Product Specifications for Recombinant Human Nidogen-1/Entactin Protein, CF
Mouse myeloma cell line, NS0-derived human Nidogen-1/Entactin protein Leu29-Lys1114 (Gln1113Arg), with an N-terminal 9-His tag
>95%, by SDS-PAGE under reducing conditions and visualized by silver stain
<1.0 EU per 1 μg of the protein by the LAL method.
N-terminal sequence Analysis
Predicted Molecular Mass
130 kDa, reducing conditions
Measured by the ability of the immobilized protein to support the adhesion of SVEC4‑10 mouse vascular endothelial cells. When 4 x 104 cells/well are added to rhNidogen-1 coated plates (30 µg/mL with 100 µL/well), approximately 40-75% will adhere after one hour at 37 °C. Optimal dilutions should be determined by each laboratory for each application.
Formulation, Preparation and Storage
What does CF mean?
CF stands for Carrier Free (CF). We typically add Bovine Serum Albumin (BSA) as a carrier protein to our
Adding a carrier protein enhances protein stability, increases shelf-life, and allows the recombinant
protein to be stored at a more dilute concentration.
The carrier free version does not contain BSA.
What formulation is right for me?
In general, we advise purchasing the recombinant protein with BSA for use in cell or tissue culture, or
as an ELISA standard.
In contrast, the carrier free protein is recommended for applications, in which the presence of BSA
Lyophilized from a 0.2 μm filtered solution in PBS.
Reconstitute at 100 μg/mL in sterile PBS.
The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage:
Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
12 months from date of receipt, -20 to -70 °C as supplied.
1 month, 2 to 8 °C under sterile conditions after reconstitution.
3 months, -20 to -70 °C under sterile conditions after reconstitution.
Nidogen-1 (also entactin) is a 150 kDa, secreted, monomeric glycoprotein that serves as a major linking component of basement membranes (1 - 4). It is synthesized as a 1247 amino acid (aa) precursor with a 28 aa signal sequence and a 1219 aa mature protein. The molecule is modular in structure with five distinct regions. There are three globular domains (G1-3) separated by a mucin region and an extended rod-shaped segment (5 - 7). The N-terminal globular domain (G1) is 200 aa in length and seemingly unrelated to any known motif (8). The mucin region is nearly 160 aa in length and presumably O-glycosylated (2, 8). G2 and G3 are both approximately 300 aa in length. G2 is described as a Nidogen ( beta -barrel) domain, while C-terminal G3 assumes a beta -propeller configuration (1). The 250 aa rod-shaped segment has multiple EGF-like motifs and two thyroglobulin type 1 domains. Functionally, G1 is reported to bind type IV collagen (2, 7). The mucin region contains a short peptide that ligates alpha 3 beta 1 integrins (9, 10). G2 interacts with perlecan, and an RGD motif in the rod-shaped segment serves as a binding site for alpha v beta 3 integrins (9, 10). Finally, G3 is associated with laminin binding (2, 7). As a full-length molecule, the multiple extracellular matrix-binding sites of Nidogen-1 are well positioned to serve as anchor sites for basement membrane molecules. Nidogen-1 also undergoes proteolytic processing by at least two MMPs, MMP7 and MMP19 (10, 11). While this destroys the integrity of Nidogen-associated matrices, it also generates peptide fragments that are capable of inducing neutrophil chemotaxis and phagocytosis (10). Nidogen-2 is related to Nidogen-1 (≈ 50% aa identity) and shares many of the same adhesive properties as Nidogen-1 (12). Both bind perlecan plus collagens I and IV. Nidogen-2, however, does not bind fibulin-1 or 2, and shows only modest interaction with laminin. Thus, although coexpressed, Nidogen-2 serves as only a partial substitute for Nidogen-1 (2, 12). Human Nidogen-1 is 85% aa identical to both mouse and rat Nidogen-1, and 88% aa identical to canine Nidogen-1.
Hohenester, E. and J. Engel (2002) Matrix Biol. 21:115.
Miosge, N. et al. (2001) Histochem. J. 33:523.
Charonis, A. et al. (2005) Curr. Med. Chem. 12:1495.
Timpl, R. and J.C. Brown (1996) BioEssays 18:123.
Nagayoshi, T. et al. (1989) DNA 8:581.
Zimmerman, K. et al. (1995) Genomics 27:245.
Fox, J.W. et al. (1991) EMBO J. 10:3137.
Mayer, U. et al. (1995) Eur. J. Biochem. 227:681.
Gresham, H.D. et al. (1996) J. Biol. Chem. 271:30587.
Dong, L-J. et al. (1995) J. Biol. Chem. 270:15383.
Titz, B. et al. (2004) Cell. Mol. Life Sci. 61:1826.
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