Human EGFR protein (extracellular domain), biotinylated, recombinant

Description

EGFR is known as epidermal growth factor receptor, proto-oncogene c-ErbB-1 , receptor tyrosine-protein kinase erbB-1.  It is a member of the epidermal growth factor receptor (EGFR) family of receptor tyrosine kinases (RTK).  Four members of the EGFR family have been identified: EGFR (ERBB1, HER1), HER2 (ERBB2), HER3 (ERBB3) and HER4 (ERBB4).  They typically contain an extracellular ligand binding domain (ECD), a transmembrane domain (TM), and an intracellular kinase domain that can interact with a multitude of signaling molecules and exhibit both ligand-dependent and ligand-independent activity.  The human EGFR has 1210 amino acid (aa), including a 24 aa signal peptide, a 621 aa ECD, and a 23 aa TM segment. It shares 88% aa sequence identity with murine EGFR and 43-44% identity with the ECD of other 3 human EGFR members.  EGFR binds to a subset of the EGF family ligands, including EGF, amphiregulin, TGFα, β-cellulin, epiregulin, HB-EGF, and epigen.  EGFR signaling is initiated by ligand binding, leading to EGFR receptor homo-/hetero-dimerization and autophosphorylation by the intracellular kinase domain and receptor activation.  Subsequently, phosphorylation of cytoplasmic substrates occurs and a signaling cascade is initiated that drives many cellular responses, including changes in gene expression, cytoskeletal rearrangement, anti-apoptosis and increased cell proliferation.  EGFR is known to form heterodimers with the ligand-activated HER3 or HER4.  EGFR is overexpressed in a variety of tumors.  Mutations in EGFR gene are associated with lung cancer.  EGFR is the target of several anti-cancer therapeutics.

 

 

Gene Symbol: EGFR; ERBB; ERBB1; HER1; mENA; PIG61; SA7

 

NCBI Gene ID: 1956 (human)

 

Uniprot Entry: P00533 (human)

 

Construct Details: The recombinant human EGFR extracellular or ecto-domain (ECD) is expressed as a 625 amino acid protein consisting of Leu25-Thr638 region of EGFR and a C-terminal poly Histidine-tag, which exists as a monomer under reducing and non-reducing conditions

 

Source: Human cells stably expressing EGFR ECD and growing in chemical-defined media with no animal components or antibiotics

 

Deduced Amino Acid Sequence:

LEEKKVCQGTSNKLTQLGTFEDHFLSLQRMFNNCEVVLGNLEITYVQRNYDLSFLKTIQEVAGYVLIALNTVERIPLENL

QIIRGNMYYENSYALAVLSNYDANKTGLKELPMRNLQEILHGAVRFSNNPALCNVESIQWRDIVSSDFLSNMSMDFQNHL

GSCQKCDPSCPNGSCWGAGEENCQKLTKIICAQQCSGRCRGKSPSDCCHNQCAAGCTGPRESDCLVCRKFRDEATCKDTC

PPLMLYNPTTYQMDVNPEGKYSFGATCVKKCPRNYVVTDHGSCVRACGADSYEMEEDGVRKCKKCEGPCRKVCNGIGIGE

FKDSLSINATNIKHFKNCTSISGDLHILPVAFRGDSFTHTPPLDPQELDILKTVKEITGFLLIQAWPENRTDLHAFENLE

IIRGRTKQHGQFSLAVVSLNITSLGLRSLKEISDGDVIISGNKNLCYANTINWKKLFGTSGQKTKIISNRGENSCKATGQ

VCHALCSPEGCWGPEPRDCVSCRNVSRGRECVDKCNLLEGEPREFVENSECIQCHPECLPQAMNITCTGRGPDNCIQCAH

YIDGPHCVKTCPAGVMGENNTLVWKYADAGHVCHLCHPNCTYGCTGPGLEGCPTSTGHHHHHHHH

 

M.W.: Calculated molecular mass (kDa): 69.2; Estimated by SDS-PAGE under reducing condition (kDa): ~80, suggestion that the protein is glycosylated

 

Calculated PI: 6.59

 

Calculated extinction coefficients (M-1 cm-1, at 280nm):  59965.

