VARIABILITY OF MOLECULAR BIOMARKER MEASUREMENTS FROM NONLINEAR CALIBRATION CURVES

In any immunoassay experiment for the detection of molecular biomarker s, a nonlinear calibration curve is constructed to relate fixed biomar ker concentrations to observed tracer levels. The biomarker concentrat ion in an experimental sample can then be estimated by projecting the experimental tracer measurements through the inverse of the calibratio n curve. Once an estimate of the biomarker level has been calculated, it is often of interest to determine its variability. Typically, metho ds for estimating this variability assume that the biomarker variabili ty is due solely to the uncertainty in the estimation of the calibrati on curve. A more complete analysis would combine this uncertainty with the variability in the processing and measurement of the sample, incl uding, e.g., measurement error of laboratory procedures or variation i n enzymatic activity in enzyme-linked immunosorbent assays or radioact ivity counts in RIAs. In this paper, we present a method of estimating the variability of inverse estimates assuming there is variation aris ing from both the determination of the calibration curve and from the preparation and measurement of the experimental sample. Our method use s a resampling algorithm that avoids requiring many distributional ass umptions present in alternative procedures, can be easily implemented, and is generalizable to any immunoassay procedure. Methods for incorp orating our results in the estimation of variability for planning and analyzing biomarker experiments are discussed. We provide an example u sing RIA data for aflatoxin B-1 detection, These biomarkers for aflato xin exposure are used in the analysis of serum aflatoxin adduct levels in human and experimental samples, and they are important in hepatoce llular carcinoma research.