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
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
- Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
- For U.S. patents that may apply, see bd.com/patents.
- 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.
- Since applications vary, each investigator should titrate the reagent to obtain optimal results.
- 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.
- When using high concentrations of antibody, background binding of this dye to erythroid fragments produced by ammonium chloride-based lysis, such as with BD Pharm Lyse™ Lysing Buffer (Cat. No. 555899), has been observed when the antibody conjugate was present during the lysis procedure. This may cause nonspecific staining of target cells, such as leukocytes, which have bound the resulting erythroid fragments. This background can be mitigated by any of the following: titrating the antibody conjugate to a lower concentration, fixing samples with formaldehyde, or removing erythrocytes before staining (eg, gradient centrifugation or pre-lysis with wash). This background has not been observed when cells were lysed with BD FACS™ Lysing Solution (Cat. No. 349202) after staining.
- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
- An isotype control should be used at the same concentration as the antibody of interest.
- Please observe the following precautions: We recommend that special precautions be taken (such as wrapping vials, tubes, or racks in aluminum foil) to protect exposure of conjugated reagents, including cells stained with those reagents, to any room illumination. Absorption of visible light can significantly affect the emission spectra and quantum yield of tandem fluorochrome conjugates.
- Tandem fluorochromes contain both an energy donor and an energy acceptor. Although every effort is made to minimize the lot-to-lot variation in the efficiency of the fluorochrome energy transfer, differences in the residual emission from the donor may be observed. Additionally, multi-laser cytometers may directly excite both the donor and acceptor fluorochromes. Therefore, we recommend for every tandem conjugate, a matched individual single-stain control be acquired for generating a compensation or spectral unmixing matrix.
- Cy is a trademark of Global Life Sciences Solutions Germany GmbH or an affiliate doing business as Cytiva.
Companion Products
The 1AH2 monoclonal antibody specifically binds to CD137. CD137 is a member of the TNFR/NGFR superfamily that is likewise known as Tnfrsf9, 4-1BB, Ly-63, or ILA. Monomers or multimeric forms of CD137 are expressed, upon activation, on the surface of splenic T lymphocytes, thymocytes, intestinal intraepithelial T lymphocytes (IEL), and some T cell lines and clones. While stimulating T cells by IL-2, IL-4, or anti-CD28 alone does not result in the expression of CD137; addition of IL-2, IL-4, anti-CD28, or syngeneic accessory cells to splenic T cells stimulated via TCR/CD3 can result in a high level CD137 expression. CD137 is also reportedly expressed on IL-2 activated NK cells, but not on freshly isolated NK cells. CD137 physically associates with p56 [lck] through a Cys-Arg-Cys-Pro binding site in its cytoplasmic domain; the same motif in the cytoplasmic tail of the CD4 and CD8a molecules is responsible for association with p56 [lck]. A signaling function for the CD137 molecule in mouse T cells is indicated by reports in which cross-linking of CD137 with 1AH2 mAb resulted in enhanced proliferation of CD3e-activated splenic T cells and IEL and in enhanced cytolytic activity of IEL in response to immobilized anti-CD3e. In addition to extracellular matrix proteins which bind to CD137, the CD137L (4-1BBL) serves as a ligand for CD137. This molecule has also been detected on LPS-activated macrophages, and anti-IgM antibody-activated splenic B cells. Interaction between T and B cells through CD137/CD137L is reported to play a role in antigen presentation, further supporting a costimulatory role for CD137 in the immune response of T lymphocytes.
Development References (11)
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Chalupny NJ, Peach R, Hollenbaugh D, Ledbetter JA, Farr AG, Aruffo A. T-cell activation molecule 4-1BB binds to extracellular matrix proteins. Proc Natl Acad Sci U S A. 1992; 89(21):10360-10364. (Biology). View Reference
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DeBenedette MA, Chu NR, Pollok KE, et al. Role of 4-1BB ligand in costimulation of T lymphocyte growth and its upregulation on M12 B lymphomas by cAMP. J Exp Med. 1995; 181(3):985-992. (Biology). View Reference
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DeBenedette MA, Shahinian A, Mak TW, Watts TH. Costimulation of CD28- T lymphocytes by 4-1BB ligand. J Immunol. 1997; 158(2):551-559. (Biology). View Reference
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Hurtado JC, Kim SH, Pollok KE, Lee ZH, Kwon BS. Potential role of 4-1BB in T cell activation. Comparison with the costimulatory molecule CD28. J Immunol. 1995; 155(7):3360-3367. (Biology). View Reference
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Hurtado JC, Kim YJ, Kwon BS. Signals through 4-1BB are costimulatory to previously activated splenic T cells and inhibit activation-induced cell death. J Immunol. 1997; 158(6):2600-2609. (Biology). View Reference
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Kim YJ, Pollok KE, Zhou Z, et al . Novel T cell antigen 4-1BB associates with the protein tyrosine kinase p56lck1. J Immunol. 1993; 151(3):1255-1262. (Biology). View Reference
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Pollok KE, Kim SH, Kwon BS. Regulation of 4-1BB expression by cell-cell interactions and the cytokines, interleukin-2 and interleukin-4. Eur J Immunol. 1995; 25(2):488-494. (Biology). View Reference
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Pollok KE, Kim YJ, Zhou Z, et al. Inducible T cell antigen 4-1BB. Analysis of expression and function. J Immunol. 1993; 150(3):771-781. (Immunogen). View Reference
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Saoulli K, Lee SY, Cannons JL, et al. CD28-independent, TRAF2-dependent costimulation of resting T cells by 4-1BB ligand. J Exp Med. 1998; 187(11):1849-1862. (Biology). View Reference
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Takeda K, Oshima H, Hayakawa Y, et al. CD27-mediated activation of murine NK cells. J Immunol. 2000; 164(4):1741-1745. (Biology). View Reference
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Zhou Z, Pollok KE, Kim KK, Kim YJ, Kwon BS. Functional analysis of T-cell antigen 4-1BB in activated intestinal intra-epithelial T lymphocytes. Immunol Lett. 1994; 41(2-3):177-184. (Biology). View Reference
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.
Refer to manufacturer's instructions for use and related User Manuals and Technical Data Sheets before using this product 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.