48 February / March 2021
on different particle technologies and materials produce the fl exibility required for most BioLC separations and depending on the separation mode employed the small particle sizes available (between 1.9 and 3 µm) allow high resolutions and speedy run times for each mode.
1. M. Kesik-Brodacka, Progress in biopharmaceutical development, Biotechnol. Appl. Biochem., 65(3) (2018) 306-322.
2. Evaluate Pharma, World Preview 2019, Outlook to 2024, 12 (2019).
3. I. Novák, B. Buszewski, J. Garaj, D. Berek, Infl uence of pore structure of silica packing on HPLC column characteristics, Chem Papers, 44 (1) (1190), 31-43.
4. A. Goyon, S. Fekete, A. Beck, J.L. Veuthey, D. Guillarme, Unravelling the mysteries of modern size exclusion chromatography – the way to achieve confi dent characterization of therapeutic proteins, J. Chrom. B, 1092 (2018) 368–378.
Figure 5: Separation of MAbs using non-porous BioPro IEX SF column and a weak exchanger column often used for such analysis. The strong exchange column can achieve higher resolution in shorter analysis time. Eluent: A) 20 mM MES-NaOH (pH 5.6) B) 20 mM MES-NaOH (pH 5.6) containing 0.2 M NaCl; gradient: 35 %B (70 mM NaCl); Gradient slope: 0.25 %B/min (0.5 mM NaCl); Flow rate: 180 cm/hr; temperature: 30 °C; detection: UV at 280 nm; injection: 10 µL (1 mg/mL IgG1).
5. Proteins Under Pressure, Chromatography Today, 12(4) (2020) 44-46.
5 µm non-porous BioPro IEX QF/SF columns offer higher performance when compared to other IEX columns (Figure 5). Due to the low backpressures, higher fl ow rates can often be applied while maintaining the desired resolution. This fact together with the particle robustness and reproducibility makes the 5 µm non-porous exchangers an ideal choice for QC.
Conclusion
The use of smaller, modern particles usually allows shorter analysis times and therefore higher sample throughput for all separation modes typically used in BioLC. With the increasing number of therapeutic biologicals, this is welcomed by R&D as well as QC labs. Modern solutions based
6. V. D’Atri, R. Pell, A. Clarke, D. Guillarme, S. Fekete, Is hydrophobic interaction chromatography the most suitable technique to characterize site-specifi c antibody-drug conjugates?, J. Chrom. A, 1586 (2019) 149–153.
7. A. Murisier, M. Lauber, S. J. Shiner, D. Guillarme, S. Fekete, Practical considerations on the particle size and permeability of ion-exchange columns applied to biopharmaceutical separations, J. Chrom. A, 1604 (2019) 460-487.
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