Knockout Tested Rabbit Recombinant Monoclonal A RAF antibody. Suitable for WB, ICC/IF, Flow Cyt (Intra), IHC-P and reacts with Human samples. Cited in 8 publications.
IgG
Rabbit
pH: 7.2 - 7.4
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA
Liquid
Monoclonal
WB | ICC/IF | Flow Cyt (Intra) | IHC-P | |
---|---|---|---|---|
Human | Tested | Tested | Tested | Tested |
Mouse | Not recommended | Not recommended | Not recommended | Not recommended |
Rat | Not recommended | Not recommended | Not recommended | Not recommended |
Species | Dilution info | Notes |
---|---|---|
Species Human | Dilution info 1/1000 | Notes Perform heat-mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol. |
Species | Dilution info | Notes |
---|---|---|
Species Mouse, Rat | Dilution info - | Notes - |
Species | Dilution info | Notes |
---|---|---|
Species Human | Dilution info 1/250 | Notes - |
Species | Dilution info | Notes |
---|---|---|
Species Mouse, Rat | Dilution info - | Notes - |
Species | Dilution info | Notes |
---|---|---|
Species Human | Dilution info 1/100 | Notes ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody. |
Species | Dilution info | Notes |
---|---|---|
Species Mouse, Rat | Dilution info - | Notes - |
Species | Dilution info | Notes |
---|---|---|
Species Human | Dilution info 1/1000 | Notes Perform heat-mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol. |
Species | Dilution info | Notes |
---|---|---|
Species Mouse | Dilution info - | Notes Perform heat-mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol. |
Species Rat | Dilution info - | Notes - |
Select an associated product type
Involved in the transduction of mitogenic signals from the cell membrane to the nucleus. May also regulate the TOR signaling cascade. Phosphorylates PFKFB2 (PubMed:36402789).Isoform 2Serves as a positive regulator of myogenic differentiation by inducing cell cycle arrest, the expression of myogenin and other muscle-specific proteins, and myotube formation.
ARAF1, PKS, PKS2, ARAF, Serine/threonine-protein kinase A-Raf, Proto-oncogene A-Raf, Proto-oncogene A-Raf-1, Proto-oncogene Pks
Knockout Tested Rabbit Recombinant Monoclonal A RAF antibody. Suitable for WB, ICC/IF, Flow Cyt (Intra), IHC-P and reacts with Human samples. Cited in 8 publications.
IgG
Rabbit
pH: 7.2 - 7.4
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA
Liquid
Monoclonal
EPR16208
Affinity purification Protein A
Blue Ice
1-2 weeks
+4°C
-20°C
Upon delivery aliquot
Avoid freeze / thaw cycle
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
This product is a recombinant monoclonal antibody, which offers several advantages including:
For more information, read more on recombinant antibodies.
This supplementary information is collated from multiple sources and compiled automatically.
The A-Raf protein also called ARAF is a serine/threonine-protein kinase with a molecular mass of about 68 kDa. Scientists frequently refer to it with names like A-Raf or anti A-Raf when discussing its function. A-Raf is expressed in various tissues with higher levels found in the urogenital system and the central nervous system. This protein plays an important role in cellular signal transduction processes influencing cell proliferation and differentiation through phosphorylation activity.
The A-Raf protein participates in the regulation of cell growth and survival. It forms part of the MAPK/ERK signaling cascade which is essential for transmitting mitogenic signals from the cell surface to the nucleus. A-Raf acts as a modulator within this complex impacting the overall signaling output. The protein is intimately involved in the regulation of the developmental processes influencing cell fate and function.
The A-Raf protein is central to the MAPK/ERK pathway a critical signaling pathway involved in the regulation of cellular responses to growth signals. This pathway includes key components like the proteins B-Raf and C-Raf with A-Raf playing specific roles in the modulation and fine-tuning of this signal transmission. It helps mediate signals that control cell division differentiation and secretion ensuring appropriate cellular responses to external stimuli.
Aberrations in A-Raf functions have been linked to cancers particularly urogenital cancers due to its role in uncontrolled cell proliferation. Faulty A-Raf activity can also contribute to neurological disorders because of its expression in the central nervous system and involvement in neuronal signaling pathways. In these contexts it can interact with proteins like B-Raf which are important for maintaining normal cellular signaling and preventing tumorigenesis and neurodegenerative disorders.
