Protein analysis and characterisation

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Using the FIDA technology you will be able to study ternary complex formation, where the cooperativity is one of the parameters extracted. On top of the cooperativity you will also gain information about the size of the complex and the affinity of the interactions.

FIDA is a fluorescence-based technique, hence for a binding affinity assay we need a protein with intrinsic fluorescence or a fluorophore on it. This is what we call the indicator and allows us to observe the interaction between an indicator and a binding partner. The binding partner is what we call the analyte. During a binding assay, the indicator is kept at a constant concentration and the analyte is titrated in at increasing concentrations. The analyte concentration should be high enough to reach a plateau on a binding curve to ensure full complex formation. This would allow you to extract the size of the indicator alone, the size of the indicator-analyte complex, and the binding affinity between indicator and analyte.

Yes, Fida Neo can be used to measure kinetics (association kon and dissociation koff rates). It does so in solution, and with no buffer, temperature, detergent or ionic strength constraints.

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Conceptually, the dissociation constant, or KD, is an equilibrium constant corresponding to the affinity between a ligand and a receptor (or between any two binding partners, such as an antibody and an antigen). Experimentally, the dissociation constant, or KD, is the concentration of analyte (ligand) at which half of the indicator (or binding sites on the receptor) is bound.