Anti-ATF3 抗体 [EPR19488] - ChIP Grade (ab207434)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR19488] to ATF3 - ChIP Grade
- Suitable for: ChIC/CUT&RUN-seq, ChIP, WB, IP, ICC/IF
- Knockout validated
- Reacts with: Mouse, Human
Related conjugates and formulations
製品の概要
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製品名
Anti-ATF3 antibody [EPR19488] - ChIP Grade
ATF3 一次抗体 製品一覧 -
製品の詳細
Rabbit monoclonal [EPR19488] to ATF3 - ChIP Grade -
由来種
Rabbit -
特異性
Stimulation may be required to allow detection of the target protein due to low levels of endogenous expression in some samples. Please see images below for recommended treatment conditions and positive controls.
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アプリケーション
適用あり: ChIC/CUT&RUN-seq, ChIP, WB, IP, ICC/IFmore details -
種交差性
交差種: Mouse, Human -
免疫原
Recombinant full length protein. This information is proprietary to Abcam and/or its suppliers.
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ポジティブ・コントロール
- WB: HCT116, HEK-293, HeLa, A431, HepG2, LnCap, Jurkat, THP-1, RAW 264.7 cell lysates. ICC/IF: THP-1, RAW 264.7, HAP1 cells. ChIC/CUT&RUN-seq: HeLa cells
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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
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
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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 long term. Avoid freeze / thaw cycle. -
バッファー
pH: 7.2
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA -
Concentration information loading...
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精製度
Protein A purified -
ポリ/モノ
モノクローナル -
クローン名
EPR19488 -
アイソタイプ
IgG -
研究分野
関連製品
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Alternative Versions
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Isotype control
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KO cell lines
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KO cell lysates
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Related Products
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ab207434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 | Abreviews | 特記事項 |
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ChIC/CUT&RUN-seq |
Use at an assay dependent concentration.
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ChIP |
Use at an assay dependent concentration.
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WB |
1/1000. Detects a band of approximately 21 kDa (predicted molecular weight: 21 kDa).
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IP |
1/50.
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ICC/IF |
1/100.
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特記事項 |
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ChIC/CUT&RUN-seq
Use at an assay dependent concentration. |
ChIP
Use at an assay dependent concentration. |
WB
1/1000. Detects a band of approximately 21 kDa (predicted molecular weight: 21 kDa). |
IP
1/50. |
ICC/IF
1/100. |
ターゲット情報
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機能
This protein binds the cAMP response element (CRE) (consensus: 5'-GTGACGT[AC][AG]-3'), a sequence present in many viral and cellular promoters. Represses transcription from promoters with ATF sites. It may repress transcription by stabilizing the binding of inhibitory cofactors at the promoter. Isoform 2 activates transcription presumably by sequestering inhibitory cofactors away from the promoters. -
配列類似性
Belongs to the bZIP family. ATF subfamily.
Contains 1 bZIP domain. -
細胞内局在
Nucleus. - Information by UniProt
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参照データベース
- Entrez Gene: 467 Human
- Entrez Gene: 11910 Mouse
- Omim: 603148 Human
- SwissProt: P18847 Human
- SwissProt: Q60765 Mouse
- Unigene: 460 Human
- Unigene: 2706 Mouse
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別名
- Activating transcription factor 3 antibody
- ATF3 antibody
- ATF3_HUMAN antibody
see all
画像
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ChIC/CUT&RUN was performed using a pAG-MNAse at a final concentration of 700 ng/µL. 2.5X10^5 of Human ATF3 knockout HeLa cell line (ab264908) or Human wild-type HeLa cell line (ab255448) were used along with 5µg of Anti-ATF3 antibody (ab207434). Assay Quality Control was conducted using 5µg Anti-CTCF (ab188408) on the same cell lines. The resulting DNA was sequenced 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 (Chromatin Immuno-Cleavage) methods. -
All lanes : Anti-ATF3 antibody [EPR19488] - ChIP Grade (ab207434) at 1/1000 dilution
Lane 1 : Wild-type HeLa cell lysate
Lane 2 : ATF3 knockout HeLa cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 21 kDa
Observed band size: 21 kDaLanes 1- 2: Merged signal (red and green). Green - ab207434 observed at 21 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.
ab207434 was shown to react with ATF3 in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab264908 (knockout cell lysate ab257073) was used. Wild-type HeLa and ATF3 knockout HeLa cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab207434 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) overnight at 4°C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were developed 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 in 20000 dilution for 1 hour at room temperature before imaging.
