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eSPC, an Online Data AnalysisPlatform for MolecularBiophysicsThermoAffinity 1.0 UserDocumentationOctober 2022 Table of Contents1. Load input1.1. Input file (raw data)1.2. Normalization1.3. Median filter (smoothing)1.4. Hot and cold region selection2. Fitting2.1. Model2.2.Initial estimates and boundaries of the parameters2.3. Curve fitting2.4. Fitting errors OverviewThermoAffinityhassevenpanels(Figure1).Panels1-2containthenecessarystepstoanalyzeuserdata.TheSimulatedataPanelcanbeusedbeforedoinganexperimenttoanalyzetheexpectedchangeinthesignaldependingonthebindingaffinity and the protein and ligand concentrations.Figure 1.ThermoAffinity online tool panels.1. Load input1.1. Input file (raw data)ThermoAffinityacceptsasinputthespreadsheetgeneratedbyNanoTemperTechnologies.Thisfilecontainsonesheetcalled‘RawData’wherethefirstcolumnhasdifferentcellswiththefollowinglabels:‘CapillaryPosition:’,‘Ligand:’,‘LigandConcentration:’,and‘Time[s]’.Then,thesecondcolumnstorestheassociatedinformation:‘1’,‘lig...