We have tested this species and application combination and it works. It is covered by our product promise.
We have not tested this specific species and application combination in-house, but expect it will work. It is covered by our product promise.
This species and application combination has not been tested, but we predict it will work based on strong homology. However, this combination is not covered by our product promise.
We do not recommend this combination. It is not covered by our product promise.
We are dedicated to supporting your work with high quality reagents and we are here for you every step of the way should you need us.
In the unlikely event of one of our products not working as expected, you are covered by our product promise.
Full details and terms and conditions can be found here:
Terms & Conditions.
Lanes 1-4: Merged signal (red and green). Green - ab200653 observed at 68 kDa. Red - loading control Anti-GAPDH antibody [6C5] - Loading Control ab8245 observed at 36 kDa.
ab200653 Anti-A-Raf antibody [EPR16208] was shown to specifically react with A-Raf in wild-type HEK-293T cells. Loss of signal was observed when knockout cell line Human A-Raf knockout HEK-293T cell line ab266351 (knockout cell lysate Human A-Raf knockout HEK-293T cell lysate ab257838) was used. Wild-type and A-Raf knockout samples were subjected to SDS-PAGE. ab200653 and Anti-GAPDH antibody [6C5] - Loading Control (Anti-GAPDH antibody [6C5] - Loading Control ab8245) were incubated 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 (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (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.
All lanes: Western blot - Anti-A-Raf antibody [EPR16208] (ab200653) at 1/1000 dilution
Lane 1: Wild-type HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate at 20 µg
Lane 2: A-Raf knockout HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate at 20 µg
Lane 3: HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 20 µg
Lane 4: Raji (Human Burkitts lymphoma cell line) whole cell lysate at 20 µg
All lanes: Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) at 1/10000 dilution
Predicted band size: 68 kDa
Observed band size: 68 kDa
This data was developed using ab200653, the same antibody clone in a different buffer formulation.
Lanes 1-4: Merged signal (red and green). Green - ab200653 observed at 68 kDa. Red - loading control Anti-GAPDH antibody [6C5] - Loading Control ab8245 observed at 36 kDa.
ab200653 Anti-A-Raf antibody [EPR16208] was shown to specifically react with A-Raf in wild-type HEK-293T cells. Loss of signal was observed when knockout cell line Human A-Raf knockout HEK-293T cell line ab266351 (knockout cell lysate Human A-Raf knockout HEK-293T cell lysate ab257838) was used. Wild-type and A-Raf knockout samples were subjected to SDS-PAGE. ab200653 and Anti-GAPDH antibody [6C5] - Loading Control (Anti-GAPDH antibody [6C5] - Loading Control ab8245) were incubated 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 (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (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.
Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cells from cervix adenocarcinoma) cells labeling A-Raf with ab200653 at 1/250 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077) secondary antibody at 1/500 dilution (green).
Cytoplasm staining on HeLa cell line is observed.
The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/500 dilution (red).
The negative controls are as follows:
-ve control 1: ab200653 at 1/250 dilution followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/500 dilution.
-ve control 2: Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077 (Alexa Fluor®488 Goat Anti-Rabbit IgG H&L) at 1/500 dilution.
Immunohistochemical analysis of paraffin-embedded Human cervix carcinoma tissue labeling A-Raf with ab200653 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) secondary antibody at 1/500 dilution.
Cytoplasm staining on Human cervix carcinoma tissue 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) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/500 dilution.
Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Lane 1: Wild-type HAP1 cell lysate (20 μg)
Lane 2:A-Raf knockout HAP1 cell lysate (20 μg)
Lanes 1 - 2: Merged signal (red and green). Green - ab200653 observed at 68 kDa. Red - loading control, Anti-GAPDH antibody [6C5] - Loading Control ab8245, observed at 37 kDa.
This western blot image is a comparison between ab200653 and a competitor's top cited rabbit polyclonal antibody.
All lanes: Western blot - Anti-A-Raf antibody [EPR16208] (ab200653)
Predicted band size: 68 kDa
Lane 1: Wild-type HAP1 cell lysate (20 μg)
Lane 2: A-Raf knockout HAP1 cell lysate (20 μg)
Lane 3: HeLa cell lysate (20 μg)
Lane 4: Raji cell lysate (20 μg)
Lanes 1 - 4: Merged signal (red and green). Green - ab200653 observed at 68 kDa. Red - loading control, Anti-GAPDH antibody [6C5] - Loading Control ab8245, observed at 37 kDa.
ab200653 was shown to specifically react with A-Raf when A-Raf knockout samples were used. Wild-type and ProteinX knockout samples were subjected to SDS-PAGE. ab200653 and Anti-GAPDH antibody [6C5] - Loading Control ab8245 (loading control to GAPDH) were diluted 1/1000 and 1/2000 incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and) Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed (Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed ab216772) secondary antibodies at 1/10000 dilution for 1 h at room temperature before imaging.
