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What is ion exchange chromatography?

Ion exchange (IEX) chromatography is a technique that separates biomolecules from a mixture on the basis of net charge. This chromatographic separation method uses the interaction between charged molecules and oppositely charged moieties in the stationary phases of chromatography matrices. Since the charge that molecules carry is easy to manipulate by altering buffer pH, ion exchange column chromatography allows the separation of similar types of molecules.

The two main types of ion exchange separation are cation and anion exchange. The stationary phase is positively charged in anion exchange and negatively charged in cation exchange.

Steps for ion exchange chromatography in protein purification:

  1. Equilibration. The first step in IEX chromatography is the equilibration of the stationary phase. pH and ionic strength of start buffers facilitate binding of the target protein to resin.
  2. Sample application and wash This step involves binding of target molecules and washing of unbound materials.
  3. Elution. Target molecules are eluted based on buffer ph. Regeneration. The last step is a wash using a buffer with high ionic strength. Be sure to re-equilibrate ion exchange columns and start buffers before the next run.

What are the benefits of prepacked columns?

Each column goes through strict quality control to ensure low variation from batch to batch, and columns are validated for use. Other advantages include:

  • They save column-packing time.
  • They facilitate speedy chromatography process set-up times.
  • They’re packed to the same specifications and rigorously tested to support reproducible results.

Ion exchange columns FAQs

Here are answers to some frequently asked questions about IEX columns.

What is an ion exchange column?

An IEX column is a chromatography column packed with an IEX resin that enables the separation of compounds based on their net charge.

How does an ion exchange column work?

Proteins can carry positive, negative, or no charge, based on the pH of their environment. The pH at which a molecule carries no net charge is its isoelectric point (pI). Cation exchange columns carry a negative charge and capture positively charged molecules, while anionic exchange columns carry a positive charge and capture negatively charged biomolecules.

How do I choose an ion exchange column?

Choosing an ion exchange column depends on various factors that affect chromatographic separation. Consider:

  • The charge properties of the target and other molecules in the sample
  • Sample volume and concentration
  • Flow rate of the chromatography system
  • Pressure limits of the chromatographic system

Our Purify app can help you choose the right column

Which column should be used in ion exchange chromatography?

We offer a vast range of ion exchange columns for specialized applications like high-resolution separations and preparative separations. Visit our ion exchange resin and column selection guide and visit our ion exchange chromatography solutions page for more resources and information.