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Multiparameter flow cytometric analysis of CD172a/b expression on human peripheral blood leucocyte populations. Human whole blood was stained with either Alexa Fluor™ 488 Mouse IgG1, κ Isotype Control (Cat. No. 567121; Left Plot) or Alexa Fluor™ 488 Mouse Anti-Human CD172a/b antibody (Cat. No. 567973/568065; Right Plot). Erythrocytes were lysed with BD FACS™ Lysing Solution (Cat. No. 349202). The bivariate pseudocolor density plot showing the correlated expression of CD172a/b [or Ig Isotype control staining] versus side light-scatter (SSC-A) signals was derived from gated events with the forward and side light-scatter characteristics of intact leucocyte populations. Flow cytometry and data analysis were performed using a BD FACSymphony™ A5 Cell Analyzer System and FlowJo™ software.
BD Pharmingen™ Alexa Fluor™ 488 Mouse Anti-Human CD172a/b
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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. This will ensure that BD® CompBeads are appropriate for your specific cellular application.
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- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
- Alexa Fluor® 488 fluorochrome emission is collected at the same instrument settings as for fluorescein isothiocyanate (FITC).
- Source of all serum proteins is from USDA inspected abattoirs located in the United States.
- 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.
- This reagent has been pre-diluted for use at the recommended Volume per Test. We typically use 1 × 10^6 cells in a 100-µl experimental sample (a test).
- An isotype control should be used at the same concentration as the antibody of interest.
- 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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The SE5A5 monoclonal antibody specifically binds to a common epitope on CD172a/SIRPα (90 kDa) and CD172b/SIRPβ1 (50 kDa). These transmembrane glycoproteins are members of the Signal Regulatory Protein (SIRP) family that, in turn, belongs to the Immunoglobulin superfamily. The SIRP family is comprised of two subgroups, SIRPα and SIRPβ that are distinguished by the presence (α) or absence (β) of a cytoplasmic domain containing two immunoreceptor tyrosine-based inhibition motifs (ITIM). CD172a/SIRPα is expressed on CD34+ stem/progenitor cells, cardiomyocytes, monocytes, macrophages, granulocytes, dendritic cells, and in the central nervous system. It binds to CD47 and is implicated in mediating inhibitory signals via the ITIM/SHP-2 association. CD172b/SIRPβ1 does not possess a cytoplasmic domain but instead the transmembrane domain contains a positively-charged residue that can interact with another transmembrane protein, DAP-12/KARAP. DAP-12 has two immunoreceptor tyrosine-based activation motifs (ITAM) within its cytoplasmic domain that are thought to link CD172b to cellular activation signaling. CD172b is expressed on myeloid cells, including peripheral blood monocytes and granulocytes. It is not expressed on CD34+ cells. CD172a and CD172b have complementary roles in signal regulation and may work together in tuning certain cellular responses to stimuli.
Development References (7)
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Bühring HJ, Simmons DL, Vernon-Wilson E. Review—CD172—SIRP; signal regulatory protein. In: Mason D. David Mason .. et al., ed. Leucocyte typing VII : white cell differentiation antigens : proceedings of the Seventh International Workshop and Conference held in Harrogate, United Kingdom. Oxford: Oxford University Press; 2002:35.
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Dietrich J, Cella M, Seiffert M, Bühring HJ, Colonna M. Cutting edge: signal-regulatory protein beta 1 is a DAP12-associated activating receptor expressed in myeloid cells. J Immunol. 2000; 164(1):9-12. (Biology). View Reference
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Dubois NC, Craft AM, Sharma P, et al. SIRPA is a specific cell-surface marker for isolating cardiomyocytes derived from human pluripotent stem cells. Nat Biotechnol. 2011; 29:1011-1018. (Biology). View Reference
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Ghannadan M, Hauswirth AW, Schernthaner GH, et al. Detection of novel CD antigens on the surface of human mast cells and basophils. Int Arch Allergy Immunol. 2002; 127(4):299-307. (Biology: Flow cytometry). View Reference
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Seiffert M, Brossart P, Cant C, et al. Signal-regulatory protein alpha (SIRPalpha) but not SIRPbeta is involved in T-cell activation, binds to CD47 with high affinity, and is expressed on immature CD34(+)CD38(-) hematopoietic cells.. Blood. 2001; 97(9):2741-9. (Clone-specific: Immunoprecipitation, Inhibition). View Reference
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Seiffert M, Cant C, Chen Z, et al. Human signal-regulatory protein is expressed on normal, but not on subsets of leukemic myeloid cells and mediates cellular adhesion involving its counterreceptor CD47. Blood. 1999; 94(11):3633-3643. (Immunogen: Flow cytometry, Functional assay, Immunoprecipitation, Inhibition). View Reference
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Simmons DL, Vernon-Wilson E. Structure and function of the signal regulatory proteins (SIRPs). In: Mason D. David Mason .. et al., ed. Leucocyte typing VII : white cell differentiation antigens : proceedings of the Seventh International Workshop and Conference held in Harrogate, United Kingdom. Oxford: Oxford University Press; 2002:35-38.
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