protein fluorescence ligandsProtein binding heavily modulates drug activity. Therefore, the binding features need to be elucidated when chemistry researchers study new molecules to be used as drugs. This short review concerns the analysis of the binding of small molecules to a fluorescent protein (the golden standard serum albumin) by using direct fluorescence titrations. This widely used technique is valuable, but often much more tricky than it might appear at first sight. We aim to share a mechanistic approach, discussing the critical aspects for correctly designing the experiments and treating the data.

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