Calibrating ITC instruments: Problems with weak base neutralization.

Modern isothermal titration calorimetry instruments give great precision, but for comparable accuracy they require chemical calibration. For the heat factor, one recommended process is HCl into the weak base TRIS. In studying this reaction with a VP-ITC and two Nano-ITCs, we have encountered some problems, most importantly a titrant volume shortfall Δv ≈ 0.

Goodness-of-Fit Tests in Calibration: Are They Any Good for Selecting Least-Squares Weighting Formulas?

No. Most goodness-of-fit (GOF) tests attempt to discern a preferred weighting using either absolute or relative errors in the back-calculated calibration values. However, the former are predisposed to select constant weighting and the latter 1/ or 1/ weighting, no matter what the true weighting should be.

A (partial) resolution of binding enthalpy discrepancies in ITC studies of Ba crown ether complexation: The importance of calibration.

By conducting binding experiments at a range of temperatures T using isothermal titration calorimetry (ITC), one can obtain two estimates of the binding enthalpy - calorimetric (ΔH°) from the experiments at each T, and van't Hoff (ΔH°) from the T dependence of the binding constant K°. From thermodynamics it is clear that these two must be identical, but early efforts to demonstrate this for ITC data indicated significant inconsistency. In an extensive 2004 study of the Ba + 18-crown-6 ether complexation used in prior comparisons, Mizoue and Tellinghuisen found modest (10-20%) but statistically significant differences, which were tentatively attributed to problems converting the calorimetric estimates to their standard state values, as implied by the superscript ° in the notation.

Estimating Real-Time qPCR Amplification Efficiency from Single-Reaction Data.

Methods for estimating the qPCR amplification efficiency from data for single reactions are tested on six multireplicate datasets, with emphasis on their performance as a function of the range of cycles - included in the analysis. The two-parameter exponential growth (EG) model that has been relied upon almost exclusively does not allow for the decline of () with increasing cycle number through the growth region and accordingly gives low-biased estimates. Further, the standard procedure of "baselining"-separately estimating and subtracting a baseline before analysis-leads to reduced precision.

dPCR vs. qPCR: The role of Poisson statistics at low concentrations.

The ultimate precision in both dPCR and qPCR experiments is limited by the Poisson statistics in the total number m of template molecules in the sample, giving relative standard deviation 1/m. This means that precision is limited by sample volume at low concentrations. Accordingly qPCR instruments, used in dPCR mode, can give better precision than dPCR instruments in this limit.