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Principle of Surface Plasmon Resonance

Surface plasmon resonance (SPR) is one of the commonly used technologies for detailed and quantitative studies of protein-protein interactions and determination of their equilibrium and kinetic parameters. SPR provides excellent instrumentation for a label-free, real-time investigation of protein-protein interactions.

Now I want to introduce the principle and protocol of SPR for you.

Principle SPR can be also used for thermodynamic analysis, epitope mapping, and to determine analyte concentration. The SPR-based binding method involves immobilization of a ligand on the surface of a sensor chip which has a monolayer of carboxymethylated dextran covalently attached to a gold surface. The ligand of interest is immobilized on the surface of the sensor chip using well-defined chemistry allowing solutions with different concentrations of an analyte to flow over it and to characterize its interactions to the immobilized ligand (Fig. 1). The SPR signal originates from changes in the refractive index at the surface of the gold sensor chip. The increase in mass associated with a binding event causes a proportional increase in the refractive index, which is observed as a change in response. These changes are measured as changes in the resonance angle (δθ) of refracted light when the analyte, flowing in a microfluidic channel, binds to the immobilized ligand and increases in density at the sensor chip. Importantly, for protein-protein interactions the change in refractive index on the surface is linearly related to the number of molecules bound. The response signal is quantified in resonance units (RU) and represents a shift in the resonance angle, where 1RU is equal to a critical angle shift of 10-4 deg or 10-12 gmm-2. When a steady-state is achieved (all binding sites occupied), the maximum RU is determined (n: number of binding sites in ligand).

Source: http://www.creativebiomart.net/resource/principle-protocol-principle-and-protocol-of-surface-plasmon-resonance-spr-361.htm

Note by Caroline Green
4 months, 2 weeks ago

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