AMICA, Fc Chimera, Recombinant, Mouse (Adhesion Molecule that Interacts with CXADR Antigen 1, Junction Adhesion Molecule-Like, JAM-L, JAML)

AMICA, also known as JAML, is a 65kD, type I transmembrane glycoprotein that belongs to the junctional adhesion molecule (JAM) subset of the immunoglobulin superfamily (1). JAM family proteins contribute to intercellular connections within epithelial and endothelial cell layers and mediate their interactions with various hemopoietic cells (1). The mouse AMICA cDNA encodes a 379 aa precursor that includes a 20 aa signal sequence, a 261 aa extracellular domain (ECD) with two Ig-like domains, a 21 aa transmembrane segment, and a 77 aa cytoplasmic domain (2). In contrast to other JAM family proteins, AMICA does not contain a cytoplasmic PDZ-binding motif (3). Within the ECD, mouse AMICA shares 58% and 77% aa sequence identity with human and rat AMICA, respectively. It shares 17%-23% aa sequence identity with the ECDs of mouse JAM-A, -B, -C, and JAM4. AMICA is expressed on the surface of granulocytes and monocytes and is upregulated during the differentiation of myeloid leukemia cells (2, 3). A motif in the ECD, which promotes dimerization of other JAM family proteins, is required for surface localization of AMICA (2). AMICA mediates adhesion of monocytes to endothelial cells (2) and neutrophil migration across epithelial cell monolayers (3). This latter function involves specific interactions of AMICA with CXADR in epithelial tight junctions (3). In particular, the membrane proximal Ig-like domain of AMICA binds the membrane-distal Ig-like domain of CXADR (3). AMICA does not appear to interact homophilically, as neutrophils adhere to immobilized CXADR but not to immobilized AMICA (3).\nCD33 Signal Peptide (Met1-Ala16); Mouse AMICA (Leu23-Leu281) IEGRMDP; Mouse IgG2a (Glu98-Lys330)\n \nSource: \nA DNA sequence encoding the extracellular domain of mouse AMICA (Leu 23-Leu 281; Accession # Q80UL9) (Strausberg, R.L. et al., 2002, Proc. Natl. Acad. Sci. USA 99(26):16899) was fused to the signal peptide of human CD33 at the N-terminus and the Fc region of mouse IgG2a at the C-terminus via a polypeptide linker. The protein was expressed in a mouse myeloma cell line, NS0. \n\nMolecular Mass: \nBased on N-terminal sequencing, the recombinant mouse AMICA/Fc begins with Leu 23. The monomer of the recombinant mouse AMICA/mIgG2a/Fc has a calculated molecular mass of approximately 56.6kD. In SDS-PAGE, the recombinant protein migrates as an approximately 68-78kD protein under reducing conditions. \n\nEndotoxin Level: \n< 1.0 EU per 1ug of the protein as determined by the LAL method.\n\nActivity:\nMeasured by the ability of the immobilized protein to support the adhesion of human neutrophils. When 2 x 10e5 cells/well are added to AMICA-coated plates (10ug/ml, 100 microL/well), 30-50% of the cells will adhere after 20 minutes at 37 degrees C. \n\nReconstitution: \nIt is recommended that sterile PBS be added to the vial to prepare a working stock solution of no less than 100ug/ml. The carrier-free protein should be used immediately upon reconstitution to avoid losses in activity due to non-specific binding to the inside surface of the vial. For long term storage as a dilute solution, a carrier protein (e.g. 0.1% HSA or BSA) should be added to the vial. \n\nStorage and Stability:\nLyophilized samples are stable for up to twelve months at -20 degrees C. Upon reconstitution, this protein, in the presence of a carrier protein, can be stored under sterile conditions at 2-8 degrees C for one month or at -20 degrees C in a manual defrost freezer for three months without detectable loss of activity. Avoid repeated freeze-thaw cycles.

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SPECIFICATIONS

Catalog Number

A1372-31

Size

50ug

Form

Supplied as a lyophilized powder in PBS.

Purity

(same/more than) 95%, as determined by SDS-PAGE and visualized by silver stain.

References

1. Mandell, K.J. and C.A. Parkos, 2005, Adv. Drug Deliv. Rev. 57:857. 2. Moog-Lutz, C. et al., 2003, Blood 102:3371. 3. Zen, K. et al., 2005, Mol. Biol. Cell 16:2694.