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Flow cytometric analysis of Nkx6.1 expression in mouse pancreatic tumor (insulinoma) cells. Beta-T-C6 (ATCC, CRL-11506™) cells were fixed with 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 (Cat. No. 565572; dashed line histogram) or Alexa Fluor™ 488 Mouse Anti-Nkx6.1 antibody (Cat. No. 567608/567609; solid line histogram). Fluorescence histogram showing Nkx6.1 expression (or Ig Isotype staining) was derived from gated events with the side and forward light-scattering characteristics of intact cells. Flow cytometry and data analysis were performed using a BD LSRFortessa™ Cell Analyzer System and FlowJo™ software.
BD Pharmingen™ Alexa Fluor™ 488 Mouse Anti-Nkx6.1
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Preparation And Storage
Recommended Assay Procedures
BD® CompBeads can be used as surrogates to assess fluorescence spillover (Compensation). When fluorochrome conjugated antibodies are bound to BD® CompBeads, they have spectral properties very similar to cells. However, for some fluorochromes there can be small differences in spectral emissions compared to cells, resulting in spillover values that differ when compared to biological controls. It is strongly recommended that when using a reagent for the first time, users compare the spillover on cells and BD® CompBeads to ensure that BD® CompBeads are appropriate for your specific cellular application.
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.
- 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.
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- Alexa Fluor® 488 fluorochrome emission is collected at the same instrument settings as for fluorescein isothiocyanate (FITC).
- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
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Companion Products
The R11-560 monoclonal antibody specifically binds to human and mouse homeobox protein Nkx6.1. The transcription factor Nkx6.1 plays a role in pancreatic islet beta cell development through the regulation of HNF1a and insulin genes. Nkx6.1 is considered to be a highly specific transcription factor marking pancreatic beta islet cells, where it is expressed in both developing and mature cells. In addition, Nkx6.1 functions as a transcriptional repressor induced by the Sonic Hedgehog (Shh) signaling pathway in fate determination of several neuronal cell types. During antibody development, the purified R11-560 monoclonal antibody was found to detect Nkx6.1 by western blot and indirect immunofluorescent staining followed by flow cytometric analysis or imaging analysis.
Development References (6)
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D'Amour KA, Bang AG, Eliazer S, et al . Production of pancreatic hormone-expressing endocrine cells from human embryonic stem cells. Nat Biotechnol. 2006; 24(12):1481-1483. (Biology). View Reference
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Donelan W, Koya V, Li SW, Yang LJ. Distinct regulation of hepatic nuclear factor 1alpha by NKX6.1 in pancreatic beta cells. J Biol Chem. 2010; 285(16):12181-12189. (Biology). View Reference
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Inoue H, Rudnick A, German MS, Veile R, Donis-Keller H, Permutt MA. Isolation, characterization, and chromosomal mapping of the human Nkx6.1 gene (NKX6A), a new pancreatic islet homeobox gene. Genomics. 1997; 40:367-370. (Biology). View Reference
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Nelson SB, Schaffer AE, Sander M. The transcription factors Nkx6.1 and Nkx6.2 possess equivalent activities in promoting beta-cell fate specification in Pdx1+ pancreatic progenitor cells. Development. 2007; 134(13):2491-2500. (Biology). View Reference
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Pedersen IL, Klinck R, Hecksher-Sorensen J, et al. Generation and characterization of monoclonal antibodies against the transcription factor Nkx6.1. J Histochem Cytochem. 2006; 54(5):567-574. (Biology). View Reference
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Prakash N, Puelles E, Freude K, et. al. Nkx6-1 controls the identity and fate of red nucleus and oculomotor neurons in the mouse midbrain. Development. 2009; 136(15):2545-2555. (Biology). View Reference
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For Research Use Only. Not for use in diagnostic or therapeutic procedures.