重组Anti-CREB (phospho S133)抗体[E113] (ab32096)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [E113] to CREB (phospho S133)
- Suitable for: WB, IHC-P, IP, ICC/IF, Flow Cyt (Intra)
- Reacts with: Mouse, Rat, Human
Related conjugates and formulations
概述
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产品名称
Anti-CREB (phospho S133)抗体[E113]
参阅全部 CREB 一抗 -
描述
兔单克隆抗体[E113] to CREB (phospho S133) -
宿主
Rabbit -
特异性
This antibody is specific for CREB phosphorylated on Serine 133. The immunogen of the antibody shares 94% homology with CREB (S136) and 86% homology with ATF1 (pS63). No experiment has been performed to verify the possible cross-reactivity.
This antibody can't detect signal in mouse and rat brain related tissues.
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经测试应用
适用于: WB, IHC-P, IP, ICC/IF, Flow Cyt (Intra)more details -
种属反应性
与反应: Mouse, Rat, Human
预测可用于: Chicken, Cow, Zebrafish -
免疫原
Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
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阳性对照
- WB: A431 cell lysate; HeLa treated with anisomycin for 30 minutes ICC/IF: A431 and HeLa IHC-P: Human thyroid gland adenocarcinoma, human astrocytoma, rat cereburm and mouse cerebrum tissues. IP: HeLa treated with 25 ug/mL anisomycin for 30 minutes.
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常规说明
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.
性能
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形式
Liquid -
存放说明
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C. Stable for 12 months at -20°C. -
存储溶液
pH: 7.20
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol, 0.05% BSA -
Concentration information loading...
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纯度
Protein A purified -
克隆
单克隆 -
克隆编号
E113 -
同种型
IgG -
研究领域
相关产品
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Alternative Versions
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Assay kits
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Compatible Secondaries
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Isotype control
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Positive Controls
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Recombinant Protein
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Related Products
应用
The Abpromise guarantee
Abpromise™承诺保证使用ab32096于以下的经测试应用
“应用说明”部分 下显示的仅为推荐的起始稀释度;实际最佳的稀释度/浓度应由使用者检定。
应用 | Ab评论 | 说明 |
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WB | (7) |
1/5000. Predicted molecular weight: 37 kDa.
For unpurified use at 1/500. |
IHC-P | (3) |
1/100 - 1/2000. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
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IP |
1/40 - 1/100.
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ICC/IF | (1) |
1/100 - 1/250.
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Flow Cyt (Intra) |
Use at an assay dependent concentration.
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说明 |
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WB
1/5000. Predicted molecular weight: 37 kDa. For unpurified use at 1/500. |
IHC-P
1/100 - 1/2000. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol. |
IP
1/40 - 1/100. |
ICC/IF
1/100 - 1/250. |
Flow Cyt (Intra)
Use at an assay dependent concentration. |
靶标
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功能
This protein binds the cAMP response element (CRE), a sequence present in many viral and cellular promoters. CREB stimulates transcription on binding to the CRE. Transcription activation is enhanced by the TORC coactivators which act independently of Ser-133 phosphorylation. Implicated in synchronization of circadian rhythmicity. -
疾病相关
Defects in CREB1 may be a cause of angiomatoid fibrous histiocytoma (AFH) [MIM:612160]. A distinct variant of malignant fibrous histiocytoma that typically occurs in children and adolescents and is manifest by nodular subcutaneous growth. Characteristic microscopic features include lobulated sheets of histiocyte-like cells intimately associated with areas of hemorrhage and cystic pseudovascular spaces, as well as a striking cuffing of inflammatory cells, mimicking a lymph node metastasis. Note=A chromosomal aberration involving CREB1 is found in a patient with angiomatoid fibrous histiocytoma. Translocation t(2;22)(q33;q12) with CREB1 generates a EWSR1/CREB1 fusion gene that is most common genetic abnormality in this tumor type. -
序列相似性
Belongs to the bZIP family.
Contains 1 bZIP domain.
