Product datasheet

Anti-EHMT2/G9A antibody [EPR18894] - ChIP Grade ab185050
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4 Abreviews 33 References 画像数 18
製品の概要
製品名

Anti-EHMT2/G9A antibody [EPR18894] - ChIP Grade

製品の詳細

Rabbit monoclonal [EPR18894] to EHMT2/G9A - ChIP Grade

由来種

Rabbitアプリケーション

適用あり : ChIP-sequencing, WB, IHC-P, ICC/IF, IP, Flow Cyt (Intra), ChIC/CUT&RUN-seq

種交差性交差種 : Mouse, Rat, Human

免疫原

Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.

ポジティブ･コントロール

WB: HeLa, HEK-293, Jurkat, HepG2, NCCIT, F9, Neuro-2a, LLC1, C6, RAW 264.7, PC-12 andNIH/3T3 whole cell lysates. Human fetal heart and fetal kidney lysates. IHC-P: Human colon,
Human gastric adenocarcinoma, mouse liver and rat kidney tissues. ICC/IF: HeLa. Flow Cyt(intra): HeLa cells. IP: HeLa whole cell lysate. ChIP-seq: HeLa cells.

特記事項This product is a recombinant monoclonal antibody, which offers several advantages including:- High batch-to-batch consistency and reproducibility- Improved sensitivity and specificity- Long-term security of supply- Animal-free production
For more information see here.
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit
monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.

製品の特性
製品の状態

Liquid

保存方法

Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C longterm. Avoid freeze / thaw cycle.

バッファー

pH: 7.2
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol, 0.05% BSA

精製度

Protein A purified

ポリ /モノモノクローナル

クローン名EPR188941アイソタイプ

IgG

アプリケーション

The Abpromise guarantee

Abpromise保証は、 次のテスト済みアプリケーションにおけるab185050の使用に適用されます

アプリケーションノートには、推奨の開始希釈率がありますが、適切な希釈率につきましてはご検討ください。
アプリケーション

特記事項

Abreviews

Use 8µg for 107 cells.

ChIP-sequencing
WB

(1)

IHC-P

1/1000. Detects a band of approximately 170, 160 kDa(predicted molecular weight: 132 kDa).
1/500. Perform heat mediated antigen retrieval with Tris/EDTA
buffer pH 9.0 before commencing with IHC staining protocol.

ICC/IF(1)

1/1000.

IP

(1)

1/50.

Flow Cyt (Intra)

1/150.

ChIC/CUT&RUN-seq

Use at an assay dependent concentration. 5µg

ターゲット情報機能Histone methyltransferase that specifically mono- and dimethylates 'Lys-9' of histone H3(H3K9me1 and H3K9me2, respectively) in euchromatin. H3K9me represents a specific tag for
epigenetic transcriptional repression by recruiting HP1 proteins to methylated histones. Also
mediates monomethylation of 'Lys-56' of histone H3 (H3K56me1) in G1 phase, leading topromote interaction between histone H3 and PCNA and regulating DNA replication. Also weakly
methylates 'Lys-27' of histone H3 (H3K27me). Also required for DNA methylation, the histone
methyltransferase activity is not required for DNA methylation, suggesting that these 2 activities
function independently. Probably targeted to histone H3 by different DNA-binding proteins like
E2F6, MGA, MAX and/or DP1. May also methylate histone H1. In addition to the histone
methyltransferase activity, also methylates non-histone proteins: mediates dimethylation of 'Lys373' of p53/TP53. Also methylates CDYL, WIZ, ACIN1, DNMT1, HDAC1, ERCC6, KLF12 anditself.

組織特異性Expressed in all tissues examined, with high levels in fetal liver, thymus, lymph node, spleen andperipheral blood leukocytes and lower level in bone marrow.

配列類似性

Belongs to the class V-like SAM-binding methyltransferase superfamily. Histone-lysine
methyltransferase family. Suvar3-9 subfamily.
Contains 7 ANK repeats.
Contains 1 post-SET domain.
Contains 1 pre-SET domain.
Contains 1 SET domain.

ドメイン

The SET domain mediates interaction with WIZ.
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The ANK repeats bind H3K9me1 and H3K9me2.
翻訳後修飾

Methylated at Lys-185; automethylated.

細胞内局在Nucleus. Chromosome. Associates with euchromatic regions. Does not associate withheterochromatin.

