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 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 CompBead to ensure that BD Comp beads 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).
For optimal results, it is recommended to perform 2 washes after staining with antibodies. Cells may be prepared, stained with antibodies and washed twice with wash buffer per established protocols for immunofluorescence staining, prior to acquisition on a flow cytometer. Performing fewer than the recommended wash steps may lead to increased spread of the negative population.
Product Notices
- 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.
- 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.
- Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
Companion Products
The OX-79 antibody reacts with CD53, a 35-45-kDa member of the Transmembrane 4-pass protein superfamily (TM4SF). As in the human, mouse peripheral leukocytes express CD53 mRNA and protein, and erythrocytes and nonhematopoietic cells do not. However, the distribution of CD53 in the mouse and rat thymus differs from that in the human. In the mouse, most CD4+8- and major subsets of CD4- CD8- (double-negative) and CD4- CD8+ thymocytes express CD53, while most cortical CD4+ CD8+ (double-positive) thymocytes are CD53-low or CD53-negative. CD53 expression can be induced in double-positive thymocytes by cross-linking of their T-cell receptors with anti-TCR mAb or peptide-pulsed antigen-presenting cells. There is a strong correlation between positive selection of thymocytes and CD53 expression. Its association with CD2 and a tyrosine phosphatase in rat T lymphocytes and with CD19, CD21, HLA-DR, and other TM4SF proteins in human B lymphocytes suggests that CD53 is involved in leukocyte signal transduction.
The antibody was conjugated to BD Horizon BB515 which is part of the BD Horizon Brilliant™ Blue family of dyes. With an Ex Max near 490 nm and an Em Max near 515 nm, BD Horizon BB515 can be excited by the blue laser (488 nm) laser and detected with a 530/30 nm filter. This dye has been exclusively developed by BD Biosciences and is up to seven times brighter than FITC with less spillover into the PE channel. Due to similar excitation and emission properties, BB515, FITC, and Alexa Fluor® 488 cannot be used simultaneously. It is not recommended to use BB515 in cocktails that include Streptavidin conjugates as it may cause high background.
Development References (4)
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Angelisová P, Hilgert I, Horejsí V. Association of four antigens of the tetraspans family (CD37, CD53, TAPA-1, and R2/C33) with MHC class II glycoproteins. Immunogenetics. 1994; 39(4):249-256. (Biology). View Reference
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Carmo AM, Wright MD. Association of the transmembrane 4 superfamily molecule CD53 with a tyrosine phosphatase activity. Eur J Immunol. 1995; 25(7):2090-2095. (Biology). View Reference
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Puls KL, Hogquist KA, Reilly N, Wright MD. CD53, a thymocyte selection marker whose induction requires a lower affinity TCR-MHC interaction than CD69, but is up-regulated with slower kinetics. Int Immunol. 2002; 14(3):249-258. (Biology). View Reference
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Tomlinson MG, Hanke T, Hughes DA, et al. Characterization of mouse CD53: epitope mapping, cellular distribution and induction by T cell receptor engagement during repertoire selection. Eur J Immunol. 1995; 25(8):2201-2205. (Immunogen: Western blot). 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.
