重组Anti-Raf1抗体[EP4969] - N-terminal (ab181115)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EP4969] to Raf1 - N-terminal
- Suitable for: Flow Cyt (Intra), WB, ICC/IF
- Knockout validated
- Reacts with: Mouse, Rat, Human
Related conjugates and formulations
概述
-
产品名称
Anti-Raf1抗体[EP4969] - N-terminal
参阅全部 Raf1 一抗 -
描述
兔单克隆抗体[EP4969] to Raf1 - N-terminal -
宿主
Rabbit -
经测试应用
适用于: Flow Cyt (Intra), WB, ICC/IFmore details -
种属反应性
与反应: Mouse, Rat, Human -
免疫原
Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
-
阳性对照
- WB: HAP1, HeLa, K-562, RAW 264.7, C2C12, NIH/3T3, and PC-12 lysates. ICC/IF: HeLa cells.
-
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
-
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 long term. Avoid freeze / thaw cycle. -
存储溶液
Preservative: 0.01% Sodium azide
Constituents: PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA -
Concentration information loading...
-
纯度
Protein A purified -
克隆
单克隆 -
克隆编号
EP4969 -
同种型
IgG -
研究领域
相关产品
-
Alternative Versions
-
Isotype control
-
KO cell lines
-
KO cell lysates
-
Positive Controls
-
Recombinant Protein
应用
The Abpromise guarantee
Abpromise™承诺保证使用ab181115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 |
---|---|---|
Flow Cyt (Intra) |
1/100.
|
|
WB |
1/1000. Detects a band of approximately 73 kDa (predicted molecular weight: 73 kDa).
|
|
ICC/IF |
1/100 - 1/250.
|
说明 |
---|
Flow Cyt (Intra)
1/100. |
WB
1/1000. Detects a band of approximately 73 kDa (predicted molecular weight: 73 kDa). |
ICC/IF
1/100 - 1/250. |
靶标
-
功能
Involved in the transduction of mitogenic signals from the cell membrane to the nucleus. Part of the Ras-dependent signaling pathway from receptors to the nucleus. Protects cells from apoptosis mediated by STK3. -
组织特异性
In skeletal muscle, isoform 1 is more abundant than isoform 2. -
疾病相关
Defects in RAF1 are the cause of Noonan syndrome type 5 (NS5) [MIM:611553]. Noonan syndrome (NS) is a disorder characterized by dysmorphic facial features, short stature, hypertelorism, cardiac anomalies, deafness, motor delay, and a bleeding diathesis. It is a genetically heterogeneous and relatively common syndrome, with an estimated incidence of 1 in 1000-2500 live births.
Defects in RAF1 are the cause of LEOPARD syndrome type 2 (LEOPARD2) [MIM:611554]. LEOPARD syndrome is an autosomal dominant disorder allelic with Noonan syndrome. The acronym LEOPARD stands for lentigines, electrocardiographic conduction abnormalities, ocular hypertelorism, pulmonic stenosis, abnormalities of genitalia, retardation of growth, and deafness. -
序列相似性
Belongs to the protein kinase superfamily. TKL Ser/Thr protein kinase family. RAF subfamily.
Contains 1 phorbol-ester/DAG-type zinc finger.
Contains 1 protein kinase domain.
Contains 1 RBD (Ras-binding) domain. -
翻译后修饰
Phosphorylated upon DNA damage, probably by ATM or ATR. Phosphorylation at Thr-269 increases its kinase activity. Phosphorylation at Ser-259 induces the interaction with YWHAZ and inactivates kinase activity. Dephosphorylation of Ser-259 by the complex containing protein phosphatase 1, SHOC2 and M-Ras/MRAS relieves inactivation, leading to stimulate RAF1 activity. -
细胞定位
Cytoplasm. Cell membrane. Colocalizes with RGS14 and BRAF in both the cytoplasm and membranes. - Information by UniProt
-
数据库链接
- Entrez Gene: 5894 Human
- Entrez Gene: 110157 Mouse
- Entrez Gene: 24703 Rat
- Omim: 164760 Human
- SwissProt: P04049 Human
- SwissProt: Q99N57 Mouse
- SwissProt: P11345 Rat
- Unigene: 159130 Human
see all -
别名
- c Raf antibody
- C-raf antibody
- C-Raf proto-oncogene, serine/threonine kinase antibody
see all
图片
-
All lanes : Anti-Raf1 antibody [EP4969] - N-terminal (ab181115) at 1/1000 dilution
Lane 1 : Wild-type HCT 116 cell lysate
Lane 2 : RAF1 knockout HCT 116 cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 73 kDa
Observed band size: 73 kDaWestern blot: Anti-RAF1 antibody [EP4969] (ab181115) staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab181115 was shown to bind specifically to RAF1. A band was observed at 73 kDa in wild-type HCT 116 cell lysates with no signal observed at this size in RAF1 knockout cell line. To generate this image, wild-type and RAF1 knockout HCT 116 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.
-
All lanes : Anti-Raf1 antibody [EP4969] - N-terminal (ab181115) at 1/20000 dilution
Lane 1 : Wild-type HeLa cell lysate
Lane 2 : RAF1 knockout HeLa cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 73 kDa
Observed band size: 73 kDaLanes 1- 2: Merged signal (red and green). Green - ab181115 observed at 73 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.
ab181115 was shown to react with Raf1 in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab264978 (knockout cell lysate ab257126) was used. Wild-type HeLa and RAF1 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. ab181115 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) overnight at 4°C at a 1 in 20000 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.
