Biolc hplc

Measurements were performed on a Dionex BioLC HPLC connected to an 18-angle light-scattering detector and a differential refractometer (DAWN HELEOS-II and Optilab rEX, Wyatt). A Superdex 75 10/300 GL column (GE Healthcare) was used in 20 mM Tris pH 8.0, 50 mM NaCl at a flow rate of 0.75 ml min⁻¹. Sample volumes of 1 ml were injected at a concentration of 1.5 mg ml⁻¹. Samples eluting from the column passed through an in-line DAWN HELEOS-II laser photometer (λ = 658 nm) and an Optilab rEX refractometer with a QELS dynamic light-scattering attachment. Light-scattering intensity and eluent refractive index (concentration) were analysed using ASTRA v. 5.3.4.13 software to give a weight-averaged MM. To determine the detector delay volumes and normalization coefficients for the MALLS detector, a BSA sample (Sigma A-8531) was used as reference.
The SEC-MALLS analysis of MuRF1^B2|CC|COS samples was carried out as above but used a Superdex 200 10/300 prep-grade column equilibrated in 20 mM Tris pH 8.0, 100 mM NaCl. MuRF1^B2|CC|COS was injected at 0.7 mg ml⁻¹.

Measurements were performed on a Dionex BioLC HPLC connected to an 18-angle light scattering detector and a differential refractometer (DAWN HELEOS-II and OPTILab rEX, Wyatt). A Superdex 75 10/300 GL column (GE Healthcare) was used in 20 mm Tris-HCl, 150 mm NaCl, 1 mm CaCl₂, pH 7.5, at a flow rate of 0.75 ml/min. Sample volumes of 1 ml were injected at a concentration of 1.5 mg/ml. Samples eluting from the column passed through an in-line DAWN HELEOS-II laser photometer (λ = 658 nm) and an OPTILab rEX refractometer with a QELS dynamic light scattering attachment. Light scattering intensity and eluant refractive index (concentration) were analyzed using ASTRA version 5.3.4.13 software to give a weight-averaged molecular mass. To determine the detector delay volumes and normalization coefficients for the MALLS detector, a BSA sample (Sigma A-8531) was used as reference.