = 6). (J) Relative organoids viability and LDH levels were detected in WT-organoid cultured with and without ILC2 after H/R (n = 6). (K–M) The relative fluorescence intensity quantification analysis of ZO-1, Occludin and Ki67 in the organoids (n = 6). (N) Relative mRNA level of Lgr5 in the organoids was measured by quantitative PCR (n = 6). The results are expressed as the mean ± SEM (E–H, J–N) . * p < 0.05, ** p < 0.01, *** p < 0.001 by one-way ANOVA (Tukey’s test) (B–H) . * p < 0.05, ** p < 0.01, *** p < 0.001, NS means No statistically significant difference by two-tailed Student’s t test (J–N) . PA, Pravastatin; I/R, ischemia/reperfusion; H/R, hypoxia/reoxygenation; ILC2, type II innate lymphoid cells; LDH, lactate dehydrogenase.

    Journal: Frontiers in Immunology

    Article Title: Gut Microbial Metabolite Pravastatin Attenuates Intestinal Ischemia/Reperfusion Injury Through Promoting IL-13 Release From Type II Innate Lymphoid Cells via IL−33/ST2 Signaling

    doi: 10.3389/fimmu.2021.704836

    Figure Lengend Snippet: Depletion of ILC2 abolished the protective effect of IL-33 on intestinal I/R injury and organoids H/R injury. (A–C) Intestinal lamina propria cells were analyzed by flow cytometry for the ratio of ILC2/ILCs and IL-13 + ILC2/ILC2 in Rag1 -/- mice (n = 3-4). (D) HE staining, ZO-1, Occludin and Ki67 immunofluorescent staining in Rag1 -/- mice, scale bar is 100 μm (n = 8). (E–G) Pathological damage score and the relative fluorescence intensity quantification analysis of ZO-1, Occludin and Ki67 in the ileum (n = 8). (H) The relative mRNA level of Lgr5 in the ileum was measured by quantitative PCR (n = 8). (I) HE staining, ZO-1, Occludin and Ki67 immunofluorescent staining in WT-organoid cultured with and without ILC2, scale bar is 20 μm (n = 6). (J) Relative organoids viability and LDH levels were detected in WT-organoid cultured with and without ILC2 after H/R (n = 6). (K–M) The relative fluorescence intensity quantification analysis of ZO-1, Occludin and Ki67 in the organoids (n = 6). (N) Relative mRNA level of Lgr5 in the organoids was measured by quantitative PCR (n = 6). The results are expressed as the mean ± SEM (E–H, J–N) . * p < 0.05, ** p < 0.01, *** p < 0.001 by one-way ANOVA (Tukey’s test) (B–H) . * p < 0.05, ** p < 0.01, *** p < 0.001, NS means No statistically significant difference by two-tailed Student’s t test (J–N) . PA, Pravastatin; I/R, ischemia/reperfusion; H/R, hypoxia/reoxygenation; ILC2, type II innate lymphoid cells; LDH, lactate dehydrogenase.

    Article Snippet: To explore the role of IL-33/ST2 signaling in the protective effect of PA against intestinal I/R injury, wild type (WT) mice were randomly divided into the following groups ( ): I/R; ( ) I/R + PA; ( ) I/R + Anti-IL-33, in which WT mice were injected i.p. with 60 μg/kg Anti-IL-33 neutralizing antibody (R&D Systems, Inc., Minneapolis, USA) 2 h before establishing the intestinal I/R model; ( ) I/R + PA + Anti-IL-33, in which WT mice were injected i.p. with 60 μg/kg Anti-IL-33 neutralizing antibody 2 h before intestinal I/R and 2 mg/kg PA 1 h before intestinal I/R in mice; ( ) I/R + PA + Anti-ST2, in which WT mice were injected i.p. with 2 mg/kg PA and 1.5 mg/kg Anti-ST2 neutralizing antibody (R&D Systems, Inc.) 1 and 2 h, respectively, before establishing the intestinal I/R model in mice.

    Techniques: Flow Cytometry, Staining, Fluorescence, Real-time Polymerase Chain Reaction, Cell Culture, Two Tailed Test

