title: Optimizing isothermal titration calorimetry protocols for the study of 1:1 binding: Keeping it simple.
abstract: BACKGROUND
  Successful ITC experiments require conversion of cell reagent (titrand M) to product and production or consumption of heat. These conditions are quantified for 1:1 binding, M+X ⇔ MX.
  
  
  METHODS
  Nonlinear least squares is used in error-propagation mode to predict the precisions with which the key quantities - binding constant K, reaction enthalpy ΔH°, and stoichiometry number n - can be estimated over a wide range of the dimensionless quantity that governs isotherm shape, c=K[M]0. The measurement precision σq is estimated from analysis of water-water blanks.
  
  
  RESULTS
  When the product conversion exceeds 90%, the parameter relative standard errors are proportional to σq/qtot, where the total heat qtot ≈ ΔH° [M]0V0. Specifically, σK/K×qtot/σq ≈ 25 for c=10(-3)-10, ≈ 11 c(1/3) for c=10-10(4). For c>1, n and ΔH° are more precise than K; this holds also at smaller c for the product n×ΔH° and for ΔH° when n can be held fixed. Use of as few as 10 titrant injections can outperform the customary 20-40 while also improving productivity.
  
  
  CONCLUSIONS
  These principles are illustrated in experiment design using the program ITC-PLANNER15.
  
  
  CONCLUSIONS
  Simple quantitative guidelines replace the "c rules" that have dominated the literature for decades.
authors.affiliation: Department of Chemistry, Vanderbilt University,Nashville,Tennessee 37235. Electronic address: joel.tellinghuisen@vanderbilt.edu.
authors.forename: Joel
authors.initials: J
authors.lastname: Tellinghuisen
mesh_terms: D017609:Barium Compounds / Q000737:chemistry*; D002151:Calorimetry / Q000592:standards*; D002712:Chlorides / Q000737:chemistry*; D043844:Crown Ethers / Q000737:chemistry*; D006358:Hot Temperature; D007700:Kinetics; D016018:Least-Squares Analysis; D009566:Nitrates / Q000737:chemistry; D012107:Research Design; D012984:Software; D013696:Temperature; D013816:Thermodynamics
chemical_list: D017609:Barium Compounds; D002712:Chlorides; D043844:Crown Ethers; D009566:Nitrates; C024986:barium chloride; C015762:18-crown-6; C023545:barium nitrate
keywords: Data analysis; Experiment design; ITC; Nonlinear least squares; Statistical errors