Dynabeads myone carboxylic acid beads

Recombinantly-expressed RBD was covalently loaded onto Dynabeads^® MyOne^™ Carboxylic Acid beads (Thermo 65011, Thermo 65012) according to the manufacturer-provided protocol for “Two-Step Coating Procedure using NHS”. In brief, for a single preparation, 1.5 mL of bead suspension was dispensed and washed twice with 1.5 mL of 25 mM MES, pH 6.0 for 10 min. at room temperature. Following washes, bead chemistry was activated by suspending the beads into 1.5 mL of freshly-prepared 50 mg/mL N-Hydroxysuccinimide (NHS), mixing in 1.5 mL of freshly-prepared 50 mg/mL 1-Ethyl-3-(3dimethylaminopropyl) carbodiimide (EDC), and incubating for 30 min. at room temperature. Following activation, the beads were again washed twice with 1.5 mL of 25 mM MES, pH 6.0 for 10 min. at room temperature. Activated beads were suspended into 1 mL 25 mM MES, pH 6.0 to which 1.5 mg of RBD protein was added. The sample was mixed and incubated for 30 min. at room temperature. Next, the beads were pulled down, the supernatant was discarded, and loading was quenched by incubating the beads in 1 mL of 50 mM Tris, pH 6.8, for 15 min. at room temperature. After quenching, beads were washed four times in 1 mL 1x PBS + 0.1% human serum albumin (HSA), and finally suspended into 3 mL 1x PBS + 0.1% HSA for storage at 4°C.