画像

ChIC/CUT&RUN was performed using a pAG-MNAse at a final
concentration of 700 ng/mL. 2.5X10^5 of Human wild-type HeLa
cell line (ab255928) or EHMT2 (G9A) knockout HeLa cell line(ab265149) were used along with 5µg of ab185050 [EPR18894].
Assay Quality Control was conducted using 5µg Anti-CTCF(ab188408) on the same cell lines. The resulting DNA wassequenced on the Illumina NovaSeq 6000 to a depth of 10 million
reads. The negative IgG control ab172730 is also shown.
Additional screenshots of mapped reads can be downloaded here.
The University of Geneva owns patents relevant to ChIC (ChromatinImmuno-Cleavage) methods.
ChIC/CUT&RUN sequencing - Anti-EHMT2/G9A
antibody [EPR18894] - ChIP Grade (ab185050)

All lanes : Anti-EHMT2/G9A antibody [EPR18894] - ChIP Grade(ab185050) at 1/1000 dilution

Lane 1 : Wild-type HeLa cell lysate
Lane 2 : EHMT2/G9A knockout HeLa cell lysate
Lane 3 : HEK-293 cell lysate
Lane 4 : HepG2 cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Western blot - Anti-EHMT2/G9A antibody[EPR18894] - ChIP Grade (ab185050)

Predicted band size: 132 kDaLanes 1-4: Merged signal (red and green). Green - ab185050
observed at 160 kDa. Red - loading control ab8245 observed at 37kDa.
ab185050 Anti-EHMT2/G9A antibody [EPR18894] was shown tospecifically react with EHMT2/G9A in wild-type HeLa cells. Loss of3

signal was observed when knockout cell line ab265149 (knockoutcell lysate ab257080) was used. Wild-type and EHMT2/G9Aknockout samples were subjected to SDS-PAGE. ab185050 andAnti-GAPDH antibody [6C5] - Loading Control (ab8245) wereincubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000
dilution respectively. Blots were developed with Goat anti-Rabbit
IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat antiMouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776)
secondary antibodies at 1 in 20000 dilution for 1 hour at room
temperature before imaging.

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1%
Triton X-100 permeabilized HeLa (Human epithelial cells fromcervix adenocarcinoma) cells labeling EHMT2/G9A with ab185050
at 1/1000 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor®
488) (ab150077) secondary antibody at 1/1000 dilution (green).
Confocal image showing nuclear staining on HeLa cell line.
The nuclear counterstain is DAPI (blue).
Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at
1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mousesecondary) at 1/1000 dilution (red).
Immunocytochemistry/ Immunofluorescence - Anti-

The negative controls are as follows:-EHMT2/G9A antibody [EPR18894] - ChIP Grade-ve control 1: ab185050 at 1/1000 dilution followed by ab150120(ab185050)(AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 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 1/1000 dilution.

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Chromatin was prepared from HeLa cells. Cells were fixed with 1%
formaldehyde for 10 minutes. ChIP was performed with 107 HeLacells and 8 µg of ab185050 [EPR18894]. ChIP DNA wassequenced on the Illumina NovaSeq 6000 to a depth of 30 million
reads.
Additional screenshots of mapped reads can be downloaded here.

ChIP-sequencing - Anti-EHMT2/G9A antibody[EPR18894] - ChIP Grade (ab185050)

Immunohistochemical analysis of paraffin-embedded Rat kidney
tissue labeling EHMT2/G9A with ab185050 at 1/500 dilution,
followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500dilution.
Nucleus staining on rat kidney is observed.
Counter stained with Hematoxylin.
Secondary antibody only control: Used PBS instead of primary
antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP)(ab97051) at 1/500 dilution.
Perform heat mediated antigen retrieval with Tris/EDTA buffer pH
Immunohistochemistry (Formalin/PFA-fixed paraffin-

9.0 before commencing with IHC staining protocol.

embedded sections) - Anti-EHMT2/G9A antibody[EPR18894] - ChIP Grade (ab185050)

All lanes : Anti-EHMT2/G9A antibody [EPR18894] - ChIP Grade(ab185050) at 1/1000 dilution

Lane 1 : Wild-type HAP1 cell lysate
Lane 2 : EHMT2/G9A knockout HAP1 cell lysate
Lane 3 : HEK293 cell lysate
Lane 4 : HepG2 cell lysate
Lysates/proteins at 20 µg per lane.

