alexa A bottom-up approach in estimating the measurement uncertainty and other important considerations for quantitative analyses in drug testing for horses.
Immunology

Immunology

Immunochemistry & Immunopathology

Author(s): Leung GN, Ho EN, Kwok WH, Leung DK, Tang FP,

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Abstract Quantitative determination, particularly for threshold substances in biological samples, is much more demanding than qualitative identification. A proper assessment of any quantitative determination is the measurement uncertainty (MU) associated with the determined value. The International Standard ISO/IEC 17025, "General requirements for the competence of testing and calibration laboratories", has more prescriptive requirements on the MU than its superseded document, ISO/IEC Guide 25. Under the 2005 or 1999 versions of the new standard, an estimation of the MU is mandatory for all quantitative determinations. To comply with the new requirement, a protocol was established in the authors' laboratory in 2001. The protocol has since evolved based on our practical experience, and a refined version was adopted in 2004. This paper describes our approach in establishing the MU, as well as some other important considerations, for the quantification of threshold substances in biological samples as applied in the area of doping control for horses. The testing of threshold substances can be viewed as a compliance test (or testing to a specified limit). As such, it should only be necessary to establish the MU at the threshold level. The steps in a "Bottom-Up" approach adopted by us are similar to those described in the EURACHEM/CITAC guide, "Quantifying Uncertainty in Analytical Measurement". They involve first specifying the measurand, including the relationship between the measurand and the input quantities upon which it depends. This is followed by identifying all applicable uncertainty contributions using a "cause and effect" diagram. The magnitude of each uncertainty component is then calculated and converted to a standard uncertainty. A recovery study is also conducted to determine if the method bias is significant and whether a recovery (or correction) factor needs to be applied. All standard uncertainties with values greater than 30\% of the largest one are then used to derive the combined standard uncertainty. Finally, an expanded uncertainty is calculated at 99\% one-tailed confidence level by multiplying the standard uncertainty with an appropriate coverage factor (k). A sample is considered positive if the determined concentration of the threshold substance exceeds its threshold by the expanded uncertainty. In addition, other important considerations, which can have a significant impact on quantitative analyses, will be presented. This article was published in J Chromatogr A and referenced in Immunochemistry & Immunopathology

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