Streptavidin sa maleimide

Recombinant wild-type (WT) monomeric VWF 1238-A1 (residues 1238-1471), 1261-A1 (residues 1261-1471) and A3 (residues 1687-1874) domains as well as the A1 N-terminal flanking region polypeptide (Lp, residues 1238-1260) were generated by E.coli as previously described [9 (link),11 (link),14 (link)]. Plasma VWF (Calbiochem, San Diego, CA), human placenta type III collagen (Sigma) and streptavidin (SA)-maleimide (Sigma) were purchased. Two anti-GPIbα mAbs were used: AK2 was purchased (Abcam, Cambridge, MA) and WM23 was a gift from Dr. Renhao Li (Emory University, Atlanta, GA). Mouse anti-human anti-A1 mAbs (5D2 and 6G1, IgG1 subtype) were gifts from Dr. Michael Berndt (Curtin University, WA, Australia). The epitope for 5D2 has not been defined and appear to be conformation-dependent; while the epitope for 6G1 has been mapped to a region at the C-terminus (E1463-H1472) of A1 where ristocetin binds [31 (link)]. PE-conjugated goat F(ab′)2 anti-mouse IgG (ab7002) was from Abcam (Cambridge, MA). All other reagents, if not specified, were purchased from Sigma.