Predicted band size: 132 kDa
Western blot - Anti-EHMT2/G9A antibody[EPR18894] - ChIP Grade (ab185050)

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Lanes 1 - 4: Merged signal (red and green). Green - ab185050
observed at 160 kDa. Red - loading control, ab8245, observed
at 37 kDa.
ab185050 was shown to recognize EHMT2/G9A
when EHMT2/G9A knockout samples were used, along withadditional cross-reactive bands. Wild-type and EHMT2/G9A
knockout samples were subjected to SDS-PAGE. ab185050 andab8245 (loading control to GAPDH) were diluted 1/1000 and 1/10000 respectively and incubated overnight at 4°C. Blots weredeveloped with Goat anti-Rabbit IgG H&L (IRDye® 800CW)
preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye®
680RD) preadsorbed (ab216776) secondary antibodies at 1/10000 dilution for 1 h at room temperature before imaging.

All lanes : Anti-EHMT2/G9A antibody [EPR18894] - ChIP Grade(ab185050) at 1/1000 dilution

Lane 1 : HEK-293 (Human epithelial cells from embryonic kidney)whole cell lysate

Lane 2 : Jurkat (Human T cell leukemia cells from peripheral blood)
whole cell lysate

Lane 3 : HepG2 (Human liver hepatocellular carcinoma) whole celllysate

Lane 4 : NCCIT (Human pluripotent embryonic carcinoma) wholecell lysate
Western blot - Anti-EHMT2/G9A antibody[EPR18894] - ChIP Grade (ab185050)

Lysates/proteins at 20 µg per lane.

SecondaryAll lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at
1/100000 dilution

Predicted band size: 132 kDa
Observed band size: 160,170 kDa

Exposure time: 30 seconds

Blocking/Dilution buffer: 5% NFDM/TBST.
The expression profile/ molecular weight observed is consistent
with what has been described in the literature (PMID 16702210).

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Anti-EHMT2/G9A antibody [EPR18894] - ChIP Grade (ab185050)
at 1/1000 dilution + Human fetal heart lysate at 10 µg

SecondaryAnti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at
1/10000 dilution

Predicted band size: 132 kDa
Observed band size: 160,170 kDa

Western blot - Anti-EHMT2/G9A antibody

Exposure time: 3 minutes[EPR18894] - ChIP Grade (ab185050)

Blocking/Dilution buffer: 5% NFDM/TBST.
The expression profile/ molecular weight observed is consistent
with what has been described in the literature (PMID 16702210).

Anti-EHMT2/G9A antibody [EPR18894] - ChIP Grade (ab185050)
at 1/1000 dilution + Human fetal kidney lysate at 10 µg

SecondaryAnti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at
1/10000 dilution

Predicted band size: 132 kDa
Observed band size: 160,170 kDa

Western blot - Anti-EHMT2/G9A antibody

Exposure time: 8 seconds[EPR18894] - ChIP Grade (ab185050)

Blocking/Dilution buffer: 5% NFDM/TBST.
The expression profile/ molecular weight observed is consistent
with what has been described in the literature (PMID 16702210).

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All lanes : Anti-EHMT2/G9A antibody [EPR18894] - ChIP Grade(ab185050) at 1/1000 dilution

Lane 1 : F9 (Mouse embyro testicular cancer cell line) whole celllysate

Lane 2 : Neuro-2a (Mouse neuroblastoma cells) whole cell lysate
Lane 3 : LLC1 (Mouse lung carcinoma cell line) whole cell lysate
Lysates/proteins at 10 µg per lane.

SecondaryWestern blot - Anti-EHMT2/G9A antibody

All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at[EPR18894] - ChIP Grade (ab185050)

1/100000 dilution

Predicted band size: 132 kDa
Observed band size: 160,170 kDa

Exposure time: 5 seconds

Blocking/Dilution buffer: 5% NFDM/TBST.
The expression profile/ molecular weight observed is consistent
with what has been described in the literature (PMID 16702210).

8All lanes : Anti-EHMT2/G9A antibody [EPR18894] - ChIP Grade(ab185050) at 1/1000 dilution

Lane 1 : C6 (Rat glial tumor cells) whole cell lysate
Lane 2 : RAW 264.7 (Mouse macrophage cells transformed withAbelson murine leukemia virus) whole cell lysate

Lane 3 : PC-12 (Rat adrenal gland pheochromocytoma) whole celllysate

Lane 4 : NIH/3T3 (Mouse embyro fibroblast cells) whole cell lysate
Lysates/proteins at 10 µg per lane.
Western blot - Anti-EHMT2/G9A antibody[EPR18894] - ChIP Grade (ab185050)

SecondaryAll lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at
1/100000 dilution

Predicted band size: 132 kDa
Observed band size: 160,170 kDa

Exposure time: 8 seconds

Blocking/Dilution buffer: 5% NFDM/TBST.
The expression profile/ molecular weight observed is consistent
with what has been described in the literature (PMID 16702210).

