Hydrophobic interaction chromatography (HIC) is a must for nucleic acid separations. When combined with the advantages of monolithic chromatography, HIC meets this need while also providing an excellent solution for the purification of large biomolecules including adenoassociated viruses (AAV).
HIC separates and purifies protein molecules on the basis of their hydrophobicity – it is more popular than other chromatography techniques for the separation of proteins as it employs a less denaturing environment compared to them. Therefore the biological activity of the proteins is kept intact.
HIC can also be effectively used to remove impurities or product aggregate species in aqueous solutions as it exploits the difference in hydrophobic properties of the aggregates and the target molecules. It is often used in combination with techniques such as ion exchange or gel filtration chromatography.
When analyzing, you need to pay attention to several points: the pH value of the solution, the effect of different pH values on protein purification will also be different. If you want the protein to be eluted smoothly, the pH value can be far away from its isoelectric point, the hydrophobicity is reduced, and the protein is not easy to bind to the stationary phase ligand, which is conducive to the elution of the protein. If you want to improve the resolution of a chromatography column, you can start by increasing the column temperature. The increase in column temperature can increase the hydrophobic effect and the adsorption capacity, thereby improving the resolution.
Whether the separation effect of the chromatographic column is good or not is not only the influence of these factors, but also the usual maintenance and use methods. After using for a period of time, you can rinse the column to wash away impurities to prevent impurities from affecting the analysis results, such as the appearance of impurity peaks. A chromatographic protection column can be added in front of the column, which can retain some strong residues, strong acidic and strong alkaline substances in the sample to prevent them from contaminating and destroying the packing in the chromatographic column. UHPLCS various protection column and HPLC guard cartridge suitable for different HPLC column.
2.1 HPLC analytical guard column
• Guard columns remove particulate contaminants and highly adsorptive compounds from samples
• Modular and cartridge replacement is easy and tool free
• Holders are available in stainless steel and PEEK materials, both designs optimized to reduce dead volume and prevent extra column band broadening
• Holders for the cartridges are reusable indefinitely
Hydrophobic interaction chromatography has many advantages, of course, there are also disadvantages. For example, some proteins have reduced solubility in high salt, so its application capacity will be limited to a certain extent, and it is more suitable for the purification and separation of hydrophobic proteins.
Post time: Aug-21-2021
