Mixed mode cation exchange-hydrophobic interaction chromatography resins with two different particle sizes were used to reduce host cell protein (HCP), leached protein A, DNA, and aggregates from a mAb stream after a protein A operation.
After labeling, samples were pooled, desalted and concentrated using a mixed mode cation exchange (MCX) cartridge (Waters), and fractionated by preparative isoelectric focusing using a Free Flow Electrophoresis (FFE, BD Biosciences, Inc).
The extraction yields OC with the mixed mode cation exchange sorbent were in most cases close to 100%, regardless of matrix subjected to extraction, and the precision was acceptable with RSD below 21%.
The Strata-X-C (200 mg, 6 mL) mixed mode cation exchange sorbent (Phenomenex, email: internationalphenomenex.com) used for the solid phase extraction (SPE) was conditioned and equilibrated by 2.0 ml of methanol and 2.0 ml of deionized water.
The Strata-X-C (200 mg, 6 mL) mixed mode cation exchange sorbent (Phenomenex, email: [email protected]) used for the solid phase extraction (SPE) was conditioned and equilibrated by 2.0 ml of methanol and 2.0 ml of deionized water.
Usually, polypropylene cartridges with a fixed sorbent bed (e.g., mixed mode cation exchange cartridges) are applied.
Four unit operations comprising of Protein A capture, viral inactivation, flow-through anion exchange (AEX), and mixed-mode cation exchange chromatography (MMCEX) were integrated across two Cadence BioSMB PD multi-column chromatography systems in order to process a 25 L volume of harvested cell culture fluid (HCCF) in less than 12 h.
Resulting peptides were concentrated and purified via mixed-mode cation exchange solid-phase extraction columns (Waters).
Selective clean-up with mixed-mode anion exchange and mixed-mode cation exchange cartridges were tried under basic and acidic conditions, respectively.
Strata-XL-C, mixed-mode cation exchange SPE column showed sufficient retention for all analytes on the RP of the cartridge under acidic condition (pH 3).
The advantage of a mixed-mode cation exchange cartridge is the selective adsorption of basic matrix components at acidic pH via ionic interaction (Tölgyesi et al. 2012).
