Dawn helios 2 apparatus

Multi-angle laser light scattering in a Dawn Helios II apparatus (Wyatt Technologies) coupled to a SEC Superose 6 10/300 column (SEC-MALLS) equilibrated in buffer J at 25 °C was performed at the joint IBMB/IRB Crystallography Platform, Barcelona Science Park (Catalonia, Spain) to analyze binding of GRAB or pro-TGF-β₂ to native or induced authentic hα₂M at a molar ration of 4:1. ASTRA 7 software (Wyatt Technologies) was used for data processing and analysis, for which a dn/dc value typical for proteins (0.185 mL/g) was assumed. All experiments were performed in triplicate.

Size-exclusion chromatography coupled to multiple-angle light scattering (SEC-MALS) was performed in a Dawn Helios II apparatus (Wyatt Technologies) coupled to a SEC Superdex 75 Increase 10/3000 column (Cytiva). The column was equilibrated with 30 mM Tris·HCl, 250 mM sodium chloride, pH 8 at 25 °C and operated at a flow rate of 0.5 mL/min. Samples were simultaneously monitored with an UV detector (SPD-20, Shimadzu), a differential refractometer (OPTI-rEx, Wyatt Corp.), and a static multiangle laser light scattering detector (DAWN-HELEOS, Wyatt Corp.). Data processing and analysis proceeded with Astra 7 software (Wyatt Technologies).