Anti-Histone H3 (mono methyl R2) antibody [EPR17704] - ChIP Grade

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-Histone H3 (mono methyl R2) antibody [EPR17704] - ChIP Grade (AB176844)

Immunofluorescent analysis of 4% paraformaldehyde-fixed 0.1% Triton X-100 permeabilized HeLa (Human epithelial cells from cervix adenocarcinoma) cells labeling Histone H3 (mono methyl R2) with ab176844 at 1/1000 dilution followed by Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/500 dilution (green). Confocal image showing nuclear staining on HeLa cell line. The nuclear counter stain is DAPI (blue). Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and ab150120 (Alexa Fluor^® 594 Goat anti-Mouse secondary) at 1/500 dilution (red).
The negative controls are as follows :
-ve control 1 : ab176844 at 1/1000 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/500 dilution.
-ve control 2 : ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor®488 Goat Anti-Rabbit IgG H&L) at 1/500 dilution.

• ChIP

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ChIP - Anti-Histone H3 (mono methyl R2) antibody [EPR17704] - ChIP Grade (AB176844)

Chromatin was prepared from HeLa (Human epithelial cells from cervix adenocarcinoma) cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10 minutes. The ChIP was performed with 25μg of chromatin 2μg of ab176844 (blue) and 20μl of Anti rabbit IgG sepharose beads. 2μg of rabbit normal IgG was added to the beads control (yellow). The immunoprecipitated DNA was quantified by real time PCR (Sybr green approach). Primers and probes are located in the first kb of the transcribed region.

• WB

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Western blot - Anti-Histone H3 (mono methyl R2) antibody [EPR17704] - ChIP Grade (AB176844)

Blocking/Dilution buffer : 5%BSA /TBST.

All lanes:

Western blot - Anti-Histone H3 (mono methyl R2) antibody [EPR17704] - ChIP Grade (ab176844) at 1/5000 dilution

All lanes:

HeLa (Human epithelial cells from cervix adenocarcinoma) whole cell lysate at 10 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/1000 dilution

Predicted band size: 14 kDa,15 kDa,30 kDa

Observed band size: 14 kDa,15 kDa,31 kDa

false

Exposure time: 15s

• WB

Supplier Data

Western blot - Anti-Histone H3 (mono methyl R2) antibody [EPR17704] - ChIP Grade (AB176844)

Blocking/Dilution buffer : 5%BSA /TBST.

All lanes:

Western blot - Anti-Histone H3 (mono methyl R2) antibody [EPR17704] - ChIP Grade (ab176844) at 1/5000 dilution

All lanes:

NIH/3T3 (mouse embryo) whole cell lysate at 10 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/1000 dilution

Predicted band size: 15 kDa

Observed band size: 15 kDa

false

Exposure time: 30s

• PepArr

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Peptide Array - Anti-Histone H3 (mono methyl R2) antibody [EPR17704] - ChIP Grade (AB176844)

ab176884 was tested in Peptide Array against 501 different modified and unmodified histone peptides; each peptide is printed on the array at six concentrations (each in triplicate).
Circle area represents affinity between the antibody and a peptide : all antigen-containing peptides are displayed as red circles, all other peptides as blue circles. The affinity is calculated as area under curve when antibody binding values are plotted against the corresponding peptide concentration. Each circle area is normalized to the peptide with the strongest affinity.
The complete dataset, including full list of all peptides and information on the position of each peptide in the diagram, is available under the Product Protocol section.

• WB

CiteAb

Western blot - Anti-Histone H3 (mono methyl R2) antibody [EPR17704] - ChIP Grade (AB176844)

Histone H3 (mono methyl R2) western blot using anti-Histone H3 (mono methyl R2) antibody [EPR17704] - ChIP Grade ab176844. Publication image and figure legend from Günes Günsel, G., Conlon, T. M., et al., 2022, Nat Commun, PubMed 35288557.

ab176844 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 ab176844 please see the product overview.

PRMT7 targets RAP1A expression through histone methylation.a Western blot analysis of RAP1A/RAP1B expression in Prmt7+/+ and Prmt7null MH-S cells (repeated twice). b mRNA expression level of Rap1a and Rap1b determined by qPCR in Prmt7+/+ and Prmt7null MH-S macrophage cells (n = 4 replicates per cell line). c–f ATAC-Seq analysis of Prmt7+/+ v Prmt7null MH-S cells. c ATAC-seq signal enrichment peaks around the transcription start site (TSS) of the Rap1a gene in Prmt7+/+ and Prmt7null MH-S cells and difference in peak height across the two lines. d Heat map of tag distributions across TSSs for Prmt7+/+ and Prmt7null MH-S cells. e Peak correlation scatter plot. f Pie chart showing the genomic distribution of accessible regions in Prmt7+/+ and Prmt7null MH-S cells. g Schematic representation of the location of the Rap1a regions targeted for qPCR following ChIP, plus H3K4me1 and H3K27a enrichment in BMDM (from UCSC genome browser Track accessions : wgEncodeEM002658 and wgEncodeEM002657). h Enrichment of H3R2 mono and dimethylation at Rap1a regions in Prmt7+/+ and Prmt7null MH-S by qPCR following ChIP (n = 2 per cell line). i Western blot analysis of mono and dimethylation of H3R2 in Prmt7+/+ and Prmt7null MH-S cells (repeated twice). j Dot blot depicting Rap1a expression (log-transformed, normalized UMI counts) and percentage of cells positive for Rap1a in monocytes from mouse lung single-cell RNA-seq data following exposure to FA (n = 3) and CS for 2 (n = 5) and 4 months (n = 5). k mRNA expression levels of RAP1A determined by qPCR in the monocytes of smokers (n = 10) and non-smokers (n = 11) isolated from the peripheral blood. l MFI of ITGAL and ITGAM surface expression as determined by flow cytometry in Prmt7+/+ MH-S cells incubated with 20 μM GGTI (RAP1 inhibitor) for 2 h and analysed 6 h later (n = 2, repeated three times). m Western blot analysis of phosphorylated ERK in WT MH-S cells pretreated with the RAP1 inhibitor GGTI (20 μM) for 2 h and incubated for 15 min with 1 μg/ml LPS. Quantification relative to actin shown (repeated two times). Data shown mean ± SD, P values shown in charts determined by unpaired two-tailed Student’s t-test (b, h, k, l), one-way ANOVA Bonferroni’s multiple comparisons test (m). Source data are provided as a Source Data file.

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