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ab207434 staining ATF3 in wild-type Hap1 cells (top panel) and ATF3 knockout Hap1 cells (bottom panel). The cells were fixed with 4% paraformaldehyde (10 min) permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab207434 at 1/100 dilution and ab7291 (Tubulin) at 1/1000 dilution overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor® 594) (ab150120) at 2 μg/ml (shown in pseudo color red). Nuclear DNA was labelled in blue with DAPI.
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
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All lanes : Anti-ATF3 antibody [EPR19488] - ChIP Grade (ab207434) at 1/1000 dilution
Lane 1 : Wild-type HAP1 whole cell lysate
Lane 2 : ATF3 knockout HAP1 whole cell lysate
Lane 3 : HepG2 whole cell lysate
Lysates/proteins at 20 µg per lane.
Predicted band size: 21 kDaLanes 1 - 3: Merged signal (red and green). Green - ab207434 observed at 21 kDa. Red - loading control, ab9484, observed at 37 kDa.
ab207434 was shown to specifically react with ATF3 in wild-type HAP1 cells as signal was lost in ATF3 knockout cells. Wild-type and ATF3 knockout samples were subjected to SDS-PAGE. ab207434 and ab9484 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging. -
Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus) cells labeling ATF3 with ab207434 at 1/100 dilution, followed by Goat Anti-Rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing increased nuclear staining on RAW 264.7 cell line, after treatment with LPS (1µg/ml) for 2 hours.
The nuclear counter stain is DAPI (blue). Tubulin is detected with with Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) (ab150120) secondary antibody at 1/1000 dilution (red).
The negative controls are as follows:
-ve control 1: ab207434 at 1/100 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) (ab150120) secondary antibody at 1/1000 dilution.
-ve control 2: with Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution. -
All lanes : Anti-ATF3 antibody [EPR19488] - ChIP Grade (ab207434) at 1/1000 dilution
Lane 1 : Wild-type HCT116 cell lysate
Lane 2 : ATF3 knockout HCT116 cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 21 kDa
Observed band size: 21 kDaLanes 1-2: Merged signal (red and green). Green - ab207434 observed at 21 kDa. Red - loading control ab8245 observed at 37 kDa.
ab207434 Anti-ATF3 antibody [EPR19488] - ChIP Grade was shown to specifically react with ATF3 in wild-type HCT116 cells. Loss of signal was observed when knockout cell line ab266872 (knockout cell lysate ab257074) was used. Wild-type and ATF3 knockout samples were subjected to SDS-PAGE. ab207434 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 1000 and 1 in 20000 dilution respectively. Blots were developed 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 in 20000 dilution for 1 hour at room temperature before imaging.
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All lanes : Anti-ATF3 antibody [EPR19488] - ChIP Grade (ab207434) at 1/1000 dilution
Lane 1 : Wild-type A549 cell lysate
Lane 2 : ATF3 knockout A549 cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 21 kDa
Observed band size: 21 kDaLanes 1-2: Merged signal (red and green). Green - ab207434 observed at 21 kDa. Red - loading control ab8245 observed at 37 kDa.
ab207434 Anti-ATF3 antibody [EPR19488] - ChIP Grade was shown to specifically react with ATF3 in wild-type A549 cells. Loss of signal was observed when knockout cell line ab266955 (knockout cell lysate ab257075) was used. Wild-type and ATF3 knockout samples were subjected to SDS-PAGE. ab207434 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 1000 and 1 in 20000 dilution respectively. Blots were developed 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 in 20000 dilution for 1 hour at room temperature before imaging.
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Chromatin was prepared from Hela cells according to the Abcam Dual X-ChIP protocol*. Cells were fixed with EGS for 30 minutes, then formaldehyde for 10 minutes.
The ChIP was performed with 25 µg of chromatin, 5 µg of ab207434 (red), and 20 µl of Protein A/G sepharose beads. 5 µg of rabbit normal IgG was added to the beads control (gray). 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.
*http://www.abcam.com/resources?keywords=X%20ChIP%20protocol -
All lanes : Anti-ATF3 antibody [EPR19488] - ChIP Grade (ab207434) at 1/500 dilution
Lane 1 : 293T (Human embryonic kidney epithelial cell) whole cell lysate
Lane 2 : Human liver tissue lysate
Lane 3 : Raw264.7 (Mouse abelson murine leukemia virus-induced tumor macrophage) whole cell lysate
Lane 4 : Mouse liver tissue lysate
Lane 5 : MEF (Mouse embryonic fibroblast (immortalized)) whole cell lysate
Lane 6 : Mouse heart tissue lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 21 kDa
Observed band size: 21 kDa
Exposure time: 180 secondsBlocking/Diluting buffer and concentration: 5% NFDM/TBST.
ATF3 has a low expression level in some cell lines and tissues, but is increased under treatment (PMID: 8622660, PMID: 22053207, PMID: 20018623, PMID: 29940414).
