Mouse Angiopoietin-like Protein 2/ANGPTL2 Antibody

Detection of Angiopoietin-like 2 by Immunohistochemistry

ANGPTL2 is necessary for fibrotic vasculopathy.(A) qPCR of Angptl2 mRNA in WT and Snail-tg mouse keratinocytes (mKT). (B) Secreted ANGPTL2 (in green) in WT and Snail-tg skin at P3. The dotted line denotes the basement membrane separating the epidermis (epi) from the underlying dermis (der). (C) ELISA for secreted ANGPTL2 in WT and Snail-tg mKT conditioned media (CM). (D) Expression of Angptl2 in keloid lesion skin (n = 3) compared with nonlesion skin (n = 3) (from GSE92566) and in healthy (n = 44) compared with affected forearm skin from diffuse (n = 180) and localized SSc (n = 115) patients (from GSE181549). The expression values are from the GEO2R algorithm’s output. (E) Secreted ANGPTL2 (in green) in normal and SSc skin. Representative of n = 5 SSc samples and normal skin controls. (F) qPCR of vasculopathy and endoMT-related genes in SVEC4-10 endothelial cells treated with WT, Snail-tg, and Snail-tg/Angptl2-KO mKT conditioned media. (G) Quantification of collagen contraction assay in SVEC4-10 endothelial cells treated with WT, Snail-tg, and Snail-tg/Angptl2-KO mKT conditioned media. Nuclei are marked in blue. Scale bars: 50 μm. Data are shown as mean ± SEM; P values were calculated using paired Student’s t test (A), unpaired Welch’s t test (D), and 1-way ANOVA followed by Tukey’s post hoc analysis (F and G). *P < 0.05, **P < 0.01, ****P < 0.0001, NS > 0.05. All experiments except (D and E) are n = 3 biological replicates. Image collected and cropped by CiteAb from the following open publication (https://www.jci.org/articles/view/177123), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Angiopoietin-like 2 by Flow Cytometry

Human MAG is a new receptor for ANGPTL2. A Flow cytometric analysis of conditioned medium with/without ANGPTL2-Flag binding to 293T cells with transient transfection of plasmids of N1-hMAG-EGFP (human MAG, full-length) or N1-EGFP (vector). Percentage of binding/unbinding cells to ANGPTL2 in EGFP+ cells are shown. B Quantitative data of percentage of binding cells in Panel A (n = 3). C ANGPLT2 bound to the ECD of human MAG (human MAG-ECD-FC) in the conditioned medium (CM) of cotransfected 293T cells using co-immunoprecipitation. Tie-2-ECD-FC served as a negative control. D Representative flow cytometric analysis showing that purified ANGPTL2-Flag protein enhanced GFP expression in human MAG chimeric reporter cells (Human MAG). Control reporter cells (control) without human MAG-ECD were examined. E Quantitative data in Panel D (n = 3). F Binding kinetics of ANGPTL2-Flag to human MAG-ECD were measured using a biolayer interferometry instrument (Octed RED 96/96S). G–H Flow cytometric plots for the binding of ANGPTL2-Flag to full-length, mutant IgG 3 domain or IgG 4 domain of human MAG expressed 293T cells. Percentage of binding/unbinding cells to ANGPTL2 in EGFP+ cells (G) and quantitative data are shown (H) (n = 3). (***p < 0.001) Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/36855057), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Angiopoietin-like 2 by Flow Cytometry

Murine homologue of human MAG binds to ANGPTL2. A Flow cytometric analysis of conditioned medium with/without ANGPTL2-Flag binding to 293 T cells with transient transfection of plasmids of N1-mMAG-EGFP (murine MAG, full-length) or N1-EGFP (vector). Percentage of binding/unbinding cells to ANGPTL2 in EGFP+ cells are shown. B Quantitative data of percentage of binding cells in Panel A (n = 3). C ANGPLT2 bound to the ECD of murine MAG (murine MAG-ECD) in the conditioned medium (CM) of 293 T cotransfected cells using co-immunoprecipitation. Tie-2-ECD-FC served as a negative control. D Representative flow cytometric analysis showing that purified ANGPTL2-Flag protein enhanced GFP expression in murine MAG chimeric reporter cells (Murine MAG). Control reporter cells (control) without murine MAG-ECD were examined. E Quantitative data in Panel D (n = 3). F Binding kinetics of ANGPTL2-Flag to murine MAG-ECD were measured using a biolayer interferometry instrument (Octed RED 96/96S). (***p < 0.001) Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/36855057), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Angiopoietin-like 2 by Flow Cytometry

Murine homologue of human MAG binds to ANGPTL2. A Flow cytometric analysis of conditioned medium with/without ANGPTL2-Flag binding to 293 T cells with transient transfection of plasmids of N1-mMAG-EGFP (murine MAG, full-length) or N1-EGFP (vector). Percentage of binding/unbinding cells to ANGPTL2 in EGFP+ cells are shown. B Quantitative data of percentage of binding cells in Panel A (n = 3). C ANGPLT2 bound to the ECD of murine MAG (murine MAG-ECD) in the conditioned medium (CM) of 293 T cotransfected cells using co-immunoprecipitation. Tie-2-ECD-FC served as a negative control. D Representative flow cytometric analysis showing that purified ANGPTL2-Flag protein enhanced GFP expression in murine MAG chimeric reporter cells (Murine MAG). Control reporter cells (control) without murine MAG-ECD were examined. E Quantitative data in Panel D (n = 3). F Binding kinetics of ANGPTL2-Flag to murine MAG-ECD were measured using a biolayer interferometry instrument (Octed RED 96/96S). (***p < 0.001) Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/36855057), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Angiopoietin-like 2 by Immunohistochemistry

