Target analyte quantification by isotope dilution LC-MS/MS directly referring to internal standard concentrations--validation for serum cortisol measurement.
BACKGROUND
Isotope dilution LC-MS/MS methods used in the clinical laboratory typically involve multi-point external calibration in each analytical series. Our aim was to test the hypothesis that determination of target analyte concentrations directly derived from the relation of the target analyte peak area to the peak area of a corresponding stable isotope labelled internal standard compound [direct isotope dilution analysis (DIDA)] may be not inferior to conventional external calibration with respect to accuracy and reproducibility.


METHODS
Quality control samples and human serum pools were analysed in a comparative validation protocol for cortisol as an exemplary analyte by LC-MS/MS. Accuracy and reproducibility were compared between quantification either involving a six-point external calibration function, or a result calculation merely based on peak area ratios of unlabelled and labelled analyte.


RESULTS
Both quantification approaches resulted in similar accuracy and reproducibility.


CONCLUSIONS
For specified analytes, reliable analyte quantification directly derived from the ratio of peak areas of labelled and unlabelled analyte without the need for a time consuming multi-point calibration series is possible. This DIDA approach is of considerable practical importance for the application of LC-MS/MS in the clinical laboratory where short turnaround times often have high priority.
Institute of Laboratory Medicine, University of Munich, Marchioninistra s e 15, 81377 Munich, Germany. barbara.maier@med.uni-muenchen.de
Barbara
B
Maier
Michael
M
Vogeser
D002138:Calibration; D002851:Chromatography, High Pressure Liquid / Q000592:standards; D003903:Deuterium / Q000737:chemistry; D006801:Humans; D006854:Hydrocortisone / Q000097:blood* / Q000592:standards; D007553:Isotope Labeling; D011786:Quality Control; D053719:Tandem Mass Spectrometry / Q000592:standards
D003903:Deuterium; D006854:Hydrocortisone