Reference methods and Certified Reference Materials are necessary to assess and improve comparability and reliability of medical tests. However, development of reference measurement systems is more complex for some biomarkers than for others. While the state of the art in metrology is mature enough to address the case of most metabolites and electrolytes quite easily, absolute quantification of protein biomarker is much more difficult due to many factors. Hyphenated quantification techniques are often necessary to measure low concentrations of large proteins in complex matrices. As different isoforms can have different clinical significance, structural heterogeneity of proteins complicates the situation even further and requires to carefully define the measurand. Due to the relationship between folding and activity, absolute quantification of some complexity proteins remains a challenge and some biomarkers are so large that they can even be considered as bionanoparticles. Through different case studies, this presentation will highlight a number of challenges associated with absolute quantification of biological entities and establishment of traceability chains in clinical diagnostics.
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