    The protection of PA/IL-33 to alleviate intestinal I/R injury was mediated by IL-13 released from ILC2. (A) IL-13 immunohistochemical staining and intensity quantification analysis in the ileum, scale bar is 100 μm (n = 8). (B) The relative protein level of IL-13 in the organoid supernatant determined by ELISA in WT-organoid cultured with and without ILC2 (n = 6). (C) HE staining, ZO-1, Occludin and Ki67 immunofluorescent staining of ileum in WT mice, scale bar is 100 μm (n = 8). (D–F) Pathological damage score and the relative fluorescence intensity quantification analysis of ZO-1, Occludin and Ki67 in the ileum (n = 8). (G) The relative mRNA level of Lgr5 in the ileum was measured by quantitative PCR (n = 8). (H) Changes in survival rate have been shown. Mice were submitted to ischemia of SMA for 60 min, and survival was monitored on day 7 after reperfusion (n = 20). (I) HE staining, ZO-1, Occludin and Ki67 immunofluorescent staining in a co-culture system of organoids and ILC2 after H/R, scale bar is 20 μm (n = 6). (J) Relative organoids viability and LDH levels were detected in a co-culture system of organoids and ILC2 after H/R (n = 6). (K–M) The relative fluorescence intensity quantification analysis of ZO-1, Occludin and Ki67 in the organoids (n = 6). (N) Relative mRNA level of Lgr5 in the organoids was measured by quantitative PCR (n = 6). The results are expressed as the mean ± SEM (B, D–G, J–N) . * p < 0.05, ** p < 0.01, *** p < 0.001, NS means No statistically significant difference by one-way ANOVA (Tukey’s test) and Log-Rank test (H) . PA, pravastatin; I/R, ischemia/reperfusion; H/R, hypoxia/reoxygenation; ILC2, type II innate lymphoid cells; SMA, superior mesenteric artery; LDH, lactate dehydrogenase.

    Journal: Frontiers in Immunology

    Article Title: Gut Microbial Metabolite Pravastatin Attenuates Intestinal Ischemia/Reperfusion Injury Through Promoting IL-13 Release From Type II Innate Lymphoid Cells via IL−33/ST2 Signaling

    doi: 10.3389/fimmu.2021.704836

    Figure Lengend Snippet: The protection of PA/IL-33 to alleviate intestinal I/R injury was mediated by IL-13 released from ILC2. (A) IL-13 immunohistochemical staining and intensity quantification analysis in the ileum, scale bar is 100 μm (n = 8). (B) The relative protein level of IL-13 in the organoid supernatant determined by ELISA in WT-organoid cultured with and without ILC2 (n = 6). (C) HE staining, ZO-1, Occludin and Ki67 immunofluorescent staining of ileum in WT mice, scale bar is 100 μm (n = 8). (D–F) Pathological damage score and the relative fluorescence intensity quantification analysis of ZO-1, Occludin and Ki67 in the ileum (n = 8). (G) The relative mRNA level of Lgr5 in the ileum was measured by quantitative PCR (n = 8). (H) Changes in survival rate have been shown. Mice were submitted to ischemia of SMA for 60 min, and survival was monitored on day 7 after reperfusion (n = 20). (I) HE staining, ZO-1, Occludin and Ki67 immunofluorescent staining in a co-culture system of organoids and ILC2 after H/R, scale bar is 20 μm (n = 6). (J) Relative organoids viability and LDH levels were detected in a co-culture system of organoids and ILC2 after H/R (n = 6). (K–M) The relative fluorescence intensity quantification analysis of ZO-1, Occludin and Ki67 in the organoids (n = 6). (N) Relative mRNA level of Lgr5 in the organoids was measured by quantitative PCR (n = 6). The results are expressed as the mean ± SEM (B, D–G, J–N) . * p < 0.05, ** p < 0.01, *** p < 0.001, NS means No statistically significant difference by one-way ANOVA (Tukey’s test) and Log-Rank test (H) . PA, pravastatin; I/R, ischemia/reperfusion; H/R, hypoxia/reoxygenation; ILC2, type II innate lymphoid cells; SMA, superior mesenteric artery; LDH, lactate dehydrogenase.

    Article Snippet: To explore the role of IL-33/ST2 signaling in the protective effect of PA against intestinal I/R injury, wild type (WT) mice were randomly divided into the following groups ( ): I/R; ( ) I/R + PA; ( ) I/R + Anti-IL-33, in which WT mice were injected i.p. with 60 μg/kg Anti-IL-33 neutralizing antibody (R&D Systems, Inc., Minneapolis, USA) 2 h before establishing the intestinal I/R model; ( ) I/R + PA + Anti-IL-33, in which WT mice were injected i.p. with 60 μg/kg Anti-IL-33 neutralizing antibody 2 h before intestinal I/R and 2 mg/kg PA 1 h before intestinal I/R in mice; ( ) I/R + PA + Anti-ST2, in which WT mice were injected i.p. with 2 mg/kg PA and 1.5 mg/kg Anti-ST2 neutralizing antibody (R&D Systems, Inc.) 1 and 2 h, respectively, before establishing the intestinal I/R model in mice.

    Techniques: Immunohistochemical staining, Staining, Enzyme-linked Immunosorbent Assay, Cell Culture, Fluorescence, Real-time Polymerase Chain Reaction, Co-Culture Assay