These results indicate that DMA can be used as a small volume, high-throughput technique, relative to pressure-flow analysis, for the mechanical characterisation of chromatography media. The same trends were observed – The Capto family showed the highest resistance to deformation (Capto Adhere- 2.7, Capto Q- 1.92 1/%min-1) through to Sepharose CL-6B, Sepharose 4 Fast Flow and Sepharose CL-4B which exhibited the lowest slurry resistances (0.59, 0.4, 0.3 1/%min-1 respectively). The technique was applied to the nine studied resins and correlated with the results obtained using the pressure-flow technique. Dynamic mechanical analysis (DMA) was therefore developed as a novel technique in this field to address these limitations and allowed for further mechanical characterisation based on the viscoelastic properties using 1ml of resin. There were practical limitations in using the pressure-flow technique alone to for mechanical characterisation, including the large quantity of chromatography resin and buffers and the stringent criteria required to pack a column. The results showed that the Capto family had the highest critical velocities (Capto Adhere- 492, Capto Q- 477 cm/hr), whilst Sepharose CL-6B, Sepharose 4 Fast Flow and Sepharose CL-4B had the lowest critical velocity values (283, 204, 149 cm/hr respectively). Scanning electron microscopy (SEM) was used to image the structural properties of nine widely used agarose-based chromatography resins before use while pressure-flow analysis was used to characterise the mechanical properties of the same fresh resins. By understanding this, there is significant potential for facilitating timely and improved decisions in large-scale chromatographic operations, maximising resin lifetime whist maintaining acceptable column performance. Wash with Capto DEAE At least 10 CV start buffer or until eluent pH and conductivity have reached the required values.This thesis, completed in collaboration with Eli Lilly & Co., aims to understand and assess the structural and mechanical changes that occur as agarose-based chromatography resins are exposed to different bioprocessing conditions in an attempt to explore the mechanisms by which different resins age. Wash with at least 2 CV ultra pure water.ĥ. Wash with at least 2 column volumes (CV) of 2.0 M NaCl.Ĥ. Stable to commonly used aqueous buffers, 1.0 M NaOH 8 M Urea, 6 M guanidine hydrochloride, 30% isopropanol, and 70% ethanol.ġ. This prevents clogging and increases the life time of the column when loading large sample volumes. Filter the sample through a 0.45 µm filter or centrifuge at 10 000 × g for 10 min immediately before loading it to the column. Exchange buffer using a HiPrep 26/10 Desalting, HiTrap Desalting or PD-10 Desalting column.Ģ. Adjust the sample to the composition of the start buffer, using one of these methods: Dilute the sample with start buffer. These columns are easily connected in series to increase bed height to 20 cm.įast results and minimal consumption of sample and buffer consumption through the small bed volume.īioProcess columns packed with the same chromatography resin and using the same linear fluid velocity can produce scalable, reproducible results.ġ. Prepacked with Capto Q strong anion exchange chromatography resin.Ĭonvenient method optimization and parameter screening because of the 10cm bed height. HiScreen Capto Q is a column packed with a strong anion exchange modern resin, combining speed and capacity.
0 Comments
Leave a Reply. |
AuthorWrite something about yourself. No need to be fancy, just an overview. ArchivesCategories |