Intra-assay total uncertainty of results in immunoassay techniques

Uncertainty is a parameter associated with the result of a measurement; this parameter characterizes the dispersion of the values that could reasonably be attributed to the sample. Data processing methods do not take into account the influence of the imprecision and deviation of the experimental points of the calibration system and their impact on the final result of a sample analysis. The aim of this work is: (a) to propose, for each run, a simple method to calculate the uncertainty due to the calibration system (Uc); and (b) to present a method to determine the "intra-assay total uncertainty" (Ut) and evaluate its impact on the final result for an analyte. Ten replicates of standards, controls, and two serum-male and female samples were measured in the same run with a manual kit for determination of testosterone. To calculate Ut, random duplicate responses were selected. For controls and samples, Ut was affected by Uc (2.91% to 6.59%) and by the uncertainty of the measurement of the sample (Us) (1.01 to 8.73%); this allowed us to determine that Ut had values from 3.73% to 9.87%. While Us affects the result of a given sample, Uc affects the result of all the samples with a similar response (cpm). In the method proposed, Ut involves Us and Uc, both factors that introduce variations into the result of a sample by random causes. Intraassay total uncertainty includes the most probable result for the analytical methodology selected.