Anti-Interferon beta antibody [EPR22186-266]

• IP

Supplier Data

Immunoprecipitation - Anti-Interferon beta antibody [EPR22186-266] (AB218229)

Interferon beta was immunoprecipitated from 0.35 mg of RAW 264.7 (mouse macrophage cell line transformed with Abelson murine leukemia virus) treated with 50μg/ml DMXAA (ab143018) for 1h, then together with 300ng/ml brefeldin A (BFA) for another 6h whole cell lysate with ab218229 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab218229 at 1/500 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/1000 dilution.

Lane 1 : RAW 264.7 (treated with DMXAA (ab143018)/BFA) whole cell lysate 10 μg (Input).
Lane 2 : ab218229 IP in RAW 264.7 (treated with DMXAA (ab143018)/BFA) whole cell lysate.
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab218229 in RAW 264.7 (treated with DMXAA (ab143018)/BFA) whole cell lysate.

Blocking and dilution buffer : 5% NFDM/TBST.

All lanes:

Immunoprecipitation - Anti-Interferon beta antibody [EPR22186-266] (ab218229)

Predicted band size: 22 kDa,30 kDa

Observed band size: 24 kDa,26 kDa,30 kDa

false

Exposure time: 10s

• WB

Supplier Data

Western blot - Anti-Interferon beta antibody [EPR22186-266] (AB218229)

Exposure time : Lanes 1 and 2 : 26 seconds; Lanes 3 and 4 : 3 minutes.

Blocking/Dilution buffer : 5% NFDM/TBST.

Interferon beta can be induced to increase the expression by DMXAA or LPS in RAW264.7 cell (PMID : 23027866, PMID : 16020513).

The doublet is likely due to isoforms (PMID : 7352261).

All lanes:

Western blot - Anti-Interferon beta antibody [EPR22186-266] (ab218229) at 1/1000 dilution

Lane 1:

Untreated RAW 264.7 (mouse macrophage cell line transformed with Abelson murine leukemia virus) whole cell lysate at 20 µg

Lane 2:

RAW 264.7 treated with 50 μg/ml DMXAA (<a href='/en-us/products/unavailable/dmxaa-asa404-vegfr-inhibitor-ab143018'>ab143018</a>) for 7 hours, then with 300 ng/ml Brefeldin A (BFA) added after 1 hour, whole cell lysate at 20 µg

Lane 3:

Untreated RAW 264.7 whole cell lysate at 20 µg

Lane 4:

RAW264.7 treated with 100 ng/ml lipopolysaccharide (LPS) for 6 hours, then with 300 ng/ml Brefeldin A (BFA) added after 3 hours, whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 22 kDa

Observed band size: 24 kDa,26 kDa

false

• WB

CiteAb

Western blot - Anti-Interferon beta antibody [EPR22186-266] (AB218229)

Interferon beta western blot using anti-Interferon beta antibody [EPR22186-266] ab218229. Publication image and figure legend from Chen, H., Chew, G., et al., 2022, Nat Commun, PubMed 36450710.

ab218229 was used in this publication in western blot. This may not be the same as the application(s) guaranteed by Abcam. For a full list of applications guaranteed by Abcam for ab218229 please see the product overview.

WWP2 regulates IRF7 network activity in macrophages in NICM.a Scatter plot of the relative activity of transcription factor (TF)-regulons in cardiac macrophages from WWP2Mut/Mut (Mut/Mut) and WWP2wt/wt (WT) mice treated with Ang-II (7 days). For each regulon, the difference in expression score between Mut/Mut and WT and its statistical significance score are shown on the x axis and y axis, respectively. The number of downstream target genes in each regulon is proportional to the circle size (more details in Methods). The IRF7 regulon showed the largest and most significant expression score difference between Mut/Mut and WT cardiac macrophages. b Average expression score of the IRF7 regulon across cardiac macrophage clusters from WT and Mut/Mut mice treated with Ang-II (7 days). (P values by two-sided Wilcoxon test). c Breakdown of IRF7 targets enrichment in the profibrotic hECM-network (n = 237 genes) previously identified in LV of dilated cardiomyopathy patients39. 80 (33.8%) of these hECM-network genes are targets of IRF7. d, e qRT-PCR (d) and WB (e) of representative IFN signaling genes (IFNα, IFNβ, and IFNγ) in WT and WWP2-/- (-/-) BMDMs with and without LPS stimulation. n = 5 per experimental group. f. IRF7 binding site motif derived from previously published IRF7 ChIP-seq analysis67. g IRF7 ChIP-qPCR analysis and schematic showing the IRF7 binding site motifs (P < 10-4) within the Ccl5 and Irf7 genes. *Motifs matching to (Ccl5, Irf7) PCR products (purple thick line), which were used in the ChIP-qPCR analysis. h,i qRT-PCR (h) and WB (i) of IRF7 mRNA and protein expression in WT and WWP2-/- (-/-) BMDMs. n = 3–6 for each group. j ChIP-qPCR analysis of IRF7 occupancy sites on Ccl5 (left) or Irf7 (right) in WT and WWP2-/- BMDMs treated with LPS. Enrichment is normalized to input DNA, and represented as fold-enrichment relative to vehicle controls in WT BMDMs. In each case, LPS stimulation (100 ng/ml, 4 hrs). Unless otherwise indicated, data are shown as dot-plots with mean ± SD, and statistical significance is assessed by the non-parametric Mann–Whitney U test.

false