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
- 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.
- 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.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
- Since applications vary, each investigator should titrate the reagent to obtain optimal results.
- 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.
- 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 http://regdocs.bd.com to access safety data sheets (SDS).
- For U.S. patents that may apply, see bd.com/patents.
Companion Products
The monoclonal antibodies TH6 and 12A5 recognize Folate Receptor 4 (FR4), also known as the membrane folate-binding protein 3 (FBP3). FR4 is a heavily glycosylated 35 kD receptor expressed exclusively in lymphoid tissue and an isoform of the family of receptors that recognize the essential nutrient folic acid. Natural T regulatory cells constitutively express high levels of FR4. Differential expression of FR4 in combination with CD25 can distinguish four functionally distinct CD4+ T cell subpopulations; Natural Tregs, effector T cells, memory-like T cells and Naïve T cells. FR4hi CD25+ expressing CD4+ T cells also express high amounts of Foxp3, GITR and CTLA-4.
Monoclonal antibody TH6 and 12A5 stained CD25+CD4+ T cells at a higher level than other CD4+ or CD8+ T cells. In addition, in vivo injection of TH6 monoclonal antibody reduced the number of CD25+CD4+ T cells and CD25-CD4+ T cells in peripheral blood. Clone 12A5 has been demonstrated to work in western blot. Clones TH6 and 12A5 do not block one another in a flow cytometric assay.
Development References (3)
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Rothberg KG, Ying YS, Kolhouse JF, Kamen BA, Anderson RG. The glycophospholipid-linked folate receptor internalizes folate without entering the clathrin-coated pit endocytic pathway. J Cell Biol. 1990; 110(3):637-649. (Biology). View Reference
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Spiegelstein O, Eudy JD, Finnell RH. Identification of two putative novel folate receptor genes in humans and mouse. Gene. 2000; 258(1-2):117-125. (Biology). View Reference
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Yamaguchi T, Hirota K, Nagahama K, et al. Control of immune responses by antigen-specific regulatory T cells expressing the folate receptor.. Immunity. 2007; 27(1):145-59. (Immunogen: Flow cytometry, Western blot). View Reference
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For Research Use Only. Not for use in diagnostic or therapeutic procedures.