Candidate reference measurement procedure for serum 17-hydroxyprogesterone quantification via isotope dilution liquid chromatography–tandem mass spectrometry

Background:Accurate quantification of 17-hydroxyprogesterone(17-OHP)in serum is vital for clinical and research applications.However,inter-laboratory variability in test results exists owing to the lack of a standardized reference measurement procedure(RMP).Therefore,in this study,we developed a highly accurate,cost-effective,and user-friendly candidate RMP(cRMP)for analyzing 17-OHP in serum.Methods:We quantified 17-OHP in serum using a one-step liquid–liquid extraction method with the addition of 17-OHP-^(13)C_(3),followed by liquid chromatography–tandem mass spectrometry.The ability of these methods to suppress interference was evaluated by chromatographic analysis.We assessed accuracy,specificity,the lower limit of quantitation,linearity,and matrix effects by following the standards specified by the Clinical and Laboratory Standards Institute.The consistency between the developed cRMP and the chemiluminescence method was evaluated through experiments with 120 clinical samples.Results:The developed cRMP required only 8 min for accurate quantification of serum 17-OHP without bias from matrix effects or interference from 19 metabolites added as potential interferents.The method exhibited favorable measurement performance,with a quantitation limit of 0.086 ng/mL,linear range of 0.1–400 ng/mL,a total imprecision of≤2.90%,spike recovery of 100.1%–100.6%,and relative deviations from assigned target values(RfB Institution)of−2.91%to 1.10%.The cRMP demonstrated good consistency with the conventional assay(chemiluminescence method),with a correlation coefficient R of 0.96977.Conclusion:A cRMP with high accuracy,cost-effectiveness,and convenient operation was developed for quantifying 17-OHP in serum.Its simplicity and robust performance make it an invaluable addition to the arsenal of analytical tools available for laboratories.This method can enhance the accuracy and reliability of 17-OHP measurements across various laboratories.

国际比对CCQM-P77a血清中氢化可的松测定

The concept of metrological traceability in laboratory medicine has increasingly received considerable recognition worldwide.The Consultative Committee for Amount of Substance (CCQM) which concerned international comparisons and establishment of international equivalence between national laboratories had organized international comparison CCQM-P77a: determination of serum cortisol in 2006.National Institute of Metrology (NIM) took part in the comparison as the representative of China and received good performance according to the results published by CCQM.

高效液相色谱-同位素稀释质谱法测定血清中的苯丙氨酸

Phenylalanine in serum was determined by HPLC isotope dilution mass spectrometry method with D8 labeled phenylalanine as internal standard.Phenylalanine and its isotope labeled analogue were monitored at the transitions m/z=166→120 and 172→126 in multiple reaction monitoring (MRM) mode,respectively,which effectively eliminated the matrix effect.The RSD for 5 repeats measurement of the same sample was 0.5%.Uncertainty was evaluated,which showed the main uncertainty was from the purity assessment of phenylalanine reference material and the variation of method.The presented method can be applied into value assessment of reference material of phenylalanine in serum,and it can be also used for traceability establishment of the measurement of phenylalanine in serum.