Flow cytometric analysis of FoxA2 in definitive endoderm derived from human embryonic stem (ES) cells. H9 human ES cells (WiCell, Madison, WI) grown on an irradiated mouse embryonic feeder layer were differentiated to definitive endoderm for 3 days (D'Amour et al, 2005) in RPMI medium supplemented with 0.5% FBS, 1x L-glutamine, and 100 ng/ml Activin A (R&D Systems). Control ES cells (left panel) and day-3 differentiated cells (right panel) were fixed with BD Cytofix buffer (Cat. No. 554655) and permeabilized with BD™ Phosflow Perm buffer III (Cat. No. 558050). The cells were stained with either PE Mouse IgG1, κ isotype control (dashed lines, Cat. No.554680) or PE Mouse Anti-human FoxA2 antibody (solid lines) at matched concentrations. The histograms were derived from gated events based on light scattering characteristics of the human ES and H9-derived endoderm cells, respectively. Flow cytometry was performed on a BD LSR™ II flow cytometry system. This antibody can also be used on fixed cells permeabilized with 0.1% Triton X-100.
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Flow cytometric analysis of FoxA2 in definitive endoderm derived from human embryonic stem (ES) cells. H9 human ES cells (WiCell, Madison, WI) grown on an irradiated mouse embryonic feeder layer were differentiated to definitive endoderm for 3 days (D'Amour et al, 2005) in RPMI medium supplemented with 0.5% FBS, 1x L-glutamine, and 100 ng/ml Activin A (R&D Systems). Control ES cells (left panel) and day-3 differentiated cells (right panel) were fixed with BD Cytofix buffer (Cat. No. 554655) and permeabilized with BD™ Phosflow Perm buffer III (Cat. No. 558050). The cells were stained with either PE Mouse IgG1, κ isotype control (dashed lines, Cat. No.554680) or PE Mouse Anti-human FoxA2 antibody (solid lines) at matched concentrations. The histograms were derived from gated events based on light scattering characteristics of the human ES and H9-derived endoderm cells, respectively. Flow cytometry was performed on a BD LSR™ II flow cytometry system. This antibody can also be used on fixed cells permeabilized with 0.1% Triton X-100.
Product Details
BD Pharmingen™
Forkhead box A2, HNF3B, HNF-3B, HNF-3β, hepatocyte nuclear factor 3β
Human (QC Testing)
Mouse BALB/c IgG1, κ
Human FoxA2 Peptide
Intracellular staining (flow cytometry) (Routinely Tested)
5 µl
AB_10716057
Aqueous buffered solution containing BSA and ≤0.09% sodium azide.
RUO
Preparation And Storage
The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. The antibody was conjugated with R-PE under optimum conditions, and unconjugated antibody and free PE were removed. Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze.
Product Notices
- This reagent has been pre-diluted for use at the recommended Volume per Test. We typically use 1 × 10^6 cells in a 100-µl experimental sample (a test).
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
- An isotype control should be used at the same concentration as the antibody of interest.
- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
- Source of all serum proteins is from USDA inspected abattoirs located in the United States.
- Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
Companion Products
Stain Buffer (FBS) RUO
Size
500 mL
Cat No.
554656
Fixation Buffer RUO
Size
100 mL
Cat No.
554655
Perm Buffer III RUO
Size
125 mL
Cat No.
558050
PE Mouse IgG1, κ Isotype Control RUO
Size
0.1 mg
Cat No.
554680
561589 Rev. 1
Antibody Details
N17-280
FoxA2, forkhead box A2, is a member of the forkhead class of DNA-binding proteins that regulates gene expression in the liver, pancreatic islets, adipocytes and some neural cells. This hepatocyte nuclear factor is a transcriptional activator for liver-specific genes such as alpha fetoprotein, albumin, tyrosine aminotransferase and transthyretin. FoxA2 is expressed in embryonic endoderm, the germ layer that gives rise to the digestive system, and contributes to the specification of the pancreas and the regulation of glucose homoeostasis. FoxA2 also has roles in neural development. Specifically, FoxA2 cooperates with related FoxA1 in the specification and differentiation of midbrain dopaminergic neurons in a dosage-dependent manner.
561589 Rev. 1
Format Details
PE
R-Phycoerythrin (PE), is part of the BD family of Phycobiliprotein dyes. This fluorochrome is a multimeric fluorescent phycobiliprotein with excitation maximum (Ex Max) of 496 nm and 566 nm and an emission maximum (Em Max) at 576 nm. PE is designed to be excited by the Blue (488 nm), Green (532 nm) and Yellow-Green (561 nm) lasers and detected using an optical filter centered near 575 nm (e.g., a 575/26-nm bandpass filter). As PE is excited by multiple lasers, this can result in cross-laser excitation and fluorescence spillover on instruments with various combinations of Blue, Green, and Yellow-Green lasers. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
PE
Yellow-Green 488 nm, 532 nm, 561 nm
496 nm, 566 nm
576 nm
561589 Rev.1
Citations & References
Development References (7)
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Burtscher I, Lickert H.. Foxa2 regulates polarity and epithelialization in the endoderm germ layer of the mouse embryo. Development. 2009; 136(6):1029-1038. (Biology). View Reference
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Clevidence DE, Overdier DG, Tao W, et al. Identification of nine tissue-specific transcription factors of the hepatocyte nuclear factor 3/forkhead DNA-binding-domain family. Proc Natl Acad Sci U S A. 1993; 90(9):3948-3952. (Biology). View Reference
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D'Amour KA, Agulnick AD, Eliazer S, Kelly OG, Kroon E, Baetge EE. Efficient differentiation of human embryonic stem cells to definitive endoderm.. Nat Biotechnol. 2005; 23(12):1534-41. (Methodology). View Reference
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Lai E, Prezioso VR, Tao WF, Chen WS, Darnell JE Jr. Hepatocyte nuclear factor 3 alpha belongs to a gene family in mammals that is homologous to the Drosophila homeotic gene fork head. Genes Dev. 1991; 5(3):416-427. (Biology). View Reference
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Lin W, Metzakopian E, Mavromatakis YE, et al. Foxa1 and Foxa2 function both upstream of and cooperatively with Lmx1a and Lmx1b in a feedforward loop promoting mesodiencephalic dopaminergic neuron development.. Dev Biol. 2009; 333(2):386-396. (Biology). View Reference
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Monaghan AP, Kaestner KH, Grau E, Schütz G. Postimplantation expression patterns indicate a role for the mouse forkhead/HNF-3 alpha, beta and gamma genes in determination of the definitive endoderm, chordamesoderm and neuroectoderm.. Development. 1993; 119(3):567-578. (Biology). View Reference
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Thomson JA, Itskovitz-Eldor J, Shapiro SS, et al. Embryonic stem cell lines derived from human blastocysts. Science. 1998; 282:1145-1147. (Methodology). View Reference
561589 Rev. 1
Please refer to Support Documents for Quality Certificates
Global - Refer to manufacturer's instructions for use and related User Manuals and Technical data sheets before using this products as described
Comparisons, where applicable, are made against older BD Technology, manual methods or are general performance claims. Comparisons are not made against non-BD technologies, unless otherwise noted.
For Research Use Only. Not for use in diagnostic or therapeutic procedures.
