Cochlin, also known as Coch-5B2, is an N-glycosylated protein that constitutes the major protein component of inner ear extracellular matrix. The predominant species in human, mouse, rat, and bovine are the 63kD full length protein, two 40 and 46kD isoforms that lack the LCCL domain, and a 16kD isoform that lacks both VWF-A domains. Cochlin is also susceptible to proteolysis between the LCCL and VWF-A1 domains. Mature full length human Cochlin shares 96% aa sequence identity with bovine, mouse, and rat Cochlin. Cochlin is secreted by inner ear fibrocytes and accumulates in acidophilic deposits of the cochlea and vestibule. Several point mutations in the LCCL domain are associated with the autosomal hearing loss disorder DFNA9, and the resulting mutant proteins exhibit altered aggregation and matrix association properties. The 63 and 16kD isoforms are also present in perilymph fluid. Cochlin is a target of IFNg producing T cells in autoimmune sensorineural hearing loss (ASNHL). Cochlin associates with the choline transporter CTL2 which itself is a target of autoimmune-mediated hearing loss. Cochlin, absent in normal trabecular meshwork (TM) of the eye, is upregulated in glaucoma and is deposited around TM cells. It promotes the in vitro aggregation of TM cells and TM cell adhesion to collagen. Polymorphisms within the second VFW-A domain are associated with glaucoma.
A DNA sequence encoding the mature human Cochlin Glu25-Gln550 was fused with a 6X-His tag at the N-terminus and expressed in a mouse myeloma cell line, NS0. Swiss/UniProt Accession: O43405.
Molecular Mass:Based on N-terminal amino acid sequencing, the recombinant human Cochlin starts with 6 histidines followed by Glu25 and has a calculated molecular mass of 58kD. The recombinant protein migrates as an approximately 64-66kD protein in SDS-PAGE under reducing conditions.
Activity:Measured by its ability to support the adhesion of SW1353 cells (human chondrosarcoma cells, ATCC: HTB-94). When 5x10e4 cells per well were added to rhCochlin coated plates (10ug/ml, 100ul/well) ~70-90% will adhere after 60 minutes at 37ºC. The protein also enhances IFNg secretion by anti-CD3 activated human T cells.
Storage and Stability:Lyophilized and reconstituted products are stable for 12 months after receipt at -20°C. Reconstitute with sterile PBS. Aliquot to avoid repeated freezing and thawing. Store at -20°C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.
仕様
Size：50ug
Source Antigen：Recombinant, A DNA sequence encoding the mature human Cochlin (Glu25-Gln550; Accession #O43405) was fused with a 6X-his tag at the N-terminus and expressed in a mouse myeloma cell line, NS0.
Grade：Purified
Purity：≥90% by SDS-PAGE under reducing conditions and visualized by silver stain. Endotoxin: ≤0.1EU/1ug (LAL)
Form：Supplied as a lyophilized powder from PBS. Reconstitute with 500ul sterile PBS.
