Preparation And Storage
Recommended Assay Procedures
For optimal and reproducible results, BD Horizon Brilliant Stain Buffer should be used anytime two or more BD Horizon Brilliant dyes (including BD Optibuild Brilliant reagents) are used in the same experiment. Fluorescent dye interactions may cause staining artifacts which may affect data interpretation. The BD Horizon Brilliant Stain Buffer was designed to minimize these interactions. More information can be found in the Technical Data Sheet of the BD Horizon Brilliant Stain Buffer (Cat. No. 563794/566349) or the BD Horizon Brilliant Stain Buffer Plus (Cat. No. 566385).
For Immunofluorescence Applications:
The use of a mounting reagent (eg, ProLong® Gold) is highly recommended to maximize the photostability of BV480. For confocal microscopy systems, a 440 nm laser is the optimal excitation source and the recommended emission filter is a 485/20 nm bandpass filter.
For epifluorescence microscopes with broad spectrum excitation sources, the recommended excitation and emission filters are 445/20 nm and 485/20 nm bandpass filters, respectively. For specific multicolor imaging applications, the exact filter configurations should be optimized by the end user. For additional instrument/filter configuration information, please visit http://www.bdbiosciences.com/research/cellularimaging.
Product Notices
- This antibody was developed for use in flow cytometry.
- The production process underwent stringent testing and validation to assure that it generates a high-quality conjugate with consistent performance and specific binding activity. However, verification testing has not been performed on all conjugate lots.
- Researchers should determine the optimal concentration of this reagent for their individual applications.
- An isotype control should be used at the same concentration as the antibody of interest.
- 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.
- BD Horizon Brilliant Stain Buffer is covered by one or more of the following US patents: 8,110,673; 8,158,444; 8,575,303; 8,354,239.
- BD Horizon Brilliant Violet 480 is covered by one or more of the following US patents: 8,575,303; 8,354,239.
- ProLong® is a registered trademark of Thermo Fisher Scientific, Inc. Waltham, MA.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
Companion Products
The 5D3/CD338 monoclonal antibody specifically binds to an epitope of ABCG2 (BCRP1), a multi-drug resistance protein that is a member of ATP binding cassette (ABC) transporters. It is highly expressed on primitive stem cells as identified by the "side-population" (SP) phenotype. This SP phenotype is based on the efflux of fluorescent dyes such as Rhodamine 123 and Hoechst 33342. The expression of ABCG2 appears to be highly conserved as it has been identified in various species. Studies show that highly purified murine stem cells express BCRP1 mRNA and this expression declines sharply as the stem cells express CD34. The highest levels of BCRP1 mRNA expression have been seen in KDR+ human stem cells. ABCG2/BCRP1 was clustered as CD338 in the HLDA8 workshop.
The antibody was conjugated to BD Horizon™ BV480 which is part of the BD Horizon Brilliant™ Violet family of dyes. With an Ex Max of 436-nm and Em Max at 478-nm, BD Horizon BV480 can be excited by the violet laser and detected in the BD Horizon BV510 (525/40-nm) filter set. BV480 has less spillover into the BV605 detector and, in general, is brighter than BV510.
Development References (3)
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Bunting KD. ABC transporters as phenotypic markers and functional regulators of stem cells. Stem Cells. 2002; 20(1):11-20. (Biology). View Reference
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Ozvegy-Laczka C, Laczkó R, Hegedus, et al. Interaction with the 5D3 monoclonal antibody is regulated by intramolecular rearrangements but not by covalent dimer formation of the human ABCG2 multidrug transporter. J Biol Chem. 2008; 283(38):26059-26070. (Immunogen: Flow cytometry). View Reference
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Zhou S, Schuetz JD, Bunting KD, 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; 7(9):1028-1034. (Biology). View Reference
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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.