All lanes: Western blot - Anti-A-Raf antibody [EPR16208] (ab200653)
Predicted band size: 68 kDa
Intracellular flow cytometric analysis of 2% paraformaldehyde-fixed HeLa (Human epithelial cells from cervix adenocarcinoma) cells labeling A-Raf with ab200653 at 1/100 dilution (red) compared with a rabbit monoclonal IgG isotype control (black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody; blue). Goat anti rabbit IgG (FITC) at 1/150 dilution was used as the secondary antibody.
Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HEK293 (Human embryonic kidney) cells labeling A-Raf with ab200653 at 1/250 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077) secondary antibody at 1/500 dilution (green).
Cytoplasm staining on HEK293 cell line is observed.
The nuclear counter stain is DAPI (blue).
Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/500 dilution (red).
The negative controls are as follows:
-ve control 1: ab200653 at 1/250 dilution followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/500 dilution.
-ve control 2: Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077 (Alexa Fluor®488 Goat Anti-Rabbit IgG H&L) at 1/500 dilution.
Blocking/Dilution buffer: 5% NFDM/TBST.
All lanes: Western blot - Anti-A-Raf antibody [EPR16208] (ab200653) at 1/10000 dilution
Lane 1: HeLa (Human epithelial cells from cervix adenocarcinoma) whole cell lysate at 20 µg
Lane 2: Raji (Human Burkitt's lymphoma cell line) whole cell lysate at 20 µg
All lanes: Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/1000 dilution
Predicted band size: 68 kDa
Observed band size: 68 kDa
Exposure time: 1min
Blocking/Dilution buffer: 5% NFDM/TBST.
All lanes: Western blot - Anti-A-Raf antibody [EPR16208] (ab200653) at 1/1000 dilution
Lane 1: HT-29 (Human colorectal adenocarcinoma cells) whole cell lysate at 10 µg
Lane 2: A375 (Human malignant melanoma) whole cell lysate at 10 µg
Lane 3: Human fetal heart lysate at 10 µg
Lane 4: Human fetal kidney lysate at 10 µg
Lane 5: Human bladder lysate at 10 µg
All lanes: Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/1000 dilution
Predicted band size: 68 kDa
Observed band size: 68 kDa
Exposure time: 1min
Blocking/Dilution buffer: 5% NFDM/TBST.
All lanes: Western blot - Anti-A-Raf antibody [EPR16208] (ab200653) at 1/1000 dilution
All lanes: Human fetal brain lysate at 10 µg
All lanes: Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/1000 dilution
Predicted band size: 68 kDa
Observed band size: 68 kDa
Exposure time: 3min
Immunohistochemical analysis of paraffin-embedded Human kidney tissue labeling A-Raf with ab200653 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) secondary antibody at 1/500 dilution.
Cytoplasm staining on Human kidney tissue 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) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/500 dilution.
Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Anti-A-Raf antibody [EPR16208] staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (Anti-GAPDH antibody [6C5] - Loading Control ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab200653 was shown to bind specifically to A-Raf. A band was observed at 67 kDa in wild-type HCT 116 cell lysates with no signal observed at this size in A-Raf knockout cell line Human A-Raf knockout HCT116 cell line ab286752. To generate this image, wild-type and A-Raf 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: Western blot - Anti-A-Raf antibody [EPR16208] (ab200653) at 1/1000 dilution
Lane 1: Wild-type HCT 116 cell lysate at 20 µg
Lane 2: A-Raf knockout HCT 116 cell lysate at 20 µg
Lane 3: HEK-293 cell lysate at 20 µg
Lane 4: Raji cell lysate at 20 µg
All lanes: Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution
Performed under reducing conditions.
Predicted band size: 68 kDa
Observed band size: 67 kDa
Please note: All products are 'FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC OR THERAPEUTIC PROCEDURES'.
For licensing inquiries, please contact partnerships@abcam.com