-
Immunocytochemistry/ Immunofluorescence analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling Raf1 with purified ab181115 at 1/100 dilution (10 µg/ml). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with Ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) 1/200 (2.5 µg/ml). Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) was used as the secondary antibody at 1/1000 (2 µg/ml) dilution. DAPI (blue) was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.
-
Lanes 1-2 : Anti-Raf1 antibody [EP4969] - N-terminal (ab181115) at 1/1000 dilution
Lanes 3-4 : Anti-beta Actin antibody [mAbcam 8226] - Loading Control (ab8226) at 1/1000 dilution
Lanes 1 & 3 & 5 : Wild-type HAP1 cell lysate
Lanes 2 & 4 & 6 : Raf1 knockout HAP1 cell lysate
Lysates/proteins at 20 µg per lane.
Predicted band size: 73 kDaLanes 1 and 2: Green signal from target – ab181115 observed at 74 kDa
Lanes 3 and 4: Red signal from loading control – ab8226 observed at 42 kDa
Lanes 5 and 6: Merged (red and green) signalUnpurified ab181115 was shown to specifically react with Raf1 when Raf1 knockout samples were used. Wild-type and Raf1 knockout samples were subjected to SDS-PAGE. ab181115 and ab8226 (loading control to beta actin) were both diluted 1/1000 and incubated overnight at 4ºC. 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 1/10000 dilution for 1 hour at room temperature before imaging.
-
Lanes 1: Wild-type HAP1 cell lysate (20 µg)
Lanes 2: Raf1 knockout HAP1 cell lysate (20 µg)
Lanes 1 and 2: Merged signal (red and green).Green - Target observed at 74 kDa. Red - loading control, ab8226, observed at 42 kDa
This western blot image is a comparison between unpurified ab181115 and a competitor's top cited discontinued rabbit polyclonal antibody.
-
Intracellular Flow Cytometry analysis of K-562 (Human chronic myelogenous leukemia lymphoblast) cells labelling with unpurified ab181115 at 1/100 dilution (10.79µg/mL) (red). Cells were fixed with 4% paraformaldehyde . Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) was used as the secondary antibody at 1/2000 dilution. Isotype control - 90% methanol . Unlabeled control - Rabbit monoclonal IgG (ab172730) / Black.
-
Anti-Raf1 antibody [EP4969] - N-terminal (ab181115) at 1/1000 dilution (Purified) + PC-12 (Rat adrenal gland pheochromocytoma) whole cell lysates at 20 µg
Secondary
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 73 kDa
Observed band size: 73 kDa -
All lanes : Anti-Raf1 antibody [EP4969] - N-terminal (ab181115) at 1/1000 dilution (Purified)
Lane 1 : RAW 264.7(Mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysates
Lane 2 : C2C12 (Mouse myoblasts myoblast) whole cell lysates
Lane 3 : NIH/3T3 (Mouse embryonic fibroblast) whole cell lysates
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: 73 kDa
Observed band size: 73 kDa -
All lanes : Anti-Raf1 antibody [EP4969] - N-terminal (ab181115) at 1/1000 dilution (Purified)
Lane 1 : HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysates
Lane 2 : K-562 (Human chronic myelogenous leukemia lymphoblast) whole cell lysates
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: 73 kDa
Observed band size: 73 kDa -
All lanes : Anti-Raf1 antibody [EP4969] - N-terminal (ab181115) at 1/1000 dilution ((unpurified))
Lane 1 : HeLa cell lysate
Lane 2 : K562 cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugate at 1/1000 dilution
Predicted band size: 73 kDa
Observed band size: 73 kDa -
Immunofluorescent analysis of acetone-fixed K562 cells labeling Raf1 with unpurified ab181115 at 1/250 dilution, followed by Goat anti rabbit IgG (Alexa Fluor® 488) secondary antibody at 1/200 dilution. Counter stained with Dapi.
实验方案
数据表及文件
-
SDS download
-
Datasheet download
Certificate of Compliance
文献 (5)
ab181115 被引用在 5 文献中.
- Kong D et al. Procoxacin bidirectionally inhibits osteoblastic and osteoclastic activity in bone and suppresses bone metastasis of prostate cancer. J Exp Clin Cancer Res 42:45 (2023). PubMed: 36759880
- Hou J et al. Caudatin blocks the proliferation, stemness and glycolysis of non-small cell lung cancer cells through the Raf/MEK/ERK pathway. Pharm Biol 60:764-773 (2022). PubMed: 35387566
- Li J et al. METTL3 Facilitates Tumor Progression by COL12A1/MAPK Signaling Pathway in Esophageal Squamous Cell Carcinoma. J Cancer 13:1972-1984 (2022). PubMed: 35399719
- Jin X et al. Cinobufagin Triggers Defects in Spindle Formation and Cap-Dependent Translation in Liver Cancer Cells by Inhibiting the AURKA-mTOR-eIF4E Axis. Am J Chin Med 48:651-678 (2020). PubMed: 32349518
- Youngworth I & Delany ME A Premature Stop Codon in RAF1 Is the Priority Candidate Causative Mutation of the Inherited Chicken Wingless-2 Developmental Syndrome. Genes (Basel) 10:N/A (2019). PubMed: 31075853