Recombinant Anti-Eph receptor B4/HTK antibody [EPR23221-54] (ab254300)
![Western blot - Anti-Eph receptor B4/HTK antibody [EPR23221-54] (ab254300)](https://www.abcam.com/ps/products/254/ab254300/Images/ab254300-395537-anti-eph-receptor-b4htk-antibody-epr23221-54-western-blot-human-knockout-lysate.jpg "Western blot - Anti-Eph receptor B4/HTK antibody [EPR23221-54] (ab254300)")
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR23221-54] to Eph receptor B4/HTK
- Suitable for: WB, IP
- Knockout validated
- Reacts with: Human
Related conjugates and formulations
Overview
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Product name
Anti-Eph receptor B4/HTK antibody [EPR23221-54]
See all Eph receptor B4/HTK primary antibodies -
Description
Rabbit monoclonal [EPR23221-54] to Eph receptor B4/HTK -
Host species
Rabbit -
Tested applications
Suitable for: WB, IPmore details
Unsuitable for: Flow Cyt,ICC/IF or IHC-P -
Species reactivity
Reacts with: Human -
Immunogen
Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
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Positive control
- WB: HEK293T, HT-29, HUVEC, HCT 116 and MCF7 lysates. IP: HT-29 and HUVEC whole cell lysates.
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General notes
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.
Properties
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Form
Liquid -
Storage instructions
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. -
Storage buffer
pH: 7.2
Preservative: 0.01% Sodium azide
Constituents: PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA -
Concentration information loading... -
Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EPR23221-54 -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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Compatible Secondaries
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Isotype control
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KO cell lines
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KO cell lysates
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab254300 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
| Application | Abreviews | Notes |
|---|---|---|
| WB |
1/1000. Detects a band of approximately 120 kDa (predicted molecular weight: 108 kDa).
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| IP |
1/30.
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| Notes |
|---|
|
WB
1/1000. Detects a band of approximately 120 kDa (predicted molecular weight: 108 kDa). |
|
IP
1/30. |
Target
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Function
Receptor for members of the ephrin-B family. Binds to ephrin-B2. May have a role in events mediating differentiation and development. -
Tissue specificity
Abundantly expressed in placenta and in a range of primary tissues and malignant cell lines. Expressed in fetal, but not adult, brain, and in primitive and myeloid, but not lymphoid, hematopoietic cells. -
Sequence similarities
Belongs to the protein kinase superfamily. Tyr protein kinase family. Ephrin receptor subfamily.
Contains 2 fibronectin type-III domains.
Contains 1 protein kinase domain.
Contains 1 SAM (sterile alpha motif) domain. -
Post-translational
modificationsAutophosphorylated. -
Cellular localization
Membrane. - Information by UniProt
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Database links
- Entrez Gene: 2050 Human
- Omim: 600011 Human
- SwissProt: P54760 Human
- Unigene: 437008 Human
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Alternative names
- Ephb4 antibody
- EPHB4_HUMAN antibody
- Ephrin receptor EphB4 antibody
see all
Images
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Western blot - Anti-Eph receptor B4/HTK antibody [EPR23221-54] (ab254300)All lanes : Anti-Eph receptor B4/HTK antibody [EPR23221-54] (ab254300) at 1/1000 dilution
Lane 1 : Wild-type HEK293T cell lysate
Lane 2 : EPHB4 knockout HEK293T cell lysate
Lane 3 : MCF7 cell lysate
Lane 4 : HT-29 cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) at 1/10000 dilution
Predicted band size: 108 kDa
Observed band size: 125 kDa why is the actual band size different from the predicted?Lanes 1-4: Merged signal (red and green). Green - ab254300 observed at 125 kDa. Red - loading control ab8245 observed at 36 kDa.
