N α maleimidoacetoxy succinimide ester amas

Amino-dextran was synthesized as previously reported (Nakamura et al., 2010 (link)). The synthesis and chemical characterization of dextran conjugates were described in details in our previous paper (Morimoto et. al., Bioconjugate Chem. 2014). In short, dextran (500 mg, average molecular weight 35,000–45,000, from Leuconostoc mesenteroids, Sigma) was dissolved in 50 mL of anhydrous DMF and treated with 126 mg of N,N′-carbonyldiimidazole and 250 µL of ethylenediamine. Aminodextran was treated with biotin-NHS ester in DMSO at 4 °C for overnight to conjugate biotin. COPA or peptide ligands were conjugated to the biotinylated-dextran polymer through N-(α-Maleimidoacetoxy) succinimide ester (AMAS, from Thermo scientific) in DMSO for two hours in the dark. The unreacted reactants were removed by filtering through Amicon Ultra 4 mL centrifugal filter unit (10,000 NMWL, from Millipore). The ligand-dextran conjugate in the filter unit was recovered in 1×PBS according to the manufacturer's protocol. The concentration of biotin in the solution was quantitatively determined using the Fluorescence Biotin Quantitation Kit (Thermo Scientific). The procedure to estimate the number of ligands conjugated to dextran polymer is described in details in Supplemental experimental procedures.