RESULTSįor all conditions tested, the chromatography medium was saturated with protein after 45 minutes of incubation (Figure 1). Buffers were identical to those used in the plate-based experiments. Columns used were tricorn™ 5/100 with 2 mL column volume. The binding capacity ( q) was calculated thusly: q = (C initial – C flowthrough) × V sample/V medium.Ĭolumn Experiments: The dynamic binding capacity at 10% breakthrough (QB10%) at 2 minutes residence time (∼90 minutes loading time) was determined using frontal analysis. Samples were collected in UV-readable collection plates, and the concentration of protein was determined spectrophotometrically using lambert- Beer’s law. Using a suitable plate layout, 200 µL of sample at a concentration of 3.5 mg/mL protein solution was added to each well and incubated for 60 minutes at 1,100 rpm. A prestudy was run to determine the effect of contact time on binding capacity. Plate Experiments: PreDictor Capto S plates filled with 2 µL of chromatography medium per well were used.

The factors investigated, associated ranges, and response (buffer was sodium acetate at ∼20 mM) The binding capacity for conalbumin on Capto S was investigated at various ionic strengths (20, 52, 85, 117, and 150 mM) and pH values (4.25, 4.75, and 5.25) using design of experiments (Table 1). Corresponding column experiments were run to verify that data correlated between formats. In this study, Predictor plates were used for screening of loading conditions for conalbumin (a protein showing nontraditional behavior) to the cation exchanger Capto™ S. PreDictor™ 96-well filter plates prefilled with GE Healthcare chromatography media allow parallel screening of conditions for binding, wash, or elution. The introduction of high-throughput techniques for process development partially addresses these challenges. This understanding/knowledge can be obtained if a larger experimental space is considered and/or more detailed studies are performed. In addition, the FDA’s new Quality by design initiative may increase demands on process development work because a higher degree of process understanding must be acquired. Finding optimal conditions for a downstream purification process is critical to achieving high productivity and a robust biopharmaceutical manufacturing process at large scale.

Capto s