18 research outputs found
Towards Protein Crystallization as a Process Step in Downstream Processing of Therapeutic Antibodies: Screening and Optimization at Microbatch Scale
Crystallization conditions of an intact monoclonal IgG4 (immunoglobulin G, subclass 4) antibody were established in vapor diffusion mode by sparse matrix screening and subsequent optimization. The procedure was transferred to microbatch conditions and a phase diagram was built showing surprisingly low solubility of the antibody at equilibrium. With up-scaling to process scale in mind, purification efficiency of the crystallization step was investigated. Added model protein contaminants were excluded from the crystals to more than 95%. No measurable loss of Fc-binding activity was observed in the crystallized and redissolved antibody. Conditions could be adapted to crystallize the antibody directly from concentrated and diafiltrated cell culture supernatant, showing purification efficiency similar to that of Protein A chromatography. We conclude that crystallization has the potential to be included in downstream processing as a low-cost purification or formulation step
Rapid SDS-GEL capillary electrophoresis for the analysis of recombinant NADP + -dependent formate dehydrogenase during expression in Escherichia coli cells and its purification
Abstract The level of expression in Escherichia coli cells and different steps of purification of the recombinant NADP + -dependent formate dehydrogenase (EC 1.2.1.2, FDH) from bacterium Pseudomonas sp.101 was analyzed by rapid SDS-Gel capillary electrophoresis (SDS-Gel CE) and compared with SDS polyacrylamide gel electrophoresis (SDS PAGE). First standard proteins were separated in the short capillary and the calibration curve generated, then fractions taken during the fermentation and purification process were analyzed. The main advantages of SDS-Gel CE are short analysis time, high sensitivity, the possibility to quantify proteins at different ultraviolet wavelength, and small injection volumes. The data for each step of the fermentation process and during the purification were controlled by spectrophotometric analysis of enzyme activity and protein concentration as well as standard SDS PAGE. The molecular mass of the purified FDH was determined as 44 078 Da by matrix-assisted laser desorption/ ionisation time of flight mass spectrometry Β© 1997 Elsevier Science B.V
Improved high-performance liquid chromatographic method in the analysis of adenovirus particles
NRC publication: Ye
Continuous Bed HPLC in the detection and quantification of adenoviruses
