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AB266733

Human EPHB4 (Eph receptor B4/HTK) knockout HEK-293T cell line

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EPHB4 KO cell line available to order. KO validated by Western blot. Free of charge wild type control available. Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp insertion in exon 3. To order both knockout and wild-type control cells: select '2 x 1000000 Cells/vial'. To order only knockout cells: select '1000000 Cells/vial'.
5 Images
Immunocytochemistry/ Immunofluorescence - Human EPHB4 (Eph receptor B4/HTK) knockout HEK-293T cell line (AB266733)
  • ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Human EPHB4 (Eph receptor B4/HTK) knockout HEK-293T cell line (AB266733)
Western blot - Human EPHB4 (Eph receptor B4/HTK) knockout HEK-293T cell line (AB266733)
  • WB

Unknown

Western blot - Human EPHB4 (Eph receptor B4/HTK) knockout HEK-293T cell line (AB266733)

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° 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.

All lanes:

Western blot - Anti-Eph receptor B4/HTK antibody [EPR23221-54] (<a href='/en-us/products/primary-antibodies/eph-receptor-b4-htk-antibody-epr23221-54-ab254300'>ab254300</a>) at 1/1000 dilution

Lane 1:

Wild-type HEK293T cell lysate at 20 µg

Lane 2:

EPHB4 knockout HEK293T cell lysate at 20 µg

Lane 2:

Western blot - Human EPHB4 (Eph receptor B4/HTK) knockout HEK-293T cell line (ab266733)

Lane 3:

MCF7 cell lysate at 20 µg

Lane 4:

HT-29 cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/10000 dilution

Predicted band size: 108 kDa

Observed band size: 125 kDa

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Western blot - Human EPHB4 (Eph receptor B4/HTK) knockout HEK-293T cell line (AB266733)
  • WB

Lab

Western blot - Human EPHB4 (Eph receptor B4/HTK) knockout HEK-293T cell line (AB266733)

Lanes 1-4 : Merged signal (red and green). Green - ab254301 observed at 125 kDa. Red - loading control ab8245 observed at 36 kDa.

ab254301 Anti-Eph receptor B4/HTK antibody [EPR23222-24] 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. ab254301 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4° 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.

All lanes:

Western blot - Anti-Eph receptor B4/HTK antibody [EPR23222-24] (<a href='/en-us/products/primary-antibodies/eph-receptor-b4-htk-antibody-epr23222-24-ab254301'>ab254301</a>) at 1/1000 dilution

Lane 1:

Wild-type HEK293T cell lysate at 20 µg

Lane 2:

EPHB4 knockout HEK293T cell lysate at 20 µg

Lane 2:

Western blot - Human EPHB4 (Eph receptor B4/HTK) knockout HEK-293T cell line (ab266733)

Lane 3:

MCF7 cell lysate at 20 µg

Lane 4:

HT-29 cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/10000 dilution

Predicted band size: 108 kDa

Observed band size: 125 kDa

false

Cell Culture - Human EPHB4 (Eph receptor B4/HTK) knockout HEK-293T cell line (AB266733)
  • Cell Culture

Lab

Cell Culture - Human EPHB4 (Eph receptor B4/HTK) knockout HEK-293T cell line (AB266733)

Representative images EPHB4 knockout HEK293T cells, low and high confluency examples (top left and right respectively) and wild-type HEK293T cells, low and high confluency (bottom left and right respectively) showing typical adherent, epithelial-like morphology. Images were captured at 10X magnification using a EVOS M5000 microscope.

Sanger Sequencing - Human EPHB4 (Eph receptor B4/HTK) knockout HEK-293T cell line (AB266733)
  • Sanger seq

Unknown

Sanger Sequencing - Human EPHB4 (Eph receptor B4/HTK) knockout HEK-293T cell line (AB266733)

Homozygous : 1 bp insertion in exon3

Key facts

Cell type

HEK-293T

Species or organism

Human

Tissue

Kidney

Form

Liquid

form

Knockout validation

Sanger Sequencing,Western blot

Mutation description

Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp insertion in exon 3

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Complementary Products

Cell Lines & Lysates

AB257429

Human EPHB4 (Eph receptor B4/HTK) knockout HEK-293T cell lysate

Western blot - Human EPHB4 (Eph receptor B4/HTK) knockout HEK-293T cell lysate (AB257429)

  • 0
  • 0
Primary Antibodies

AB254301

Anti-Eph receptor B4/HTK antibody [EPR23222-24]

BOND RX™ Validated|RabMAb|Recombinant|KO Validated

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Eph receptor B4/HTK antibody [EPR23222-24] (AB254301)

  • 0
  • 0
Proteins & Peptides

AB217407

Recombinant human Vitronectin/S-Protein (Animal Free)

  • 0
  • 0
Proteins & Peptides

AB313361

Recombinant Human WFDC2 Protein (His tag)

Mass Spectrometry - Recombinant Human WFDC2 Protein (His tag) (AB313361)

  • 0
  • 0
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Reactivity data

{ "title": "Reactivity Data", "filters": { "stats": ["", "Reactivity", "Dilution Info", "Notes"] }, "values": { "WB": { "reactivity":"TESTED_AND_REACTS", "dilution-info":"", "notes":"<p></p>" } } }
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Product details

We will provide viable cells that proliferate on revival.