Rabbit monoclonal [EPR16891] to GAPDH (ab181602) used as loading control.
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All lanes : Anti-ATF3 antibody [EPR19488] - ChIP Grade (ab207434) at 1/1000 dilution
Lane 1 : HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate
Lane 2 : HEK-293 (Human embryonic kidney epithelial cell) whole cell lysate
Lane 3 : Jurkat (Human T cell leukemia T lymphocyte) whole cell lysate
Lane 4 : A431 (Human epidermoid carcinoma epithelial cell) whole cell lysate
Lane 5 : HepG2 (Human hepatocellular carcinoma epithelial cell) whole cell lysate
Lane 6 : LnCap (Human prostate carcinoma epithelial cell) whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 21 kDa
Observed band size: 21,23 kDa why is the actual band size different from the predicted?
Exposure time: 180 secondsBlocking/Diluting buffer and concentration 5% NFDM /TBST
ATF3 migrates as a 21 and 23 kDa doublet band due to an alternative ATG usage (PMID: 12225289, PMID: 8649793)
The mRNA and protein expression of ATF3 is low or undetectable in most cells, but its expression is rapidly induced by a large variety of cellular stresses including DNA damage, wounds, and cellular injury (PMID: 19136462, 20651982, 20592017).
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All lanes : Anti-ATF3 antibody [EPR19488] - ChIP Grade (ab207434) at 1/1000 dilution
Lane 1 : Untreated THP-1 (Human monocytic leukemia cell line) whole cell lysate
Lane 2 : THP-1 (Human monocytic leukemia cell line) treated with 80nM TPA overnight, then treated with 1 µg/ml LPS for 8 hours whole cell lysate
Lysates/proteins at 10 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution
Predicted band size: 21 kDa
Observed band size: 21 kDa
Exposure time: 3 minutesBlocking/Dilution buffer: 5% NFDM/TBST.
The expression profile observed is consistent with what has been described in the literature (PMID: 24973221).
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All lanes : Anti-ATF3 antibody [EPR19488] - ChIP Grade (ab207434) at 1/1000 dilution
Lane 1 : Untreated RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus) whole cell lysate
Lane 2 : RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus) treated with 1 µg/ml LPS for 2 hours whole cell lysate
Lysates/proteins at 10 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution
Predicted band size: 21 kDa
Observed band size: 21 kDa
Exposure time: 1 minuteBlocking/Dilution buffer: 5% NFDM/TBST.
The expression profile observed is consistent with what has been described in the literature (PMID: 24973221, PMID: 19136462).
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Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized THP-1 (Human monocytic leukemia cell line) cells labeling ATF3 with ab207434 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing increased nuclear staining on THP-1 cell line, after treatment with TPA (80nM) for overnight, followed by LPS (1µg/ml) for 8 hours.
The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) (ab150120) secondary antibody at 1/1000 dilution (red).
The negative controls are as follows:
-ve control 1: ab207434 at 1/100 dilution followed by followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) (ab150120) secondary antibody at 1/1000 dilution.
-ve control 2: Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution. -
ab207434 at 1/50 immunoprecipitating ATF3 in HeLa (human cervix adenocarcinoma) cells.
Lane 1 (input): HeLa whole cell lysate 10μg
Lane 2 (+): ab207434 + HeLa whole cell lysate
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab207434 in HeLa (human cervix adenocarcinoma) whole cell lysate
For western blotting, ab207434 (1:500) as primary antibody and VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/1000 dilution.
Blocking buffer and concentration: 5% NFDM/TBST.
Diluting buffer and concentration: 5% NFDM /TBST.
プロトコール
データシートおよび資料
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SDS download
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Datasheet download
Certificate of Compliance
参考文献 (30)
ab207434 は 30 報の論文で使用されています。
- Wang Y et al. Unveiling E2F4, TEAD1 and AP-1 as regulatory transcription factors of the replicative senescence program by multi-omics analysis. Protein Cell 13:742-759 (2022). PubMed: 35023014
- Cheng Y et al. Transcription factor network analysis identifies REST/NRSF as an intrinsic regulator of CNS regeneration in mice. Nat Commun 13:4418 (2022). PubMed: 35906210
- Kiryu-Seo S et al. Impaired disassembly of the axon initial segment restricts mitochondrial entry into damaged axons. EMBO J 41:e110486 (2022). PubMed: 36004759
- Tierney JA et al. High-fat diet causes mechanical allodynia in the absence of injury or diabetic pathology. Sci Rep 12:14840 (2022). PubMed: 36050326
- Horan NL et al. The impact of tumor immunogenicity on cancer pain phenotype using syngeneic oral cancer mouse models. Front Pain Res (Lausanne) 3:991725 (2022). PubMed: 36172037