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Two-color flow cytometric analysis of CD45RA expression on mouse splenocytes. Splenic leucocytes were preincubated with Purified Rat Anti-Mouse CD16/CD32 antibody (Mouse BD Fc Block™) (Cat. No. 553141/553142). The cells were then stained with FITC Hamster Anti-Mouse CD3e antibody (Cat. No. 553062/553061/561827) and either PerCP-Cy™5.5 Rat IgG2b, κ Isotype Control (Cat. No. 550764; Left Panel) or PerCP-Cy™5.5 Rat Anti-Mouse CD45RA antibody (Cat. No. 564359; Right Panel). Two-color flow cytometric contour plots showing the correlated expression of CD45RA (or Ig Isotype control staining) versus CD3e were derived from gated events with the forward and side light-scatter characteristics of viable leucocytes. Flow cytometric analysis was performed using a BD™ LSR II Flow Cytometer System.
BD Pharmingen™ PerCP-Cy™5.5 Rat Anti-Mouse CD45RA
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- 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.
- Please observe the following precautions: Absorption of visible light can significantly alter the energy transfer occurring in any tandem fluorochrome conjugate; therefore, we recommend that special precautions be taken (such as wrapping vials, tubes, or racks in aluminum foil) to prevent exposure of conjugated reagents, including cells stained with those reagents, to room illumination.
- PerCP-Cy5.5–labelled antibodies can be used with FITC- and R-PE–labelled reagents in single-laser flow cytometers with no significant spectral overlap of PerCP-Cy5.5, FITC, and R-PE fluorescence.
- PerCP-Cy5.5 is optimized for use with a single argon ion laser emitting 488-nm light. Because of the broad absorption spectrum of the tandem fluorochrome, extra care must be taken when using dual-laser cytometers, which may directly excite both PerCP and Cy5.5™. We recommend the use of cross-beam compensation during data acquisition or software compensation during data analysis.
- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
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- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
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The 14.8 monoclonal antibody specifically recognizes an exon A-dependent epitope of the CD45 protein, which is found at high density on B cells and at low density on peripheral T cytotoxic/suppressor cells and a very small subset of thymocytes. Nearly all B-lineage cells, including B-cell precursors in fetal liver and adult bone marrow and Ig-secreting cells, but not hematopoietic stem cells or myeloid progenitors, have been reported to be detectable by mAb 14.8. CD45 is a member of the Protein Tyrosine Phosphatase (PTP) family: Its intracellular (COOH-terminal) region contains two PTP catalytic domains, and the extracellular region is highly variable due to alternative splicing of exons 4, 5, and 6 (designated A, B, and C, respectively), plus, differing levels of glycosylation. The CD45 isoforms detected in the mouse are cell type-, maturation-, and activation state-specific. The CD45 isoforms play complex roles in T-cell and B-cell antigen receptor signal transduction. mAb 14.8 has been reported to enhance the proliferative effect of PHA on purified spleen T cells, possibly by replacing a signal normally delivered by accessory cells, to enhance isotype switching during in vitro B-cell responses, and to inhibit antigen-induced p21 [ras] activation.
研发参考 (10)
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George A, Rath S, Shroff KE, Wang M, Durdik JM. Ligation of CD45 on B cells can facilitate production of secondary Ig isotypes. J Immunol. 1994; 152(3):1014-1021. (Clone-specific: Functional assay, Stimulation). 查看参考
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Goff LK, Huby RD. Characterization of constitutive and strain-dependent subsets of CD45RA+ cells in the thymus. Int Immunol. 1992; 4(11):1303-1311. (Biology). 查看参考
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Hathcock KS, Laszlo G, Dickler HB, et al. Expression of variable exon A-, B-, and C-specific CD45 determinants on peripheral and thymic T cell populations. J Immunol. 1992; 148(1):19-28. (Clone-specific: Flow cytometry). 查看参考
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Inoue T, Asano Y, Matsuoka S, et al. Distinction of mouse CD8+ suppressor effector T cell clones from cytotoxic T cell clones by cytokine production and CD45 isoforms. J Immunol. 1993; 150(6):2121-2128. (Clone-specific: Flow cytometry). 查看参考
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Johnson P, Maiti A, Ng DHW. CD45: A family of leukocyte-specific cell surface glycoproteins. In: Herzenberg LA, Weir DM, Herzenberg LA, Blackwell C , ed. Weir's Handbook of Experimental Immunology, Vol 2. Cambridge: Blackwell Science; 1997:62.1-62.16.
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Kawauchi K, Lazarus AH, Rapoport MJ, Harwood A, Cambier JC, Delovitch TL. Tyrosine kinase and CD45 tyrosine phosphatase activity mediate p21ras activation in B cells stimulated through the antigen receptor. J Immunol. 1994; 152(7):3306-3316. (Clone-specific: Blocking, Functional assay, Immunoprecipitation). 查看参考
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Kincade PW, Lee G, Watanabe T, Sun L, Scheid MP. Antigens displayed on murine B lymphocyte precursors. J Immunol. 1981; 127(6):2262-2268. (Clone-specific: Depletion, Flow cytometry). 查看参考
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Kincade PW. Formation of B lymphocytes in fetal and adult life. Adv Immunol. 1981; 31:177-245. (Immunogen). 查看参考
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Marvel J, Poirier G, Lightstone E. Anti-CD45RA antibodies increase the proliferation of mouse T cells to phytohemagglutinin through the interleukin 2/interleukin 2 receptor pathway. Eur J Immunol. 1989; 19(11):2005-2010. (Biology). 查看参考
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Rogers PR, Pilapil S, Hayakawa K, Romain PL, Parker DC. CD45 alternative exon expression in murine and human CD4+ T cell subsets. J Immunol. 1992; 148(12):4054-4065. (Clone-specific: Flow cytometry). 查看参考
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For Research Use Only. Not for use in diagnostic or therapeutic procedures.