ANGPTL2 promotes oligodendrocyte differentiation and remyelination under pathological conditions. A Representative images of the corpus callosum region stained with oil red O in Angptl2+/+ and Angptl2−/− mice after the withdrawal of cuprizone for two additional weeks. Myelin was stained in red color. Meanwhile, more nonspecific oil red O positive deposits (a phenomenon that oil red O deposits in damaged myelin region) were observed in Angptl2−/− mice (arrows). B Quantitative data of the demyelinating areas in Panel A; six sections from each mouse were analyzed (n = 7). C Immunofluorescence staining for MBP, MOG, MAG and GFAP in the corpus callosum region from Angptl2+/+ and Angptl2−/− mice after the withdrawal of cuprizone for two additional weeks. D Quantification of the fluorescence intensity of MBP, MOG, MAG and GFAP in the corpus callosum region in Panel C. Four sections from each mouse were analyzed (n = 6–7). E Representative electron microscopy images of the corpus callosum from Angptl2+/+ and Angptl2−/− mice after the withdrawal of cuprizone for two additional weeks; red arrow indicates uncompacted myelin lamella; F Quantification of the G ratios of the remyelinated axons of the corpus callosum in Panel E. Approximately 70 axons were counted per mouse (n = 3). G The scatter plot for the individual G-ratio values and axonal size distribution of the corpus callosum in Panel E. H Motor coordination of Angptl2+/+ and Angptl2−/− mice after cuprizone (Cupz) treatment for five weeks and cuprizone withdrawal for two additional weeks in the rotarod test; 7–8 mice were used for each group. (*p < 0.05, **p < 0. 01, ***p < 0.001) Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/36855057), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Angiopoietin-like 2 by Flow Cytometry

Human MAG is a new receptor for ANGPTL2. A Flow cytometric analysis of conditioned medium with/without ANGPTL2-Flag binding to 293T cells with transient transfection of plasmids of N1-hMAG-EGFP (human MAG, full-length) or N1-EGFP (vector). Percentage of binding/unbinding cells to ANGPTL2 in EGFP+ cells are shown. B Quantitative data of percentage of binding cells in Panel A (n = 3). C ANGPLT2 bound to the ECD of human MAG (human MAG-ECD-FC) in the conditioned medium (CM) of cotransfected 293T cells using co-immunoprecipitation. Tie-2-ECD-FC served as a negative control. D Representative flow cytometric analysis showing that purified ANGPTL2-Flag protein enhanced GFP expression in human MAG chimeric reporter cells (Human MAG). Control reporter cells (control) without human MAG-ECD were examined. E Quantitative data in Panel D (n = 3). F Binding kinetics of ANGPTL2-Flag to human MAG-ECD were measured using a biolayer interferometry instrument (Octed RED 96/96S). G–H Flow cytometric plots for the binding of ANGPTL2-Flag to full-length, mutant IgG 3 domain or IgG 4 domain of human MAG expressed 293T cells. Percentage of binding/unbinding cells to ANGPTL2 in EGFP+ cells (G) and quantitative data are shown (H) (n = 3). (***p < 0.001) Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/36855057), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Angiopoietin-like 2 by Immunohistochemistry

ANGPTL2 is necessary for fibrotic vasculopathy.(A) qPCR of Angptl2 mRNA in WT and Snail-tg mouse keratinocytes (mKT). (B) Secreted ANGPTL2 (in green) in WT and Snail-tg skin at P3. The dotted line denotes the basement membrane separating the epidermis (epi) from the underlying dermis (der). (C) ELISA for secreted ANGPTL2 in WT and Snail-tg mKT conditioned media (CM). (D) Expression of Angptl2 in keloid lesion skin (n = 3) compared with nonlesion skin (n = 3) (from GSE92566) and in healthy (n = 44) compared with affected forearm skin from diffuse (n = 180) and localized SSc (n = 115) patients (from GSE181549). The expression values are from the GEO2R algorithm’s output. (E) Secreted ANGPTL2 (in green) in normal and SSc skin. Representative of n = 5 SSc samples and normal skin controls. (F) qPCR of vasculopathy and endoMT-related genes in SVEC4-10 endothelial cells treated with WT, Snail-tg, and Snail-tg/Angptl2-KO mKT conditioned media. (G) Quantification of collagen contraction assay in SVEC4-10 endothelial cells treated with WT, Snail-tg, and Snail-tg/Angptl2-KO mKT conditioned media. Nuclei are marked in blue. Scale bars: 50 μm. Data are shown as mean ± SEM; P values were calculated using paired Student’s t test (A), unpaired Welch’s t test (D), and 1-way ANOVA followed by Tukey’s post hoc analysis (F and G). *P < 0.05, **P < 0.01, ****P < 0.0001, NS > 0.05. All experiments except (D and E) are n = 3 biological replicates. Image collected and cropped by CiteAb from the following open publication (https://www.jci.org/articles/view/177123), licensed under a CC-BY license. Not internally tested by R&D Systems.