NRC publication: Ye
Improved high-performance liquid chromatographic method in the analysis of adenovirus particles
NRC publication: Ye
Purification of a Fab'Fragment by expanded bed absorption chromatography
NRC publication: Ye
Metod farmakologicheskoy korrektsii metabolizma kostnoy tkani dlya uluchsheniya rezul'tatov endoprotezirovaniya tazobedrennogo sustava
ΠΠΎΡΠΎΠ±ΠΈΠ΅ Π΄Π»Ρ Π²ΡΠ°ΡΠ΅ΠΉ ΠΏΠΎΡΠ²ΡΡΠ΅Π½ΠΎ ΡΠ»ΡΡΡΠ΅Π½ΠΈΡ ΡΠ΅Π·ΡΠ»ΡΡΠ°ΡΠΎΠ² ΡΠ½Π΄ΠΎΠΏΡΠΎΡΠ΅Π·ΠΈΡΠΎΠ²Π°Π½ΠΈΡ ΠΏΠ°ΡΠΈΠ΅Π½ΡΠΎΠ², ΡΡΡΠ°Π΄Π°ΡΡΠΈΡ
ΠΎΡΡΠ΅ΠΎΠΏΠΎΡΠΎΠ·ΠΎΠΌ ΠΈΠ»ΠΈ Π΄ΡΡΠ³ΠΈΠΌΠΈ Π·Π°Π±ΠΎΠ»Π΅Π²Π°Π½ΠΈΡΠΌΠΈ, ΠΏΡΠΈ ΠΊΠΎΡΠΎΡΡΡ
Π½Π°ΡΡΡΠ°Π΅ΡΡΡ ΡΠ΅ΠΌΠΎΠ΄Π΅Π»ΠΈΡΠΎΠ²Π°Π½ΠΈΠ΅ ΠΊΠΎΡΡΠ½ΠΎΠΉ ΡΠΊΠ°Π½ΠΈ (Π±ΠΎΠ»Π΅Π·Π½Ρ ΠΠ΅Π΄ΠΆΠ΅ΡΠ°, Π³ΠΈΠΏΠ΅ΡΠΏΠ°ΡΠ°ΡΠΈΡΠ΅ΠΎΠΈΠ΄Π½Π°Ρ ΠΎΡΡΠ΅ΠΎΠ΄ΠΈΡΡΡΠΎΡΠΈΡ ΠΏΠΎΡΠ»Π΅ ΡΠ΄Π°Π»Π΅Π½ΠΈΡ Π°Π΄Π΅Π½ΠΎΠΌΡ ΠΎΠΊΠΎΠ»ΠΎΡΠΈΡΠΎΠ²ΠΈΠ΄Π½ΠΎΠΉ ΠΆΠ΅Π»Π΅Π·Ρ, ΠΎΡΡΠ΅ΠΎΠΌΠ°Π»ΡΡΠΈΡ). Π Π°Π·ΡΠ°Π±ΠΎΡΠ°Π½Π½ΡΠΉ ΡΠΏΠΎΡΠΎΠ± ΡΠ°ΡΠΌΠ°ΠΊΠΎΠ»ΠΎΠ³ΠΈΡΠ΅ΡΠΊΠΎΠΉ ΠΊΠΎΡΡΠ΅ΠΊΡΠΈΠΈ Π½Π°ΡΡΡΠ΅Π½ΠΈΠΉ ΡΠ΅ΠΌΠΎΠ΄Π΅Π»ΠΈΡΠΎΠ²Π°Π½ΠΈΡ ΠΊΠΎΡΡΠ½ΠΎΠΉ ΡΠΊΠ°Π½ΠΈ Π·Π°ΡΠΈΡΠ΅Π½ ΠΏΠ°ΡΠ΅Π½ΡΠΎΠΌ (ΠΏΠ°ΡΠ΅Π½Ρ Π Π€ Π½Π° ΠΈΠ·ΠΎΠ±ΡΠ΅ΡΠ΅Π½ΠΈΠ΅ β 2176519 ΠΎΡ 10.12.01) ΠΈ ΠΎΡΠ½ΠΎΠ²Π°Π½ Π½Π° ΠΊΠΎΠΌΠ±ΠΈΠ½ΠΈΡΠΎΠ²Π°Π½Π½ΠΎΠΉ Π»Π΅ΠΊΠ°ΡΡΡΠ²Π΅Π½Π½ΠΎΠΉ ΡΠ΅ΡΠ°ΠΏΠΈΠΈ Ρ ΠΈΡΠΏΠΎΠ»ΡΠ·ΠΎΠ²Π°Π½ΠΈΠ΅ΠΌ ΠΊΠ°Π»ΡΡΠΈΡΠΎΠ½ΠΈΠ½Π° (ΠΌΠΈΠ°ΠΊΠ°Π»ΡΡΠΈΠΊΠ° ΠΏΡΠΈΠΌΠ΅Π½ΡΠ΅ΠΌΠΎΠ³ΠΎ ΠΈΠ½ΡΡΠ°Π½Π°Π·Π°Π»ΡΠ½ΠΎ ΠΈΠ»ΠΈ Π² Π²ΠΈΠ΄Π΅ Π²Π½ΡΡΡΠΈΠΌΡΡΠ΅ΡΠ½ΡΡ
ΠΈΠ½ΡΠ΅ΠΊΡΠΈΠΉ), Π°Π»ΡΡΠ°ΠΊΠ°Π»ΡΡΠΈΠ΄ΠΎΠ»Π° (Π°Π»ΡΡΠ°-D3 Π’ΠΠΠ, ΠΡΠ°Π»ΡΡΠ° ΠΎΠΊΡΠΈΠ΄Π΅Π²ΠΈΡΠ°) ΠΈ ΠΏΡΠ΅ΠΏΠ°ΡΠ°ΡΠΎΠ² ΠΊΠ°Π»ΡΡΠΈΡ.
ΠΡΠΈΠ²Π΅Π΄Π΅Π½Ρ ΡΠ°ΠΊΡΠΎΡΡ ΡΠΈΡΠΊΠ° Π°ΡΠ΅ΠΏΡΠΈΡΠ΅ΡΠΊΠΎΠΉ Π½Π΅ΡΡΠ°Π±ΠΈΠ»ΡΠ½ΠΎΡΡΠΈ ΡΠ½Π΄ΠΎΠΏΡΠΎΡΠ΅Π·ΠΎΠ² ΠΈ ΡΡ
Π΅ΠΌΡ ΠΏΡΠΎΡΠΈΠ»Π°ΠΊΡΠΈΠΊΠΈ ΠΈ Π»Π΅ΡΠ΅Π½ΠΈΡ Π² Π·Π°Π²ΠΈΡΠΈΠΌΠΎΡΡΠΈ ΠΎΡ ΡΡΠ΅ΠΏΠ΅Π½ΠΈ ΡΠΈΡΠΊΠ° Π΅Π΅ ΡΠ°Π·Π²ΠΈΡΠΈΡ. ΠΠ΅ΡΠΎΠ΄ ΠΏΡΠ΅Π΄Π½Π°Π·Π½Π°ΡΠ΅Π½ Π΄Π»Ρ ΡΠ»ΡΡΡΠ΅Π½ΠΈΡ ΡΠ΅Π·ΡΠ»ΡΡΠ°ΡΠΎΠ² ΡΠ½Π΄ΠΎΠΏΡΠΎΡΠ΅Π·ΠΈΡΠΎΠ²Π°Π½ΠΈΡ ΠΈ ΡΠ΅ΡΠ½Π΄ΠΎΠΏΡΠΎΡΠ΅Π·ΠΈΡΠΎΠ²Π°Π½ΠΈΡ ΡΠ°Π·ΠΎΠ±Π΅Π΄ΡΠ΅Π½Π½ΠΎΠ³ΠΎ ΡΡΡΡΠ°Π²Π°
Anything but Conventional Chromatography Approaches in Bioseparation
While packed bed chromatography, known as conventional chromatography, has been serving the biopharmaceutical industry for decades as the bioseparation method of choice, alternative approaches are likely to take an increasing leading role in the next few years. The high number of new biological drugs under development, and the need to make biopharmaceuticals widely accessible, has been driving the academia and industry in the quest of anything but conventional chromatography approaches. In this perspective paper, these alternative approaches are discussed in view of current and future challenges in the downstream processing field