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Agile BioScience: High-Performance Analysis with Liquid Chromatography
Liquid Chromatography
Liquid chromatographic profiling is the use of chromatographic techniques to separate molecules by different physicochemical properties. It forms a key part of any protein, glycoprotein or peptide structural characterization investigation and it is a requirement of ICH Topic Q6B.
Chromatographic Techniques at Agile Bioscience
Using a wide variety of chromatographic techniques, Agile Bioscience has vast experience separating compounds based on their distinct physicochemical features. We perform this for our clients as a purity and impurity assessment of their product as well as to help them meet the ICH Q6B criteria for structural characterization. separate chromatographic separation techniques are employed to fully evaluate a sample's purity, with the expectation that any components not resolved by one method will likely
Reverse phase chromatography
This method uses the interaction of a protein, glycoprotein, or peptide with a stationary phase coated with hydrocarbon to separate components. To encourage the elution of every component in the sample, a liquid gradient with an increasing organic composition is employed. Depending on the components being studied, different hydrocarbon chain lengths are selected on the stationary phase.
Ion exchange chromatography
This technique separates molecules based on their charge. Proteins, glycoproteins and peptides can carry either a net negative or net positive charge, depending on the amino acid sequence, post-translational modifications and glycosylation. Separation of charged species is brought about through the use of either salt or pH gradients in the liquid phase.
Ion exchange chromatography is complementary to imaging capillary isoelectric focusing (icIEF) which is used for charge isoform analysis.
Size exclusion chromatography (SEC)
Molecules are separated according to size during SEC as they go through a gel column made of beads with a specific pore size. To put it simply, because larger molecules are excluded from or have limited access to pore space than smaller molecules, they will travel through the matrix more quickly. Smaller molecules have a longer pathlength in the column because they can pass through the pores and around the beads.
Hydrophobic interaction chromatography (HIC)
Hydrophobicity and hydrophilicity are the criteria used to separate molecules during HIC. The protein's exposed hydrophobic domains associate with the stationary phase, which functions as a high salt buffer, to accomplish this. Components are eluted using a decreasing salt gradient; more hydrophobic species elute at lower salt concentrations.
