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BB700 Mouse Anti-Mouse CD45.2
BB700 Mouse Anti-Mouse CD45.2
Flow cytometric analysis of CD45.2 expression on C57BL/6 mouse splenocytes. Splenic leucocytes from an SJL mouse (Left Plot) or a C57BL/6 mouse (Right Plot) were preincubated with Purified Rat Anti-Mouse CD16/CD32 antibody (Mouse BD Fc Block™) (Cat. No. 553141/553142). The cells were then stained with either BD Horizon™ BB700 Mouse IgG2a, κ Isotype Control (Cat. No. 566419; dashed line histograms) or BD Horizon BB700 Mouse Anti-Mouse CD45.2 antibody (Cat. No 566496/566497; solid line histogram) at 1 µg/test. BD Pharmingen™ DAPI Solution (Cat. No. 564907) was added to cells right before analysis. The fluorescence histogram showing CD45.2 expression (or Ig Isotype control staining) was derived from DAPI negative-gated events with the forward and side light-scatter characteristics of viable leucocytes. Flow cytometric analysis was performed using a BD FACSCelesta™ Flow Cytometer System. Data shown on this Technical Data Sheet are not lot specific.
Flow cytometric analysis of CD45.2 expression on C57BL/6 mouse splenocytes. Splenic leucocytes from an SJL mouse (Left Plot) or a C57BL/6 mouse (Right Plot) were preincubated with Purified Rat Anti-Mouse CD16/CD32 antibody (Mouse BD Fc Block™) (Cat. No. 553141/553142). The cells were then stained with either BD Horizon™ BB700 Mouse IgG2a, κ Isotype Control (Cat. No. 566419; dashed line histograms) or BD Horizon BB700 Mouse Anti-Mouse CD45.2 antibody (Cat. No 566496/566497; solid line histogram) at 1 µg/test. BD Pharmingen™ DAPI Solution (Cat. No. 564907) was added to cells right before analysis. The fluorescence histogram showing CD45.2 expression (or Ig Isotype control staining) was derived from DAPI negative-gated events with the forward and side light-scatter characteristics of viable leucocytes. Flow cytometric analysis was performed using a BD FACSCelesta™ Flow Cytometer System. Data shown on this Technical Data Sheet are not lot specific.
Product Details
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BD Horizon™
Ly-5.2; T200; LCA; Leukocyte common antigen; Ptprc
Mouse (QC Testing)
Mouse SJL IgG2a, κ
B10.S mouse thymocytes and splenocytes
Flow cytometry (Routinely Tested)
0.2 mg/ml
19264
AB_2744398
Aqueous buffered solution containing ≤0.09% sodium azide.
RUO


Preparation And Storage

Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze. The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. The antibody was conjugated with BD Horizon™ BB700 under optimum conditions, and unconjugated antibody and free BD Horizon BB700 were removed.

Recommended Assay Procedures

     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 for the BD Horizon Brilliant Stain Buffer (Cat. No. 563794/566349) or the BD Horizon Brilliant Stain Buffer Plus (Cat. No. 566385).

     When setting up compensation, it is recommended to compare spillover values obtained from cells and BD™ CompBeads to ensure that beads will provide sufficiently accurate spillover values.

     For optimal results, it is recommended to perform two washes after staining with antibodies. Cells may be prepared, stained with antibodies and washed twice with wash buffer per established protocols for immunofluorescent 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

  1. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
  2. An isotype control should be used at the same concentration as the antibody of interest.
  3. 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.
  4. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  5. The manufacture, use, sale, offer for sale, or import of this product is subject to one or more patents or pending applications. This product, and only in the amount purchased by buyer, may be used solely for buyer’s own internal research, in a manner consistent with the accompanying product literature. No other right to use, sell or otherwise transfer (a) this product, or (b) its components is hereby granted expressly, by implication or by estoppel. Diagnostic uses require a separate license.
  6. 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.
  7. BD Horizon Brilliant Blue 700 is covered by one or more of the following US patents: 8,455,613 and 8,575,303.
  8. Cy is a trademark of GE Healthcare.
  9. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
566496 Rev. 1
Antibody Details
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104

The 104 monoclonal antibody recognizes the CD45 (Leukocyte Common Antigen) present on all leucocytes of most mouse strains (eg, A, AKR, BALB/c, CBA/Ca, CBA/J, C3H/He, C57BL, C57BR, C57L, C58, DBA/1, DBA/2, NZB, SWR, 129). This alloantigen was originally named Ly-5.1, and this was the designation at the time that the antibody was characterized. The designation was later changed from Ly-5.1 to Ly-5.2 to conform with the convention that the .2 alloantigen designations be assigned to the C57BL/6 strain. mAb 104 has been reported not to react with leucocytes of the mouse strains expressing the CD45.1 alloantigen (eg, RIII, SJL/J, STS/A, and DA). 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. The 104 antibody has been reported to inhibit some responses of B cells, from mice expressing the CD45.2 alloantigen, to certain antigens and LPS.  In addition, reduction of serum IgG levels and amelioration of autoimmune renal pathology were reported in mAb 104-treated systemic lupus erythematosus-prone mice.

