United Kingdom (UK)
Recombinant monoclonal antibody to ABCG2. Manufactured using AbAb’s Recombinant Platform with variable regions (i.e. specificity) from the hybridoma 5D3.
UniProt Accession Number of Target Protein: Q9UNQ0
Alternative Name(s) of Target: CD338; CDw338; ABCP; BCRP; BCRP1; MXR; Breast cancer resistance protein; mitoxantrone resistance; Urate exporter; Broad substrate specificity ATP-binding cassette transporter ABCG2; ATP-binding cassette sub-family G member 2; Mitoxantrone resistance-associated protein; Placenta-specific ATP-binding cassette transporter
Immunogen: The original antibody was generated by immunizing mice with intact mouse 3T3 fibroblasts expressing the human ABCG2 protein.
Specificity: This antibody recognizes and binds a conformation-sensitive external epitope of ABCG2 on living cells. ABCG2 is a broad substrate specificity ATP-dependent transporter from the ATP-binding cassette (ABC) family that actively extrudes a wide variety of physiological compounds, dietary toxins and xenobiotics from cells. (PMID:11306452, 12958161, 19506252, 20705604, 28554189, 30405239, 31003562). The human ABCG2 protein is an important primary active transporter for hydrophobic compounds in several cell types, and its overexpression causes multidrug resistance in tumors (PMID: 15557326). ABCG2 is associated with drug resistance in acute myeloid leukemia (AML) (PMID: 12393637). ABCG2 expression may also have potential as a marker that identifies human limbal epithelial stem cells (PMID: 15625123).
Application Notes: This antibody is recommended for identification of human ACBG2 in a sample. This antibody was used in the immunostaining of bone marrow cells to establish a direct link between the SP phenotype and ABCG2 expression (PMID: 11533706). The IgG2b version of this antibody was used to identify AML cell lines that express ABCG2 protein using flow cytometry. This antibody inhibits ABCG2 function, presumably by binding to an external epitope and preventing conformational changes in the transporter, and therefore provides a specific inhibitor assay for ABCG2 function (PMID: 12393637). The SP (side population phenotype) analysis of limbal epithelial cells using 5D3 and TPS-A showed that both 5D3 and TPS-A effectively inhibit Hoechst 33342 dye efflux from these cells (PMID: 15135043). Conformational changes in the ABCG2 multidrug transporter result in the modification of its interaction with 5D3 antibody. This antibody therefore cannot be used to identify human ABCG2 in an immunoblot/western blot assay. The binding of this antibody was maximum in the presence of ATP, ADP, or Ko143, whereas the non-hydrolysable ATP analog, adenosine 5'-(beta, gamma-imido)triphosphate (AMP-PNP), and nucleotide trapping by vanadate decreased antibody binding. In membranes expressing the ABCG2-K86M mutant, ATP, ADP, and AMP-PNP decreased, whereas Ko143 increased the binding of this antibody. 5D3 can be used as a sensitive tool to reveal intramolecular changes, reflecting ATP binding, the formation of a catalytic intermediate, or substrate inhibition within the transport cycle of the ABCG2 protein (PMID: 15557326). This antibody was also used in a study involving localization of ABCG2 mitoxantrone resistance-associated protein in normal tissues using immunohistochemistry (PMID: 15990223). This antibody was also used in determining ABCG2 expression using immunofluorescent staining (PMID: 15625123).
Antibody first published in:
Zhou et al. The ABC transporter Bcrp1/ABCG2 is expressed in a wide variety of stem cells and is a molecular determinant of the side-population phenotype. Nat Med. 2001 Sep;7(9):1028-34. PMID:11533706
Note on publication:
Describes the use of this antibody in the immunostaining of bone marrow cells expressing ABCG2.
Immunofluorescence staining of A549 cells with anti-ABCG2 antibody 5D3 (Ab02927). Immunofluorescence analysis of paraformaldehyde-fixed A549 cells on Shi-fix™ coverslips stained with the mouse IgG2b version of 5D3 (Ab02927-3.0) (1:100 dilution) for 1h followed by Alexa Fluor® 488 secondary antibody (1:1000 dilution), showing cytoplasmic staining. The nuclear stain is DAPI (blue). Panels show, from left-right, top-bottom, Ab02927-3.0, DAPI, merged channels, and an isotype control. The isotype control was an unknown specificity antibody (Ab00178-3.0) followed by staining with Alexa Fluor® 488 secondary antibody.