>95%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.
<0.10 EU per 1 μg of the protein by the LAL method.
Measured by the ability of the immobilized protein to support the adhesion of human red blood cells. Subramanian, S. et al. (2006) Blood 107:2548. Red blood cells, added at 10 x 106 cells/mL (100 µL/well) to SIRP alpha coated plates, will adhere after 1 hour at 37 °C. The ED50 for this effect is 0.6-2.4 µg/mL. Optimal dilutions should be determined by each laboratory for each application.
Chinese Hamster Ovary cell line, CHO-derived human SIRP alpha/CD172a protein
Human SIRP alpha (Gly27-Arg370) & (Glu31-Arg370) Accession # NP_542970
Both immobilized unlabeled Recombinant Human SIRP alpha /CD172a Fc Chimera (Catalog # 4546-SA) and Biotinylated Recombinant Human SIRP alpha/CD172a Fc Chimera (Catalog # BT4546) support the adhesion of human red blood cells. The ED50 for this effect is 0.6-2.4 μg/mL. The similarity in activity highlights that the biotinylated protein is fully functional.
Background: SIRP alpha/CD172a
regulatory protein alpha (SIRP alpha, designated CD172a), also called SHPS-1 (SHP
substrate 1) and previously, MyD-1 (Myeloid/Dendritic-1), is a monomeric ~90 kDa type I transmembrane glycoprotein that belongs to the SIRP/SHPS (CD172)
family of the immunoglobulin superfamily (1-4). SIRPs are paired receptors,
with similar extracellular domains but differing C-termini and functions (1, 2). The 503 amino acid (aa) human SIRP alpha contains a 342 aa extracellular domain
(ECD), with one V-type, and two C1 type Ig domains, and three potential N
glycosylation sites. It has a 110 aa cytoplasmic sequence with ITIM motifs that
recruit tyrosine phosphatases SHP-1 and SHP-2 when phosphorylated (4). Human
SIRP alpha has more than 40 described polymorphisms, including the prominent
BIT (Brain Ig like molecule with Tyrosine-based activation motifs, also called
SIRP alpha 2 or PTPNS) (5). One reported isoform lacks aa 1-101,
which eliminates most of the V type Ig domain. Human SIRP alpha ECD shares 61%, 60%,
71%, 72% and 73% aa identity with mouse, rat, porcine, bovine and
equine SIRP alpha, respectively; it shares 84% and 76% aa identity with human
SIRP beta 1 and SIRP gamma, respectively (2). SIRP alpha is expressed mainly on myeloid cells,
including macrophages, neutrophils, dendritic and Langerhans cells (3-6). It is also found on neurons, smooth muscle and endothelial cells
(7-9). SIRP alpha shows adhesion to the ubiquitous CD47/IAP (integrin
associated protein), while SIRP gamma binds more weakly and SIRP alpha 1 does not bind at
all (1, 2). Mouse and human SIRP alpha -CD47 binding only cross-reacts for specific
polymorphisms and influences engraftment of xenotransplanted stem cells (6,
10). SIRP alpha engagement generally produces a negative regulatory signal (4). Low
SIRP alpha recognition of CD47, which occurs on aged erythrocytes or platelets or
xenogenic cells, promotes clearance of CD47low cells from
circulation (11, 13). SIRP alpha recognition of surfactants SP-A and SP-D in the
lung can inhibit alveolar macrophage cytokine production (14). The CD47
integrin-SIRP alpha interaction is reported to promote macrophage fusion during
Barclay, A.N. & M.H. Brown (2006) Nat. Rev. Immunol. 6:457.
vanBeek, E.M. et al. (2005) J. Immunol. 175:7781.
Liu, Y. et al. (2005) J. Biol. Chem. 280:36132.
Kharitonenkov, A. et al. (1997) Nature 386:181.
Swissprot Accession # P78324.
Miyashita, M. et al. (2004) Mol. Biol. Cell 15:3950.
Wang, X.X. & K.H. Pfenninger (2005) J. Cell Sci. 119:172.
Maile, L.A. et al. (2003) Mol. Biol. Cell 14:3519.
Johansen, M.L. & E.J. Brown (2007) J. Biol. Chem. 282:24219.
Takenaka, K. et al. (2007) Nat. Immunol. 8:1313.
Ishikawa-Sekigami, T. et al. (2006) Biochem. Biophys. Res. Commun. 343:1197.
Olsson, M. et al. (2005) Blood 105:3577.
Ide, K. et al. (2007) Proc. Natl. Acad. Sci. USA 104:5062.
Gardai, S.J. et al. (2003) Cell 115:13.
Lundberg, P. et al. (2007) Biochem. Biophys. Res. Commun. 352:444.
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