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PE Rat Anti-Mouse CD45R/B220
PE Rat Anti-Mouse CD45R/B220

Multicolor flow cytometric analysis of CD45R/B220 expression on mouse splenocytes. Mouse splenic leucocytes were stained with APC Hamster Anti-Mouse CD3e antibody (Cat. No. 553066/561826) and either PE Rat IgG2a, κ Isotype Control (Cat. No. 553930; Left Plot) or FITC Rat Anti-Mouse CD45R/B220 antibody (Cat. No. 561878/553090/553089; Right Plot) at 0.06 µg/test. The bivariate pseudocolor density plot showing the correlated expression of CD45R/B220 (or Ig Isotype control staining) versus CD3e was derived from gated events with the forward and side light-scatter characteristics of intact splenic leucocytes. Flow cytometry and data analysis were performed using a BD LSRFortessa™ X-20 Cell Analyzer System and FlowJo™ software. Data shown on this Technical Data Sheet are not lot specific.

Multicolor flow cytometric analysis of CD45R/B220 expression on mouse splenocytes. Mouse splenic leucocytes were stained with APC Hamster Anti-Mouse CD3e antibody (Cat. No. 553066/561826) and either PE Rat IgG2a, κ Isotype Control (Cat. No. 553930; Left Plot) or FITC Rat Anti-Mouse CD45R/B220 antibody (Cat. No. 561878/553090/553089; Right Plot) at 0.06 µg/test. The bivariate pseudocolor density plot showing the correlated expression of CD45R/B220 (or Ig Isotype control staining) versus CD3e was derived from gated events with the forward and side light-scatter characteristics of intact splenic leucocytes. Flow cytometry and data analysis were performed using a BD LSRFortessa™ X-20 Cell Analyzer System and FlowJo™ software. Data shown on this Technical Data Sheet are not lot specific.

Product Details
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BD Pharmingen™
B220; Ly-5; CD45R; LCA; Ptprc; Protein tyrosine phosphatase receptor type C
Mouse (QC Testing), Human (Reported)
Rat IgG2a, κ
Mouse Abelson Leukemia Virus-Induced pre-B tumor cells
Flow cytometry (Routinely Tested)
0.2 mg/ml
AB_394619
Aqueous buffered solution containing ≤0.09% sodium azide.
RUO


Preparation And Storage

The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze. The antibody was conjugated to the dye under optimum conditions and unconjugated antibody and free dye were removed.

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 cell and CompBead to ensure that BD® CompBeads are appropriate for your specific cellular application.

Product Notices

  1. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
  2. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
  3. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  4. 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.
  5. An isotype control should be used at the same concentration as the antibody of interest.
  6. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
561878 Rev. 17
Antibody Details
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RA3-6B2

The RA3-6B2 monoclonal antibody specifically binds to an epitope on the extracellular domain of the transmembrane CD45 glycoprotein which is dependent upon the expression of exon A and specific carbohydrate residues. It is expressed on B lymphocytes at all stages from pro-B through mature and activated B cell, but it is decreased on plasma cells and a subset of memory B cells. The levels of CD45R expression on the B-cell lineage appear to be developmentally regulated. It is also reportedly found on the abnormal T cells involved in the lymphadenopathy of lpr/lpr and gld/gld mutant mice, on lytically active subsets of lymphokine-activated killer cells (NK cells and non-MHC-restricted CTL), on apoptotic T lymphocytes of mice injected with bacterial superantigen, on a population of NK-cell precursors in the bone marrow, and on B-lymphocyte, T-lymphocyte, and macrophage progenitors in fetal liver. The CD45R antigen has been reported not to be on hematopoietic stem cells, naive T lymphocytes, or MHC-restricted CTL. 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. CD45R is commonly used as a pan B-cell marker; however, CD19 expression, detectable by the rat anti-mouse CD19 antibody (clone 1D3), is reported to be more restricted to the B-cell lineage. The rat anti-mouse CD45R antibody (clone RA3-6B2) has been reported to enhance isotype switching during in vitro B-cell responses and to inhibit in vivo B-cell responses. Cross-reaction of the RA3-6B2 clone with activated human T lymphocytes has also been reportedly observed.