 

Purity: >95% judged by SDS-PAGE under reducing condition (see the gel image above)

 

Formulation: Supplied at 0.5 mg/ml in sterile PBSpH7.4 (carrier free). The purified recombinant protein was labeled with Biotin (3-5 Biotin per molecule) using the standard procedure.

 

Endotoxin Level: <0.1 EU per 1 μg of purified recombinant protein determined by the LAL method

 

Biological Activity: The purified recombinant protein binds anti-EGFR mAbs (SKU#MAB0118, #MAB0520, #MAB0169) with high affinity (K_D < 1 nM - see the Technical Data) and therapeutic mAbs such as Erbitux®, Vectbix® with high affinity (estimated K_D < 20 nM) measured by ELISA with 20 ng of this protein as ligand coated on the plate.

 

 

 

FOR RESEARCH USE ONLY. NOT FOR DIAGNOSTIC OR THERAPEUTIC USE IN HUMAN.

Restriction: This product is not transferable or re-sellable.  Customer obtain no right to transfer, assign, or sublicense its use rights, or to transfer, resell, package, or otherwise distribute the product, or to use the product for the benefit of any third party or for any commercial purpose.  Customer may only use the product in compliance with applicable local, state and federal laws, regulations and rules.  Customer may not directly or indirectly use the product or allow the transfer, transmission, export or re-export of all or any part of the product in violation of any export control law or regulation of the united states or any other relevant jurisdiction.  Your use of this product constitutes acceptance of the terms of this limited use agreement.  Please refer to our “terms & conditions” for details.  If you are not willing to accept the limitation of this agreement, G&P Biosciences will accept return of the product for a full/partial refund.

 

 

 

Recombinant EGFR ECD protein (SKU#FCL0169) was immobilized at 20 ng/well on a 96-well ELISA plate and incubated with serially diluted anti-EGFR human IgG1 monoclonal antibody (SKU#MAB0118). Anti-HER2 (SKU#MAB0203) human IgG1 monoclonal antibody was used as a control to demonstrate the specificity of EGFR binding under the experimental conditions.  The bound antibodies were detected by goat anti-human IgG HRP-conjugated secondary antibody.  The ELISA binding curve was fit and K_D was calculated for anti-EGFR : EGFR binding (affinity/avidity) to be 13 ± 0.7 pM. 

 

 

 

FOR RESEARCH USE ONLY. NOT FOR DIAGNOSTIC OR THERAPEUTIC USE IN HUMAN.

 

 

The product is shipped at 4°C. Upon receipt, centrifuge the product briefly before opening the vial. It is recommended to store small aliquots at the temperature below –20°C for long-term storage and the product is stable for 3 months. The undiluted protein can be stored at 4°C for no more than 2 weeks.

 

Use a manual defrost freezer and avoid repeated freeze-thaw cycles.

 

 

 

FOR RESEARCH USE ONLY. NOT FOR DIAGNOSTIC OR THERAPEUTIC USE IN HUMAN.

 

 

1.  Science 306: 1506-1507, 2004.
2.  Nature 309: 418-425, 1984
3.  New Eng. J. Med. 350: 2129-2139, 2004.
4.  Cell 125: 1137-1149, 2006.

 

 

 

Product datasheet (pdf) can be downloaded here: FCL0169B-PDS.pdf

 

 

 

Additional supporting documents, including COA and MSDS are available upon request.

 

 

 

FOR RESEARCH USE ONLY. NOT FOR DIAGNOSTIC OR THERAPEUTIC USE IN HUMAN.

Restriction: This product is not transferable or re-sellable.  Customer obtain no right to transfer, assign, or sublicense its use rights, or to transfer, resell, package, or otherwise distribute the product, or to use the product for the benefit of any third party or for any commercial purpose.  Customer may only use the product in compliance with applicable local, state and federal laws, regulations and rules.  Customer may not directly or indirectly use the product or allow the transfer, transmission, export or re-export of all or any part of the product in violation of any export control law or regulation of the united states or any other relevant jurisdiction.  Your use of this product constitutes acceptance of the terms of this limited use agreement.  Please refer to our “terms & conditions” for details.  If you are not willing to accept the limitation of this agreement, G&P Biosciences will accept return of the product for a full/partial refund.