Table 7.
5-mL validation runs of the fully optimised process for both FabscFv-KiH and Fab2scFv-KiH
| Load | Monomer concentration (mg/mL) | Step monomer recovery (%) | Overall monomer recovery (%) | Leached Protein A (ppm) | HCP (ppm) | hcDNA (ppm) | Purity (%) | ||||
|---|---|---|---|---|---|---|---|---|---|---|---|
| HMW | Mono | LMW | |||||||||
| CCS | – | 0.56 | - | - | N.D | 1,307,373 | 67,006.3 | 30.8 | 34.0 | 35.2 | |
| FabscFv-KiH |
Post-Protein A eluate (pH 4.7 wash) |
31.5 mg/mL-R | 3.68 | 91.1 | 91.1 | 5 | 2350 | 0.5 | 4.8 | 94.7 | 0.5 |
|
Polishing FT mode: Capto Butyl ImpRes, pH 4.0 |
40 mg/mL-R | 2.25 | 92.7 | 84.4 | N.D | 189 | 0.5 | 1.8 | 98.1 | 0.1 | |
|
Polishing FT mode: Capto adhere, pH 6.8 |
30 mg/mL-R | 0.76 | 88.7 | 74.9 | N.D | 87 | 0.2 | 0.6 | 99.3 | 0.1 | |
| CCS | – | 0.72 | - | - | N.D | 1,077,136 | 54,133.9 | 33.5 | 31.1 | 35.4 | |
| Fab2scFv-KiH |
Post-Protein A eluate (pH 4.1 wash) |
30.5 mg/mL-R | 1.75 | 78.4 | 78.4 | 6 | 2064 | 0.5 | 5.1 | 93.2 | 1.7 |
|
Polishing FT mode: Capto Butyl ImpRes, pH 4.0 |
60 mg/mL-R | 1.52 | 93.0 | 72.9 | N.D | 153 | 0.3 | 2.6 | 96.6 | 0.8 | |
|
Polishing FT mode: Capto adhere, pH 6.8 |
45 mg/mL-R | 0.73 | 82.1 | 59.9 | N.D | 59 | 0.1 | 0.3 | 99.2 | 0.5 | |
Representative post-Protein A eluates are reported in the table, as multiple post-Protein A eluates from 5-mL column are pooled together to generate sufficient materials for the subsequent polishing steps