HPLC-SEC Analysis of OAg Molecular Size

HPLC—SEC analysis was used to estimate the molecular size distribution of OAg populations [34 (link),36 (link)]. Samples were run, without pre-treatment, on a TSK gel G3000 PWXL column (30 cm x_7.8 mm; particle size 7 um; cod. 808021) with a TSK gel PWXL guard column (4.0 cm _x 6.0 mm; particle size 12 um; cod. 808033) (Tosoh Bioscience, Tokyo, Japan). The mobile phase was 0.1 M NaCl, 0.1 M NaH₂PO₄, 5% CH₃CN, pH 7.2, at the flow rate of 0.5 ml/min (isocratic method for 30 min). OAg peaks were detected by differential refractive index (dRI). Void and bed volume calibration was performed with λ-DNA (λ-DNA molecular weight (MW) Marker III 0.12–21.2 kb; Roche) and sodium azide (Merck, New Jersey, USA), respectively. OAg average MW was estimated on standard dextrans (Sigma) calibration curve.