Reversed-Phase HPLC Chromatography Services
Biopharmaceutical and related industries rely on the team at GL Technologies for their HPLC calibration and certification requirements. We have worked hard to earn the trust of our clients throughout the entire State of California with equipment calibration services and provide a full-service HPLC team of experts to help our clients with their laboratory requirements.
Reversed-phase chromatography (RPC) is a powerful technique used in the biopharmaceutical industry to purify proteins and other biological molecules. This technique exploits the differences in hydrophobicity between molecules to separate them based on their affinity for a hydrophobic stationary phase.
To speak with the experts about your Reversed Phase HPLC Calibration needs, please fill out our online form or give us a call!
The Stationary Phase
The stationary phase in RPC is typically a hydrophobic resin, such as C18 or C8, that is packed into a column. The sample is applied to the column and then mixed with a mobile phase that contains a gradient of an organic solvent, such as acetonitrile or methanol, and an aqueous buffer. As the concentration of the organic solvent increases, the hydrophobic molecules are gradually displaced from the stationary phase and removed from the column. The less hydrophobic molecules removed, they require a higher concentration of organic solvent to be displaced.
RPC is particularly useful for purifying proteins that have been expressed in a heterologous system, such as E. coli or yeast, because these proteins often contain hydrophobic domains that can interact with the stationary phase. In addition, RPC can be used in combination with other chromatographic techniques, such as ion exchange chromatography or size exclusion chromatography, to achieve higher levels of purification.
Reversed Phase Chromatography Advantages
One of the key advantages of RPC is its ability to separate closely related molecules, such as protein isoforms or variants, that have similar charge or size properties but differ in hydrophobicity. By adjusting the gradient of the mobile phase, it is possible to selectively elute specific isoforms or variants, allowing for highly specific purification.
Despite its many advantages, RPC does have some limitations. For example, the hydrophobic stationary phase can interact non-specifically with other molecules in the sample, leading to decreased selectivity and yield. In addition, the use of organic solvents can denature some proteins, leading to loss of activity or conformational changes.
Reversed phase chromatography is a very effective technique for purifying proteins and other biological molecules in the biopharmaceutical industry. Its ability to separate closely related molecules and achieve high levels of purification make it an important tool for research and development.
To speak with the experts about your Reversed Phase HPLC Calibration needs, please fill out our online form or give us a call!
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