This product is subject to limited use licenses from The Broad Institute, ERS Genomics Limited and Sigma-Aldrich Co. LLC, and is developed with patented technology. For full details of the licenses and patents please refer to our limited use license and patent pages.

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What's included?

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Properties and storage information

Gene name
EPHB4
Gene editing type
Knockout
Gene editing method
CRISPR technology
Knockout validation
Sanger Sequencing, Western blot
Zygosity
Homozygous
Shipped at conditions
Dry Ice
Appropriate short-term storage conditions
-196°C
Appropriate long-term storage conditions
-196°C
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Handling procedures

Initial handling guidelines

Upon arrival, the vial should be stored in liquid nitrogen vapor phase and not at -80°C. Storage at -80°C may result in loss of viability.

1. Thaw the vial in 37°C water bath for approximately 1-2 minutes.
2. Transfer the cell suspension (0.8 mL) to a 15 mL/50 mL conical sterile polypropylene centrifuge tube containing 8.4 mL pre-warmed culture medium, wash vial with an additional 0.8 mL culture medium (total volume 10 mL) to collect remaining cells, and centrifuge at 201 x g (rcf) for 5 minutes at room temperature. 10 mL represents minimum recommended dilution. 20 mL represents maximum recommended dilution.
3. Resuspend the cell pellet in 5 mL pre-warmed culture medium and count using a haemocytometer or alternative cell counting method seed all remaining cells into a T25.
4. Incubate the culture at 37°C incubator with 5% CO2. Check the culture one day after revival and continue to check until 80% confluent. Media change can be given if needed.
5. Once confluent passage into an appropriate flask at a density of 2x104 cells/cm2. Seeding density is given as a guide only and should be scaled to align with individual lab schedules. Cultures should be monitored daily.

Subculture guidelines
  • All seeding densities should be based on cell counts gained by established methods.
  • A guide seeding density of 2x104 cells/cm2 is recommended.
  • Cells should be passaged when they have achieved 80-90% confluence.
Culture medium

DMEM (High Glucose) + 10% FBS

Cryopreservation medium

Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.

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Supplementary information

This supplementary information is collated from multiple sources and compiled automatically.

The Eph receptor B4 also known as EphB4 or HTK is a protein belonging to the Ephrin receptor subfamily of receptor tyrosine kinases. The receptor has a molecular weight of about 120 kDa. EphB4 typically interacts with its binding partner ephrin-B2 facilitating bidirectional signaling. These complexes reside in the cell membrane and are mainly found in vascular endothelial cells and various tissues including the heart lungs and skin. Notably this receptor plays a role in cellular processes like cell migration and adhesion.
Biological function summary

The Eph receptor B4 is a significant regulator of angiogenesis and vascular development. It forms a critical component of a signaling complex that also includes ephrin-B2. During embryonic development EphB4's interaction with ephrin-B2 guides the proper formation and organization of blood vessels. In adult organisms it remains active in maintaining vascular homeostasis. Additionally this receptor impacts neuronal development and repair through modulating axon guidance and synaptic plasticity.

Pathways

EphB4 engages in various cellular signaling cascades. It features prominently within pathways governing angiogenesis and cell adhesion. EphB4 acts as a central player in the VEGF signaling pathway modulating endothelial cell behavior. The protein's pathways frequently intersect with other proteins such as FAK and Src influencing cell motility and morphology. Furthermore in the ephrin receptor pathway it cross-talks with other Eph receptors like EphA4 assisting in diverse biological functions.

Abnormal Eph receptor B4 activity associates with several pathological conditions. Dysregulation often links to cancer due to its role in neovascularization and tumor progression. Overexpression of EphB4 in tumors like breast and colorectal cancer suggests a contribution to malignancy by supporting angiogenesis and tumor growth. In vascular disorders impaired EphB4-ephrin-B2 signaling may lead to defects in blood vessel formation. Additionally alterations in this receptor are observed in neurological diseases such as Alzheimer's where EphB4 interacts with tau protein-modifying vascular and neuronal deficiencies.
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Quality control

STR analysis

CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX

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Cell culture

Biosafety level

EU: 2 US: 2

Adherent/suspension

Adherent

Gender

Female

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Product protocols

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Target data

See full target information EPHB4
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Product promise

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For full details, please see our Terms & Conditions

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