Immunohistochemical analysis of paraffin-embedded Human colon
tissue labeling EHMT2/G9A with ab185050 at 1/500 dilution,
followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500dilution.
Nucleus staining on epithelial cells of the normal Human colon isobserved.
Counter stained with Hematoxylin.
Secondary antibody only control: Used PBS instead of primary
antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP)(ab97051) at 1/500 dilution.
Immunohistochemistry (Formalin/PFA-fixed paraffin-Perform heat mediated antigen retrieval with Tris/EDTA buffer pH

embedded sections) - Anti-EHMT2/G9A antibody

9.0 before commencing with IHC staining protocol.

[EPR18894] - ChIP Grade (ab185050)

9Immunohistochemical analysis of paraffin-embedded Humangastric adenocarcinoma tissue labeling EHMT2/G9A with
ab185050 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L(HRP) (ab97051) at 1/500 dilution.
Nucleus staining on tumor cells of the gastric adenocarcinoma isobserved.
Counter stained with Hematoxylin.
Secondary antibody only control: Used PBS instead of primary
antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP)(ab97051) at 1/500 dilution.
Immunohistochemistry (Formalin/PFA-fixed paraffin-Perform heat mediated antigen retrieval with Tris/EDTA buffer pH

embedded sections) - Anti-EHMT2/G9A antibody

9.0 before commencing with IHC staining protocol.

[EPR18894] - ChIP Grade (ab185050)

Immunohistochemical analysis of paraffin-embedded Mouse livertissue labeling EHMT2/G9A with ab185050 at 1/500 dilution,
followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500
dilution.
Nucleus staining on hepatocytes of the mouse liver is observed.
Counter stained with Hematoxylin.
Secondary antibody only control: Used PBS instead of primary
antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP)(ab97051) at 1/500 dilution.
Perform heat mediated antigen retrieval with Tris/EDTA buffer pHImmunohistochemistry (Formalin/PFA-fixed paraffin-9.0 before commencing with IHC staining protocol.

embedded sections) - Anti-EHMT2/G9A antibody[EPR18894] - ChIP Grade (ab185050)

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Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1%
Triton X-100 permeabilized MCF7 (Human breast adenocarcinoma
cell line) cells labeling EHMT2/G9A with ab185050 at 1/100dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488)
(ab150077) secondary antibody at 1/1000 dilution (green).
Confocal image showing no staining on MCF7 cell line, as MCF7
cells have a very low level expression of EHMT2/G9A.
The nuclear counterstain is DAPI (blue).
Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at
1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse
Immunocytochemistry/ Immunofluorescence - Anti-secondary) at 1/1000 dilution (red).

EHMT2/G9A antibody [EPR18894] - ChIP GradeThe negative controls are as follows:-(ab185050)-ve control 1: ab185050 at 1/70 dilution followed by ab150120(AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 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 1/1000 dilution.

Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed
HeLa (Human epithelial cells from cervix adenocarcinoma) cells
labeling EHMT2/G9Awith ab185050 at 1/150 dilution (red)compared with a rabbit monoclonal IgG isotype control (ab172730;black) and an unlabelled control (cells without incubation withprimary antibody and secondary antibody; blue). Goat anti rabbit
IgG (FITC) at 1/150 dilution was used as the secondary antibody.

Flow Cytometry (Intracellular) - Anti-EHMT2/G9A
antibody [EPR18894] - ChIP Grade (ab185050)

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EHMT2/G9A was immunoprecipitated from 1mg of HeLa (Humanepithelial cells from cervix adenocarcinoma) whole cell lysate with
ab185050 at 1/50 dilution.
Western blot was performed from the immunoprecipitate using
ab185050 at 1/1000 dilution.
VeriBlot for IP Detection Reagent (HRP) (ab131366) was used fordetection at 1/10000 dilution.
Lane 1: HeLa whole cell lysate 10ug (Input).
Lane 2: ab185050 IP in HeLa whole cell lysate.
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab185050 in
Immunoprecipitation - Anti-EHMT2/G9A antibody[EPR18894] - ChIP Grade (ab185050)

HeLa whole cell lysate.
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 30 seconds.

Anti-EHMT2/G9A antibody [EPR18894] - ChIP
Grade (ab185050)Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"

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