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Amine coupling

Chemistry

Amine coupling chemistry is the most widely applicable approach for attaching biomolecules covalently to the sensor surface. With this method, carboxyl groups on the surface of the sensor chip surface are first activated with a mixture of 1-ethyl-3-(3 dimethylaminopropyl) carbodiimide (EDC) and N-hydroxysuccinimide (NHS) to give reactive succinimide esters. Ligand is then passed over the surface and the esters react spontaneously with primary amine groups or other nucleophilic groups to link the ligand covalently to the dextran matrix. Excess reactive groups are deactivated with ethanolamine. This approach is generally useful for immobilization of proteins.

Most proteins contain several amine groups so that efficient attachment can be achieved without seriously affecting the biological activity of the ligand. In some instances, however, amine coupling may involve groups at or near the active site or binding site of the ligand, with the result that attachment is accompanied by loss of activity. In such cases, the ligand can be attached using alternative coupling chemistry or a capturing approach.

Amine coupling chemistry

Immobilization procedure overview

The general pattern of steps involved in immobilizing ligand is the same for essentially all covalent immobilization methods:

• Activate the surface by injection of appropriate reagents.
• Inject the ligand solution.
• Inject reagent to deactivate remaining active groups on the surface and remove non-covalently bound ligand.

There may be additional steps depending on the properties of the ligand, the type of sensor surface and the details of the chemistry used.

Lab protocol

Lab procedure Amine coupling of ligand to Biacore sensor chips is available for download on www.cytivalifesciences.com/solutions/protein-research/Interaction-analysis-with-Biacore-surface-plasmon-resonance-SPR/Get-started-with-surface-plasmon-resonance-SPR-interaction-analysis

See also

List of help files