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 CompBead to ensure that BD CompBeads are appropriate for your specific cellular application.
For optimal and reproducible results, BD Horizon Brilliant Stain Buffer should be used anytime two or more BD Horizon Brilliant dyes 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).
Product Notices
- 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.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
- 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.
- Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
- Alexa Fluor® is a registered trademark of Life Technologies Corporation.
- BD Horizon Brilliant Violet 711 is covered by one or more of the following US patents: 8,110,673; 8,158,444; 8,227,187; 8,455,613; 8,575,303; 8,354,239.
- Cy is a trademark of GE Healthcare.
Companion Products
The RAM34 monoclonal antibody specifically binds to the CD34 glycoprotein on the surface of three independently derived mouse CD34-transfected cell lines. RAM34 antibody also reacts with the mouse cell lines PA6, 416B, Swiss 3T6, NIH, 3T3, DA1, and M1, all of which are positive for expression of mouse CD34 mRNA. Cell lines shown to be negative for CD34 mRNA transcript, including WEHI-3B, EL4, 18.8, and CMT64/61, are also negative for surface expression of CD34 as determined by RAM34 staining. Normal thymocytes and splenocytes are negative for CD34 expression. In the bone marrow, 7-10% of cells are stained with the RAM34 antibody, including most of the Ly-6A/E (Sca-1)+ CD90 (Thy-1)low Lineage Marker- hematopoietic stem cell-enriched subpopulation and myeloerythroid progenitors. CD34 is also expressed on a small percentage of fetal liver cells, including NK-cell progenitors. CD34 has been reported to be expressed on the endothelium of capillaries and, in this form, to function as a ligand for L-selectin. Consistent with this observation, RAM34 antibody stains endothelial cells in spleen, thymus, and postcapillary HEVs in the lymph nodes. It is reported that RAM34 antibody can be used to select CD34+ CD117 (c-Kit)+ Ly-6A/E (Sca-1)+ Lineage Marker- bone marrow-derived hematopoietic stem cells, capable of short-term multi-lineage reconstitution of lethally irradiated mice. In contrast, the CD34- CD117+ Sca-1+ Lineage Marker- population contains self-renewing hematopoietic stem cells. Similarly, the bone marrow population with high dye-efflux capacity and highly enriched for long-term reconstituting hematopoietic stem cells, is CD34- CD117 (c-Kit)+ Ly-6A/E (Sca-1)+ Lineage Marker-.
The antibody was conjugated to BD Horizon™ BV711 which is part of the BD Horizon Brilliant™ Violet family of dyes. This dye is a tandem fluorochrome of BD Horizon BV421 with an Ex Max of 405-nm and an acceptor dye with an Em Max at 711-nm. BD Horizon BV711 can be excited by the violet laser and detected in a filter used to detect Cy™5.5 / Alexa Fluor® 700-like dyes (eg, 712/20-nm filter). Due to the excitation and emission characteristics of the acceptor dye, there may be moderate spillover into the Alexa Fluor® 700 and PerCP-Cy5.5 detectors. However, the spillover can be corrected through compensation as with any other dye combination.
Development References (11)
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Goodell MA, Rosenzweig M, Kim H, et al. Dye efflux studies suggest that hematopoietic stem cells expressing low or undetectable levels of CD34 antigen exist in multiple species. Nat Med. 1997; 3(12):1337-1345. (Clone-specific: Flow cytometry). View Reference
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Lu J, Patrene KD, Herberman RB, Boggs SS. Expression of murine CD34 by fetal liver NK cell progenitors. Exp Hematol. 1999; 27(2):272-281. (Clone-specific: Flow cytometry, Fluorescence activated cell sorting). View Reference
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Morel F, Szilvassy SJ, Travis M, Chen B, Galy A. Primitive hematopoietic cells in murine bone marrow express the CD34 antigen. Blood. 1996; 88(10):3774-3784. (Clone-specific: Flow cytometry, Fluorescence activated cell sorting). View Reference
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Osawa M, Hanada K, Hamada H, Nakauchi H. Long-term lymphohematopoietic reconstitution by a single CD34-low/negative hematopoietic stem cell. Science. 1996; 273(5272):242-245. (Immunogen: Flow cytometry). View Reference
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Suda J, Sudo T, Ito M, Ohno N, Yamaguchi Y, Suda T. Two types of murine CD34 mRNA generated by alternative splicing. Blood. 1992; 79(9):2288-2295. (Biology). View Reference
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Suzuki A, Andrew DP, Gonzalo JA, et al. CD34-deficient mice have reduced eosinophil accumulation after allergen exposure and show a novel crossreactive 90-kD protein. Blood. 1996; 87(9):3550-3562. (Clone-specific: Immunoaffinity chromatography, Immunohistochemistry, Immunoprecipitation). View Reference
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For Research Use Only. Not for use in diagnostic or therapeutic procedures.