Download use case: Global Fitting
The Indian parable of “The six blind men and the elephant” tells the story of six blind men who touch an elephant in the hope of learning what it is like. As each one can only feel a different part of the animal the individual conclusions obtained are in disagreement and none of them provides a real view of the full elephant. “only by sharing what each of you knows can you possibly reach a true understanding”; that´s the moral behind this nice story.
Fig 1: The six blind men and the elephant: only a global analysis of the overall data provides a true understanding.
The binding assay(s) achieved to characterize a molecular interaction often provides not just one, but several binding curves from which the affinity constant is obtained.
Sometimes, an individual fit of these curves yield a set of binding constants that are significantly different from them; this result can be very confusing because, in principle, these binding curves are a representation of the same binding event and should converge to provide the analogous information. Often, the explanation for this behaviour is that the different curves indeed provide only partial and/or different information of the interaction, not enough to unequivocally determine the binding affinity through individual analysis.
“It´s like feeling only a separate part of the elephant”
This is a typical scenario when facing the study of complex binding events that involve more than one equilibrium and several binding curves are obtained, i.e. from different frequencies of the spectra in a titration experiment, from data registered using different techniques (ITC, NMR, Optical Spectroscopies…) and/or from experiments performed at different concentrations of the species participating in the binding event.
Analogous to the parable of the six men and the elephant, the way to get a true understanding of the binding event consist of the global analysis of the different curves.
Fig 2: The binding curve obtained from 2D NMR titrations.
Being aware of the relevance of global analysis, in AFFINImeter we count with the possibility to perform Global fitting of multiple data to tailored binding models where one or more fitting parameters are shared between isotherms. The number and identity of the parameters shared are selected by the user.
Moreover, two or more parameters can be related through mathematical relationships designed by the user. All these features make our global fitting tool the most potent among others to perform a robust analysis of binding data of complex interactions.