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Flow cytometric analysis of fibronectin in human mesenchymal stem cells (MSC). MSC (Lonza), passage 6, were dissociated and fixed in BD Cytofix™ Fixation Buffer (Cat. No. 554655), and permeabilized with BD Phosflow™ Perm Buffer III (Cat. No. 558050). The cells were stained with either Alexa Fluor® 488 Mouse IgG1, κ isotype control (dashed line, Cat. No. 557782) or Alexa Fluor® 488 Anti-Fibronectin monoclonal antibody (solid line) at matched concentrations. Histograms were derived from gated events based on light scattering characteristics of MSC. Flow cytometry was performed on a BD LSRFortessa™ II flow cytometry system. BD Phosflow™ Perm/Wash Buffer I (Cat. No. 557885) is also suitable for permeabilization.
Immunofluorescent analysis of fibronectin in human mesenchymal stem cells (MSC). MSC (Lonza), passage 6, were fixed in BD Cytofix™ Fixation Buffer (Cat. No. 554655), permeabilized with BD Phosflow™ Perm Buffer III (Cat. No. 558050), and stained with Alexa Fluor® 488 Mouse Anti-Fibronectin monoclonal antibody (pseudo-colored green) at 1.2 µg/ml. Cell nuclei were stained with DAPI (pseudo-colored blue). The images were captured on a BD Pathway™ 435 Cell Analyzer and merged using BD Attovision™Software. 0.1% Triton™ X-100 is also suitable for permeabilization.
BD Pharmingen™ Alexa Fluor® 488 Mouse Anti-Fibronectin
BD Pharmingen™ Alexa Fluor® 488 Mouse Anti-Fibronectin
Regulatory Status Legend
Any use of products other than the permitted use without the express written authorization of Becton, Dickinson and Company is strictly prohibited.
Preparation And Storage
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).
- An isotype control should be used at the same concentration as the antibody of interest.
- Species cross-reactivity detected in product development may not have been confirmed on every format and/or application.
- Alexa Fluor® 488 fluorochrome emission is collected at the same instrument settings as for fluorescein isothiocyanate (FITC).
- Alexa Fluor® is a registered trademark of Molecular Probes, Inc., Eugene, OR.
- The Alexa Fluor®, Pacific Blue™, and Cascade Blue® dye antibody conjugates in this product are sold under license from Molecular Probes, Inc. for research use only, excluding use in combination with microarrays, or as analyte specific reagents. The Alexa Fluor® dyes (except for Alexa Fluor® 430), Pacific Blue™ dye, and Cascade Blue® dye are covered by pending and issued patents.
- Triton is a trademark of the Dow Chemical Company.
- 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.
- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
Companion Products
The 240-kDa dimeric fibronectin protein exists in two forms: a soluble protomer in body fluids and an insoluble multimer in the extracellular matrix. The latter is the primary functional form and creates a substrate for cell migration, a role which makes fibronectin vital to embryogenesis and wound response. Fibronectin mediates cytoskeletal organization, cell attachment, and cellular signaling through interactions with cellular receptors. Although various isoforms of fibronectin are derived by alternative splicing, they share a common N-terminus which is a critical region for cell surface binding in an initial step of multimer assembly. Further polymerization steps are regulated by fibronectin/integrin interactions and result in generation of the complex fibrils that constitute the fibronectin matrix.
Development References (5)
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Chen H, Mosher DF. Formation of sodium dodecyl sulfate-stable fibronectin multimers. Failure to detect products of thiol-disulfide exchange in cyanogen bromide or limited acid digests of stabilized matrix fibronectin. J Biol Chem. 1996; 271(15):9084-9089. (Biology). View Reference
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Danen EH, Sonneveld P, Brakebusch C, Fassler R, Sonnenberg A. The fibronectin-binding integrins alpha5beta1 and alphavbeta3 differentially modulate RhoA-GTP loading, organization of cell matrix adhesions, and fibronectin fibrillogenesis. J Cell Biol. 2002; 159(6):1071-1086. (Clone-specific: Immunofluorescence). View Reference
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Rhee CS, Sen M, Lu D, et al. Wnt and frizzled receptors as potential targets for immunotherapy in head and neck squamous cell carcinomas. Oncogene. 2002; 21(42):6598-6605. (Clone-specific: Western blot). View Reference
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Sechler JL, Takada Y, Schwarzbauer JE. Altered rate of fibronectin matrix assembly by deletion of the first type III repeats. J Cell Biol. 1996; 134(2):573-583. (Biology). View Reference
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Zuk A, Bonventre JV, Brown D, Matlin KS. Polarity, integrin, and extracellular matrix dynamics in the postischemic rat kidney. Am J Physiol. 1998; 275(3):C711-C731. (Clone-specific: Immunohistochemistry). View Reference
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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.