Contains 1 KID (kinase-inducible) domain. -
翻译后修饰
Stimulated by phosphorylation. Phosphorylation of both Ser-133 and Ser-142 in the SCN regulates the activity of CREB and participates in circadian rhythm generation. Phosphorylation of Ser-133 allows CREBBP binding (By similarity). Phosphorylated upon DNA damage, probably by ATM or ATR.
Sumoylated by SUMO1. Sumoylation on Lys-304, but not on Lys-285, is required for nuclear localization of this protein. Sumoylation is enhanced under hypoxia, promoting nuclear localization and stabilization. -
细胞定位
Nucleus. - Information by UniProt
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数据库链接
- Entrez Gene: 281713 Cow
- Entrez Gene: 1385 Human
- Entrez Gene: 12912 Mouse
- Entrez Gene: 81646 Rat
- Omim: 123810 Human
- SwissProt: P27925 Cow
- SwissProt: P16220 Human
- SwissProt: Q01147 Mouse
see all -
别名
- Active transcription factor CREB antibody
- cAMP response element binding protein 1 antibody
- cAMP response element binding protein antibody
see all
图片
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Immunohistochemical analysis of paraffin-embedded Rat cerebrum tissue labeling CREB with ab32096 at 1/2000 followed by a ready to use ab209101. Nuclear staining on rat cerebrum without alkaline phosphatase treatment (image A). No signal was detected when tissues were treated with alkaline phosphatase (image B). The section was incubated with ab32096 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.
Secondary antibody only control: Secondary antibody is a ready to use ab209101.
Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins.
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Immunohistochemical analysis of paraffin-embedded Mouse cerebrum tissue labeling CREB with ab32096 at 1/2000 followed by a ready to use ab209101. Nuclear staining on mouse cerebrum without alkaline phosphatase treatment (image A). No signal was detected when tissues were treated with alkaline phosphatase (image B). The section was incubated with ab32096 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.
Secondary antibody only control: Secondary antibody is a ready to use ab209101.
Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins.
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Immunohistochemical analysis of paraffin-embedded Human astrocytoma tissue labeling CREB with ab32096 at 1/2000 followed by a ready to use ab209101. Nuclear staining on human astrocytoma without alkaline phosphatase treatment (image A). No signal was detected when tissues were treated with alkaline phosphatase (image B). The section was incubated with ab32096 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.
Secondary antibody only control: Secondary antibody is a ready to use ab209101.
Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins.
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All lanes : Anti-CREB (phospho S133) antibody [E113] (ab32096) at 1/500 dilution
Lane 1 : Untreated HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysates
Lane 2 : HeLa whole cell lysates treated with anisomycin at 25ug/ml for 30 minutes
Lane 3 : HeLa whole cell lysates treated with anisomycin at 25ug/ml for 30 minutes, then the membrane was incubated with phosphatase
Lysates/proteins at 15 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 37 kDa
Observed band size: 40 kDa why is the actual band size different from the predicted?
Exposure time: 15 secondsBlocking and diluting buffer and concentration: 5% NFDM/TBST
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Immunocytochemistry/Immunofluorescence analysis of LP treated and untreated HeLa (Human epithelial cell line from cervix adenocarcinoma) labelling CREB with purified ab32096 at 1/200. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody. DAPI (blue) was used as the nuclear counterstain at a dilution of 1/200.
Confocal image showing nuclear staining on HeLa cells .The signal decreased after Lambda Protein Phosphatase treatment ( 31℃,2h).
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human thyroid carcinoma tissue labelling CREB with purified ab32096 at 1/100. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. ab97051, a goat anti-rabbit IgG H&L (HRP) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.
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Intracellular Flow Cytometry analysis of A431 (human epidermoid carcinoma) cells labeling CREB with purified ab32096 at 1/70 dilution (10µg/ml) (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. A Goat anti rabbit IgG (Alexa Fluorr® 488) (1/2000 dilution) was used as the secondary antibody. Rabbit monoclonal IgG (Black) was used as the isotype control, cells without incubation with primary antibody and secondary antibody (Blue) were used as the unlabeled control.
