Dynamic binding capacity
Each device was sanitized with 1 N NaOH for 30 min at 10 MV/min followed by equilibration with 150 MV binding buffer composed
of 150 mM NaCl in 20 mM Tris/HCl pH 7.3 ± 0.1, conductivity 16 mS/cm. 150 MV of 1 g/L BSA in binding buffer or 0.1 g/L DNA
in binding buffer were loaded. All solutions used were prefiltered with a 0.2 μm membrane filter. All steps were performed
at flow rate of 10 MV/min. Breakthrough curves were recorded by measuring the extinction at 280 nm (protein) and 260 nm (DNA)
using the ÄKTA Explorer. To compare different devices the void volume of the experimental setup was determined by injection
of acetone (2 %). The dynamic binding capacity at 10% breakthrough was calculated as shown in Equation 1,
where V10% is volume loaded at 10% breakthrough, Vv is void volume, Vm is membrane volume, and ci is initial concentration.
Each membrane adsorber device was sanitized with 1 N NaOH for 30 min at 10 MV/min followed by equilibration with 100 MV binding
buffer composed of 150 mM NaCl in 20 mM Tris/HCl pH 7.3 ± 0.1, conductivity 16 mS/cm. Protein throughput was determined using
the pressure vessel filled with a solution of 20 g/L γ-globulin in binding buffer was used to determine the protein throughput
with the membrane adsorber devices. The filtrate volume up to 1000 MV was monitored at a constant pressure of 3 bar using
Chinese hamster ovary proteins clearance
Chinese hamster ovary proteins (CHOP) clearance was determined using industrially relevant mAb feedstock. Before loading the
MAb feedstock onto the membrane adsorber, the membrane was equilibrated with 10 MV of 50 mM Tris buffer at the appropriate
pH. The conductivity of this buffer was adjusted by altering the concentration of sodium acetate. After equilibration, the
mAb feedstock was loaded onto the devices to a targeted load density of 10 kg mAb/L of membrane at a flow rate of 10 MV/min.
Pool fractions were collected during the experiment and analyzed for CHOP concentration.
Determination of log reduction value of bacteriophages
Equipment and membrane devices were sanitized with 1 M sodium hydroxide for 30 minutes. Membrane devices were further equilibrated
with 300 MV of binding buffer. The ΦX174 phage solution with a titer of 1.5x107 PFU/mL was prepared and loaded onto the devices at a flow rate of 10 MV/min. Flow-through fractions were collected after
100 and 150 MV of load for quantitative analysis.
Pump, tubing, and devices were treated with 1 M sodium hydroxide for 30 minutes at room temperature and at a flow rate of
10 MV/min before performing the experiment. Compatible vessels and materials were heated at 200 °C for 4 hours to destroy
naturally occurring endotoxins. After sufficient rinsing with reverse osmosis water, the equilibration was performed with
300 MV of binding buffer. 150 MV of endotoxin in binding buffer were loaded to the membrane at a flow rate of 10 MV/min. The
flow-through was divided into fractions of 50 MV each and was analyzed to determine the endotoxin level.