Purification of Ecoil-tagged TZMs

Filtered supernatants were loaded on MabSelect SuRe (GE Healthcare, cat# 17–5438–02) columns equilibrated in PBS. The columns were washed with PBS, and proteins were eluted with 100 mM citrate buffer (pH 3.6). The fractions containing eluted chimeric mAbs were pooled and elution buffer was exchanged for PBS using CentriPure P-100 desalting columns (emp Biotech GmbH, Germany). Final mAb titers in culture supernatants were determined by protein-A HPLC using an 800 μl POROS 20 μm Protein A ID Cartridge (Applied Biosystems, cat# 2–1001–00) according to the manufacturer’s recommendations. Purified proteins in PBS were quantified by absorbance at 280 nm using a Nanodrop spectrophotometer (Thermo Fisher Scientific) and the calculated extinction coefficient for each protein (ProtParam tool – Expasy). Purified mAbs were sterilized by passing through 0.2 mm filters, aliquoted, and stored at −80°C.