ab254300 Anti-Eph receptor B4/HTK antibody [EPR23221-54] was shown to specifically react with Eph receptor B4/HTK in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab266733 (knockout cell lysate ab257429) was used. Wild-type and Eph receptor B4/HTK knockout samples were subjected to SDS-PAGE. ab254300 and 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 (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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![Immunoprecipitation - Anti-Eph receptor B4/HTK antibody [EPR23221-54] (ab254300)](https://www.abcam.com/ps/products/254/ab254300/Images/ab254300-2-anti-eph-receptor-b4-antibody-epr23221-54-immunoprecipitation-huvec-human.jpg)
Immunoprecipitation - Anti-Eph receptor B4/HTK antibody [EPR23221-54] (ab254300)Eph receptor B4 was immunoprecipitated from 0.35 mg HUVEC (human umbilical vein endothelial cell) whole cell lysate with ab254300 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab254300 at 1/500 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366) was used at 1/1000 dilution.
Lane 1: HUVEC (human umbilical vein endothelial cell) whole cell lysate 10ug
Lane 2: ab254300 IP in HUVEC whole cell lysate
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab254300 in HUVEC whole cell lysate
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 8 seconds.
Cleaved intracellular domain (ICD) was observed at around 47kDa. (PMID: 24854540).
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![Western blot - Anti-Eph receptor B4/HTK antibody [EPR23221-54] (ab254300)](https://www.abcam.com/ps/products/254/ab254300/Images/ab254300-3-anti-eph-receptor-b4-antibody-epr23221-54-western-human.jpg)
Western blot - Anti-Eph receptor B4/HTK antibody [EPR23221-54] (ab254300)All lanes : Anti-Eph receptor B4/HTK antibody [EPR23221-54] (ab254300) at 1/1000 dilution
Lane 1 : HT-29 (human colorectal adenocarcinoma epithelial cell), whole cell lysate
Lane 2 : HUVEC (human umbilical vein endothelial cell), whole cell lysate
Lane 3 : SW620 (human colorectal adenocarcinoma epithelial cell), whole cell lysate
Lane 4 : HCT 116 (human colorectal carcinoma epithelial cell), whole cell lysate
Lane 5 : MCF7 (human breast adenocarcinoma epithelial cell), whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution
Predicted band size: 108 kDa
Observed band size: 120 kDa why is the actual band size different from the predicted?Blocking and dilution buffer: 5% NFDM/TBST.
Exposure time: 37 seconds.
Negative control: SW620 (PMID: 19366806).
EphB4 is negative in SW620 and positive in HT29 and HUVEC, which is different from expression profiles of other EphB family members.
Cleaved intracellular domain (ICD) was observed at around 47kDa. (PMID: 24854540).
The expression profile observed is consistent with what has been described in the literature (PMID: 19366806, 16840724).
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![Immunoprecipitation - Anti-Eph receptor B4/HTK antibody [EPR23221-54] (ab254300)](https://www.abcam.com/ps/products/254/ab254300/Images/ab254300-1-anti-eph-receptor-b4-antibody-epr23221-54-immunoprecipitation-ht29-human.jpg)
Immunoprecipitation - Anti-Eph receptor B4/HTK antibody [EPR23221-54] (ab254300)Eph receptor B4 was immunoprecipitated from 0.35 mg HT-29 (human colorectal adenocarcinoma epithelial cell) whole cell lysate with ab254300 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab254300 at 1/500 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366) was used at 1/1000 dilution.
Lane 1: HT-29 (human colorectal adenocarcinoma epithelial cell) whole cell lysate 10ug
Lane 2: ab254300 IP in HT-29 whole cell lysate
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab254300 in HT-29 whole cell lysate
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 8 seconds.
Cleaved intracellular domain (ICD) was observed at around 47kDa. (PMID: 24854540).
-
![Anti-Eph receptor B4/HTK antibody [EPR23221-54] (ab254300)](https://www.abcam.com/ps/products/254/ab254300/Images/ab254300-5-benefits-of-recombinant-antibodies.png)
Anti-Eph receptor B4/HTK antibody [EPR23221-54] (ab254300)
Protocols
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.
Datasheets and documents
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Datasheet download
Certificate of Compliance
References (0)
ab254300 has not yet been referenced specifically in any publications.
