Cheat sheet: Static and Kinetic SPR

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Cheat Sheet_Kin vs stat_V5 final.pdf

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Main Differences

Static SPR

Purpose: Scouting or screening prior to Kinetic SPR, yes/no binding

• Like label-free ELISA: ligand-analyte interaction depends on diffusion, concentration, and binding strength is measured as Apparent K_D

• Concentration determination of active analyte in complex media (e.g. crude lysate)

• Quick verification of protein expression

• Contact time ~ seconds to hours so slow interactions can be measured with Static SPR

• Low viscosity buffers and biomolecules 10-500 kDa are recommended²

• Up to 4 independent experiments (4 channels) for P4 SPR device¹

Flow SPR

Purpose: Sensitive, quantitative binding characterization

• Constant flow of analyte enables ligand-analyte interaction so both strength and rate of binding are measured as Quantitative K_D

• Can transfer Static SPR conditions to Kinetic SPR

• Due to controlled flow, Kinetic SPR can accommodate a broader range of molecule sizes (100+ Da)

• Suitable for low analyte concentrations, better real-time sensitivity

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Data Output Differences

Static SPR

• Association without dissociation

• End-point response units are analyzed

• Like ELISA, Apparent K_D can be estimated at ½ RU_max determined by concentration titration

Flow SPR

• Both association and dissociation

• Quantitative K_D calculated by k_off/k_on

• Steeper upward curve à faster association

• Steeper downward curve à faster dissociation

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