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ab32096 at 1/100 dilution immunoprecipitating CREB (phospho S133) in HeLa (human cervix adenocarcinoma) treated with 25ug/mL anisomycin for 30 minutes, whole cell lysate, observed at 40 kDa (lanes 1 and 2).
Lane 1 (input): HeLa treated with 25ug/mL anisomycin for 30 minutes. Whole cell lysate, 10μg.
Lane 2 (+): ab32096 + HeLa treated with 25ug/mL anisomycin for 30 minutes. Whole cell lysate.
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab32096 in treated with 25ug/mL anisomycin for 30 minutes. Whole cell lysate
For western blotting, ab32096 at 1/200 dilution followed by ab131366 VeriBlot for IP Detection Reagent (HRP) at 1/1000 for detection.
Blocking/Diluting buffer and concentration: 5% NFDM/TBST.
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All lanes : Anti-CREB (phospho S133) antibody [E113] (ab32096) at 1/200 dilution
Lane 1 : HeLa (Human cervix adenocarcinoma epithelial cell) treated with 250ng/ml anisomycin for 30 minutes whole cell lysates
Lane 2 : Mouse brain lysates
Lane 3 : Rat brain lysates
Lane 4 : Rat cerebellum lysate
Lane 5 : Mouse hippocampus lysate
Lane 6 : Rat hippocampus lysate
Lane 7 : Rat cerebral cortex lysate
Lysates/proteins at 15 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 37 kDa
Exposure time: 3 secondsBlocking and diluting buffer: 5% NFDM/TBST
This antibody can`t detect signal in mouse and rat brain related tissues.
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Immunocytochemistry/Immunofluorescence analysis of A431(human epidermoid carcinoma) cells +/- AP 37℃ 1h labelling CREB with purified ab32096 at 1/100. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/500) was used as the secondary antibody. DAPI (blue) was used as the nuclear counterstain. ab7291, a mouse anti-tubulin (1/1000) and ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/1000) were also used.
Control 1: primary antibody (1/100) and secondary antibody, ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/500).
Control 2: ab7291 (1/1000) and secondary antibody, ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/500).
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat hippocampus tissue labelling CREB with unpurified ab32096 at 1/100 dilution. Sections were subjected to antigen retrieval by autoclave prior to blocking with 8% milk for 30 minutes at 37°C. The primary antibody was diluted 1/100 with DAKO antibody diluent and incubated with the sample for 18 hours at 4°C. An LSAB-labeled Streptavidin-Biotin conjugated Goat polyclonal antibody was used undiluted as the secondary antibody.
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SK-N-SH cells were incubated at 37°C for 30 minutes with vehicle control (0 µM) and different concentrations of (S)-3,5-DHPG (ab120007). Increased expression of CREB (phospho S133) in SK-N-SH cells correlates with an increase in (S)-3,5-DHPGconcentration, as described in literature.
Whole cell lysates were prepared with RIPA buffer (containing protease inhibitors and sodium orthovanadate), 20µg of each were loaded on the gel and the WB was run under reducing conditions. After transfer the membrane was blocked for an hour using 3% milk before being incubated with unpurified ab32096 at 1/500 dilution and ab32515 at 1 µg/ml overnight at 4°C. Antibody binding was detected using an anti-rabbit antibody conjugated to HRP (ab97051) at 1/10000 dilution and visualised using ECL development solution.
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Lane 1 : Anti-CREB (phospho S133) antibody [E113] (ab32096) at 1/5000 dilution (purified)
Lane 2 : purified at 1/5000 dilution
Lane 1 : Untreated C6 cell lysate
Lane 2 : C6 treated with Calyculin A cell lysate
Lysates/proteins at 10 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/1000 dilution (Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated)
Predicted band size: 37 kDaBlocking buffer and concentration: 5% NFDM/TBST.
