Giorgia Antonelli1, Andrea Padoan2, Carlo Artusi3, Mariela Marinova3, Martina Zaninotto3, Mario Plebani4. 1. U.O.C. of Laboratory Medicine, Department of Medicine, University of Padova, Padova, Italy. Electronic address: giorgia.antonelli@unipd.it. 2. U.O.C. of Laboratory Medicine, Department of Medicine, University of Padova, Padova, Italy. 3. Department of Laboratory Medicine, University-Hospital of Padova, Padova, Italy. 4. U.O.C. of Laboratory Medicine, Department of Medicine, University of Padova, Padova, Italy; Department of Laboratory Medicine, University-Hospital of Padova, Padova, Italy.
Abstract
OBJECTIVES: The aim of this study was to implement in our routine practice an automated saliva preparation protocol for quantification of cortisol (F) and cortisone (E) by LC-MS/MS using a liquid handling platform, maintaining the previously defined reference intervals with the manual preparation. DESIGN AND METHODS: Addition of internal standard solution to saliva samples and calibrators and SPE on μ-elution 96-well plate were performed by liquid handling platform. After extraction, the eluates were submitted to LC-MS/MS analysis. The manual steps within the entire process were to transfer saliva samples in suitable tubes, to put the cap mat and transfer of the collection plate to the LC auto sampler. Transference of the reference intervals from the manual to the automated procedure was established by Passing Bablok regression on 120 saliva samples analyzed simultaneously with the two procedures. RESULTS: Calibration curves were linear throughout the selected ranges. The imprecision ranged from 2 to 10%, with recoveries from 95 to 116%. Passing Bablok regression demonstrated no significant bias. CONCLUSIONS: The liquid handling platform translates the manual steps into automated operations allowing for saving hands-on time, while maintaining assay reproducibility and ensuring reliability of results, making it implementable in our routine with the previous established reference intervals.
OBJECTIVES: The aim of this study was to implement in our routine practice an automated saliva preparation protocol for quantification of cortisol (F) and cortisone (E) by LC-MS/MS using a liquid handling platform, maintaining the previously defined reference intervals with the manual preparation. DESIGN AND METHODS: Addition of internal standard solution to saliva samples and calibrators and SPE on μ-elution 96-well plate were performed by liquid handling platform. After extraction, the eluates were submitted to LC-MS/MS analysis. The manual steps within the entire process were to transfer saliva samples in suitable tubes, to put the cap mat and transfer of the collection plate to the LC auto sampler. Transference of the reference intervals from the manual to the automated procedure was established by Passing Bablok regression on 120 saliva samples analyzed simultaneously with the two procedures. RESULTS: Calibration curves were linear throughout the selected ranges. The imprecision ranged from 2 to 10%, with recoveries from 95 to 116%. Passing Bablok regression demonstrated no significant bias. CONCLUSIONS: The liquid handling platform translates the manual steps into automated operations allowing for saving hands-on time, while maintaining assay reproducibility and ensuring reliability of results, making it implementable in our routine with the previous established reference intervals.
Authors: A Christler; E Felföldi; M Mosor; D Sauer; N Walch; A Dürauer; A Jungbauer Journal: Bioprocess Biosyst Eng Date: 2019-12-07 Impact factor: 3.210