Quantification of CD44 Isoforms in Cells

Monoclonal anti-CD44v3, anti-CD44v4,5 and anti-CD44v6 antibodies were purchased from R&D Systems (Minneapolis, MN). Monoclonal anti-CD44v7 and anti-CD44v10 antibodies were purchased from Abcam (Cambridge, MA). Monoclonal anti-CD16 F(ab′)2 and anti-CD32 F(ab′)2 antibody fragments were purchased from Fitzgerald Industries International, Inc (Concord, MA). Monoclonal anti-desmoglein mAb was purchased from Santa Cruz Biotechnology Inc. (Santa Cruz, CA). J10.4 an isotype matched binding control IgG₁ for GM35, the anti-CD11b/CD18 mAb CBRM1/29 and OE-1 the anti-CD55 mAb have been described elsewhere (13 (link), 21 (link), 22 (link)). Protein G Sepharose beads were purchased from GE healthcare. 3′,6′-bis(Acetyloxy)-5(or 6)-(acetyloxy)methoxy]carbonyl-3-oxo-spiroisobenzofuran-1(3H),9′-9Hxanthene]-2′,7′-dipropanoic acid 2′,7′-bis(acetyloxy)methyl ester (BCECF-AM) was purchased from Merck (Damstadt, Germany). Zenon ® Alexa Fluor ® 488 Mouse IgG₁, 568 Mouse IgG₁ and 568 Mouse IgG_2b labeling kits were purchased from Invitrogen Corporation (Carlsbad, CA). Chemotactic peptide N-Formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLF) and 2,2′-azino-bis(3-ethylbenzthiazoline-6-sulphonic acid (ABTS) were purchased from Sigma-Aldrich (St. Louis, MO). Full length CD44 in a pCMv6 entry vector plasmid, CD44 specific short hairpin RNA (shRNA) constructs and a Scr construct plasmid in pGFP-V-RS vectors were purchased from Origene (Rockville, MD). Recombinant human IFNγ was provided by Genentech. Human stdCD44 (CD44s) and human CD44var6 (CD44v6) instant ELISA kits were purchased from Bender Medsystems (Vienna, Austria).