The antibody was conjugated to BD Horizon BB700, which is part of the BD Horizon Brilliant™ Blue family of dyes.   It is a polymer-based tandem dye developed exclusively by BD Biosciences.  With an excitation max of 485 nm and an emission max of 693 nm, BD Horizon BB700 can be excited by the 488 nm laser and detected in a standard PerCP-Cy™5.5 set (eg, 695/40-nm filter). This dye provides a much brighter alternative to PerCP-Cy5.5 with less cross laser excitation off the 405 nm and 355 nm lasers.

566496 Rev. 1
Format Details
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BB700
The BD Horizon Brilliant™ Blue 700 (BB700) Dye is part of the BD Horizon Brilliant™ Blue family of dyes. This tandem fluorochrome is comprised of a polymer-technology dye donor with an excitation maximum (Ex Max) of 476-nm and an acceptor dye with an emission maximum (Em Max) at 695-nm. Driven by BD innovation, BB700 is designed to be excited by the blue laser (488-nm) and detected using an optical filter centered near 695-nm (e.g., a 695/20-nm bandpass filter). The donor dye can be excited by the Violet (405 nm) laser and the acceptor dye can be excited by the red (627–640 nm) laser resulting in cross-laser excitation and fluorescence spillover. BB700 Reagents are significantly brighter than equivalent PerCP or PerCP-Cy5.5 reagents and are less sensitive to photobleaching. In addition, BB700 shows much less excitation by the violet (407-nm) laser resulting in less spillover. BB700 has minimal yellow green (562-nm) excitation and is ideal for instruments with both blue (488-nm) and yellow green (562-nm) lasers. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
altImg
BB700
Blue 488 nm
476 nm
695 nm
566496 Rev.1
Citations & References
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Development References (9)

  1. Johnson P, Greenbaum L, Bottomly K, Trowbridge IS. Identification of the alternatively spliced exons of murine CD45 (T200) required for reactivity with B220 and other T200-restricted antibodies. J Exp Med. 1989; 169(3):1179-1184. (Biology). View Reference
  2. Morse HC 3rd, Shen FW, Hammerling U. Genetic nomenclature for loci controlling mouse lymphocyte antigens. Immunogenetics. 1987; 25(2):71-78. (Biology). View Reference
  3. Ogimoto M, Mizuno K, Tate G, et al. Regulation of lipopolysaccharide- and IL-4-induced immunoglobulin heavy chain gene activation: differential roles for CD45 and Lyb-2. Int Immunol. 1992; 4(6):651-659. (Biology). View Reference
  4. Shen FW, Tung JS, Boyse EA. Further definition of the Ly-5 system. Immunogenetics. 1986; 24(3):146-149. (Biology). View Reference
  5. Shen FW. Monoclonal antibodies to mouse lymphocyte differentiation alloantigens. In: Hammerling GJ, Hammerling U, Kearney JF, ed. Monoclonal Antibodies and T-cell Hybridomas; Perspectives and Technical Advances. 1981:25-31.
  6. Yakura H, Ashida T, Kawabata I, Katagiri M. Alleviation of autoimmunity in BXSB mice by monoclonal alloantibody to Ly-5 (CD45). Eur J Immunol. 1989; 19(8):1505-1508. (Clone-specific: Inhibition, In vivo exacerbation). View Reference
  7. Yakura H, Kawabata I, Ashida T, Katagiri M. Differential regulation by Ly-5 and Lyb-2 of IgG production induced by lipopolysaccharide and B cell stimulatory factor-1 (IL-4). J Immunol. 1988; 141(3):875-880. (Clone-specific: Inhibition). View Reference
  8. Yakura H, Kawabata I, Shen FW, Katagiri M. Selective inhibition of lipopolysaccharide-induced polyclonal IgG response by monoclonal Ly-5 antibody. J Immunol. 1986; 136(8):2729-2733. (Clone-specific: Inhibition). View Reference
  9. Yakura H, Shen FW, Bourcet E, Boyse EA. On the function of Ly-5 in the regulation of antigen-driven B cell differentiation. Comparison and contrast with Lyb-2. J Exp Med. 1983; 157(4):1077-1088. (Clone-specific: Inhibition). View Reference
View All (9) View Less
566496 Rev. 1

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For Research Use Only. Not for use in diagnostic or therapeutic procedures.