Establishment of a Monoclonal Antibody against CD81 that Decreases the Proliferation of Rat Glioma Cells

Abstract

CD81 belongs to a family of cell-surface proteins (tetraspanins) known to be up-regulated in rheumatoid arthritis (RA) synoviocytes. Recently, we showed that small interfering RNA targeting CD81 (CD81 siRNA) has a therapeutic effect on RA. We also showed that CD81 siRNA decreased the expression of TNF-in SW982 cells, suggesting that TNF-stimulates the expression of synoviolin via CD81. Here, we show that a monoclonal antibody (MAb) against CD81, which was raised by immunization with recombinant CD81, decreased the proliferation of rat C6 glioma cells, showing that this antibody is functionally active and probably inhibits cell proliferation through the inhibition of CD81 function on the cell surface.

Identification of Specific Protein Markers of Rheumatoid Arthritis in Synovial Fluid and Serum

“…To establish anti-CD81 MAb, we previously overexpressed and purified the large outer membrane region of CD81 (LEL) 11,12) . Although this MAb recognizes the LEL region of CD81, we detected the whole molecule (26 kDa) and did not fi nd LEL or smaller molecules in western blotting under reducing conditions, indicating that CD81 exists in its entirety on the membranes of microsomes in RA synovial fl uid, and is not processed or degraded in RA joints.…”

Establishment of Sandwich Enzyme-Linked ImmunoSorbent Assay (ELISA) System for Quantitation of CD81

“…CD81 was originally found as a target of TAPA-1 MAb 10,11) . We have already reported the establishment of anti-CD81 MAbs that are functionally active and inhibit the proliferation of mouse C6 glioma cells 12) . Also, some anti-CD81 MAbs are functionally active in binding to the hepatitis C virus (HCV) cell surface 13) and in the inhibition of proliferation of rat astrocytes 14) .…”

CD81 Mediated Regulation of Synoviolin Expression in Synovial Sarcoma Cells

“…This eluted fraction was concentrated and the titer of IgG1 MAb was analyzed by ELISA. These IgG1 MAbs showed a higher titer to the antigen compared with the titer of antiserum 11,12) .…”

“…After 10-14 days, ascites were collected from these mice. The growth rate of hybridomas and the volume of ascites from four clones is shown elsewhere 11,12) . Relative Luc Activity…”

“…Each fraction eluted from Protein G Sepharose 4 was analyzed used SDS-PAGE 11) . In SDS-PAGE under non-reducing conditions, an IgG1 MAb of about 150 kDa was clearly shown in the eluted fraction.…”