Diluting buffer and concentration: 5% NFDM /TBST. -
All lanes : Anti-CREB (phospho S133) antibody [E113] (ab32096) at 1/5000 dilution (purified)
Lane 1 : Untreated NIH/3T3 cell lysate
Lane 2 : NIH/3T3 treated with Calyculin A
Lysates/proteins at 10 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/1000 dilution (Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated)
Predicted band size: 37 kDaBlocking buffer and concentration: 5% NFDM/TBST.
Diluting buffer and concentration: 5% NFDM /TBST. -
All lanes : Anti-CREB (phospho S133) antibody [E113] (ab32096) at 1/5000 dilution (purified)
Lane 1 : Untreated HeLa cell lysate
Lane 2 : HeLa treated with Calyculin A cell lysate
Lysates/proteins at 10 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/1000 dilution (Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated)
Predicted band size: 37 kDaBlocking buffer and concentration: 5% NFDM/TBST.
Diluting buffer and concentration: 5% NFDM /TBST. -
SK-N-SH cells were incubated at 37°C for 30 minutes with vehicle control (0 µM) and different concentrations of (R,S)-CHPG (ab120039). Increased expression of CREB (phospho S133) in SK-N-SH cells correlates with an increase in (R,S)-CHPG concentration, as described in literature.
Whole cell lysates were prepared with RIPA buffer (containing protease inhibitors and sodium orthovanadate), 20µg of each were loaded on the gel and the WB was run under reducing conditions. After transfer the membrane was blocked for an hour using 3% milk before being incubated with unpurified ab32096 at 1/500 dilution and ab32515 at 1 µg/ml overnight at 4°C. Antibody binding was detected using an anti-rabbit antibody conjugated to HRP (ab97051 ) at 1/10000 dilution and visualised using ECL development solution.
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ab32096 (purified) at 1/50 immunoprecipitating CREB (phospho S133) in HeLa whole cell lysate. 10 ug of cell lysate was present in the input. For western blotting, a HRP-conjugated Veriblot for IP Detection Reagent (ab131366) (1/1,500) was used for detection. A rabbit monoclonal IgG (ab172730) was used intead of ab128913 as a negative control (Lane 3).
Blocking buffer and concentration: 5% NFDM/TBST.
Diluting buffer and concentration: 5% NFDM /TBST.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human thyroid gland adenocarcinoma tissue labelling CREB with unpurified ab32096 at 1/250 dilution.
Panel A: Cells are untreated.
Panel B: Cells are treated with Phosphatase. -
Immunocytochemistry/Immunofluorescence analysis of A431 cells labelling CREB with unpurified ab32096 at 1/250.
Panel A: Cells are untreated.
Panel B: Cells are treated with Phosphatase.
实验方案
To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.
数据表及文件
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SDS download
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Datasheet download
文献 (198)
ab32096 被引用在 198 文献中.
- Iwasa K et al. GPR137 Inhibits Cell Proliferation and Promotes Neuronal Differentiation in the Neuro2a Cells. Neurochem Res 48:996-1008 (2023). PubMed: 36436172
- Liu B et al. BHF177 Suppresses Diabetic Neuropathic Pain by Blocking PKC/CaMKII/ERK1/2/CREB Signaling Pathway through Activating GABAB Receptor. Oxid Med Cell Longev 2022:4661519 (2022). PubMed: 36439691
- Yan J et al. TREM2 activation alleviates neural damage via Akt/CREB/BDNF signalling after traumatic brain injury in mice. J Neuroinflammation 19:289 (2022). PubMed: 36463233
- Yuan P et al. Proteomics reveals the potential mechanism of Tanshinone IIA in promoting the Ex Vivo expansion of human bone marrow mesenchymal stem cells. Regen Ther 21:560-573 (2022). PubMed: 36475023
- Cai B et al. LncEDCH1 improves mitochondrial function to reduce muscle atrophy by interacting with SERCA2. Mol Ther Nucleic Acids 27:319-334 (2022). PubMed: 35024244