561878 Rev. 17
Format Details
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PE
R-Phycoerythrin (PE), is part of the BD family of Phycobiliprotein dyes. This fluorochrome is a multimeric fluorescent phycobiliprotein with excitation maximum (Ex Max) of 496 nm and 566 nm and an emission maximum (Em Max) at 576 nm. PE is designed to be excited by the Blue (488 nm), Green (532 nm) and Yellow-Green (561 nm) lasers and detected using an optical filter centered near 575 nm (e.g., a 575/26-nm bandpass filter). As PE is excited by multiple lasers, this can result in cross-laser excitation and fluorescence spillover on instruments with various combinations of Blue, Green, and Yellow-Green lasers. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
altImg
PE
Yellow-Green 488 nm, 532 nm, 561 nm
496 nm, 566 nm
576 nm
561878 Rev.17
Citations & References
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Development References (18)

  1. Allman DM, Ferguson SE, Cancro MP. Peripheral B cell maturation. I. Immature peripheral B cells in adults are heat-stable antigenhi and exhibit unique signaling characteristics. J Immunol. 1992; 149(8):2533-2540. (Biology). View Reference
  2. Asensi V, Kimeno K, Kawamura I, Sakumoto M, Nomoto K. Treatment of autoimmune MRL/lpr mice with anti-B220 monoclonal antibody reduces the level of anti-DNA antibodies and lymphadenopathies. Immunology. 1989; 68(2):204-208. (Clone-specific). View Reference
  3. Ballas ZK, Rasmussen W. Lymphokine-activated killer cells. VII. IL-4 induces an NK1.1+CD8 alpha+beta- TCR-alpha beta B220+ lymphokine-activated killer subset. J Immunol. 1993; 150(1):17-30. (Biology). View Reference
  4. Bleesing JJ, Morrow MR, Uzel G, Fleisher TA. Human T cell activation induces the expression of a novel CD45 isoform that is analogous to murine B220 and is associated with altered O-glycan synthesis and onset of apoptosis. Cell Immunol. 2001; 213(1):72-81. (Clone-specific). View Reference
  5. Coffman RL. Surface antigen expression and immunoglobulin gene rearrangement during mouse pre-B cell development. Immunol Rev. 1982; 69:5-23. (Biology). View Reference
  6. Domiati-Saad R, Ogle EW, Justement LB. Administration of anti-CD45 mAb specific for a B cell-restricted epitope abrogates the B cell response to a T-dependent antigen in vivo. J Immunol. 1993; 151(11):5936-5947. (Clone-specific). View Reference
  7. Driver DJ, McHeyzer-Williams LJ, Cool M, Stetson DB, McHeyzer-Williams MG. Development and maintenance of a B220- memory B cell compartment. J Immunol. 2001; 167(3):1393-1405. (Biology). View Reference
  8. 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). View Reference
  9. Hardy RR, Carmack CE, Shinton SA, Kemp JD, Hayakawa K. Resolution and characterization of pro-B and pre-pro-B cell stages in normal mouse bone marrow. J Exp Med. 1991; 173(5):1213-1225. (Biology). View Reference
  10. Hathcock KS, Hirano H, Murakami S, Hodes RJ. CD45 expression by B cells. Expression of different CD45 isoforms by subpopulations of activated B cells. J Immunol. 1992; 149(7):2286-2294. (Biology). View Reference
  11. 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.
  12. Kobata T, Takasaki K, Asahara H, et al. Apoptosis with FasL+ cell infiltration in the periphery and thymus of corrected autoimmune mice. Immunology. 1997; 92(2):206-213. (Biology). View Reference
  13. Krop I, de Fougerolles AR, Hardy RR, Allison M, Schlissel MS, Fearon DT. Self-renewal of B-1 lymphocytes is dependent on CD19. Eur J Immunol. 1996; 26(1):238-242. (Biology). View Reference
  14. Laouar Y, Ezine S. In vivo CD4+ lymph node T cells from lpr mice generate CD4-CD8-B220+TCR-beta low cells. J Immunol. 1994; 153(9):3948-3955. (Biology). View Reference
  15. Puzanov IJ, Bennett M, Kumar V. IL-15 can substitute for the marrow microenvironment in the differentiation of natural killer cells. J Immunol. 1996; 157(10):4282-4285. (Biology). View Reference
  16. Renno T, Hahne M, Tschopp J, MacDonald HR. Peripheral T cells undergoing superantigen-induced apoptosis in vivo express B220 and upregulate Fas and Fas ligand. J Exp Med. 1996; 183(2):431-437. (Biology). View Reference
  17. Rolink A, ten Boekel E, Melchers F, Fearon DT, Krop I, Andersson J. A subpopulation of B220+ cells in murine bone marrow does not express CD19 and contains natural killer cell progenitors. J Exp Med. 1996; 183(1):187-194. (Biology). View Reference
  18. Sagara S, Sugaya K, Tokoro Y, et al. B220 expression by T lymphoid progenitor cells in mouse fetal liver. J Immunol. 1997; 158(2):666-676. (Biology). View Reference
View All (18) View Less
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