Size-Exclusion Chromatography of Antibodies

The purity and structure of the antibodies were analyzed by Superdex 200 Increase 10/300 GL chromatography (GE Healthcare, Chicago, IL, USA) using methods previously established by our laboratory (31 (link)). The standard proteins Ferritin, Aldolase, Conalbumin, Ovalbumin, Carbonic anhydrase and Ribonuclease were used for calibration. A 500 μL sample mix containing the above proteins was loaded into the column and separated by the ÄKTA explorer machine (GE Healthcare, Chicago, IL, USA). For the antibody analysis, 100 μL of filtered antibodies 17 and 3G12 (2mg/mL) in 1 × DPBS (Dulbecco’s phosphate-buffered saline, Gibco, Waltham, MA, USA) were analyzed. Antibodies were eluted by DPBS buffer at a flow rate of 0.5 mL/min.