|= Emerging. More than 5 years before clinical availability. (9.82%)|
|= Expected to be clinically available in 1 to 4 years. (12.95%)|
|= Clinically available now. (22.77%)|
Authors: Antonelli Giorgia (1), Artusi Carlo (2), Sciacovelli Laura (2) and Plebani Mario (1,2)
The examination procedure (EP) salivary cortisol (sF) is accredited according to the ISO15189:2012 since 2016. This EP is a laboratory developed method, using home-made calibrators; according to the requirement 220.127.116.11, this method was validated, producing a validation certificate. Recently CE-IVD calibrators were available and our aim was to substitute the home-made calibrators with these ones. Comparing the obtained results, a significant bias was obtained for the salivary cortisol and cortisone ratio (sFEr), so the reference intervals (RI) were modified according to the Passing Bablok regression. The new RI had to be verified, according to CLSI EP28. Twenty healthy volunteers were recruited and saliva was collected. The results were suitable and a new validation certificate was produced.
Salivary cortisol (sF) and cortisone (sE) is determined by an home brew LC-MS/MS, with home-made calibrators and internal quality controls [1,2]. The clinicians can request the salivary cortisol and cortisone ratio (sFEr): in the report, only sF and sFEr results are described, with the appropriate reference intervals (RI). sF is an examination procedure (EP) which is accredited ISO15189:2012  since 2016. According to the requirement 18.104.22.168 , this EP was validated , producing a validation certificate. sFEr was not accredited since external quality controls are not available for sE. Recently CE-IVD calibrators for both cortisol and cortisone were available and the aim of this work was to substitute the home made calibrators with the commercial ones, since a higher standardization is guaranteed.
• Cortisol (F), cortisone (E) from Sigma (Sigma-Aldrich, St. Louis, MO, USA);
• d4-cortisol (d4-F) and d7-cortisone (d7-E) from Sigma (Sigma-Aldrich, St. Louis, MO, USA);
• Saliva CE-IVD calibrators for cortisol and cortisone (MassChrom® Chromsystem)
• 300µL saliva/calibrators/controls extracted with µ-elution Oasis® HLB plate (Waters, MA;USA) - on Tecan Freedom EVO 150 liquid handling platform (Tecan, Männedorf, Switzerland)
• Agilent HPLC series 1200 (Agilent Technologies, Palo Alto, USA)
• Triple quadrupole mass spectrometer Agilent 6430 (Agilent Technologies, Palo Alto, USA)
• Saliva from 20 healthy volunteers
Saliva samples from routine analysis (n=160) were used to compare home-made calibrators with CE-IVD calibrators. No significant bias was observed for sF. A significant bias resulted for sFEr (Bland - Altman -33,6%, Passing Bablok regression y=0,71x+0,01). The RI for sF was not changed, while those for sFEr were recalculated according to the obtained regression. The new RI were verified according to CLSI EP28 . Saliva samples were collected from 20 healthy volunteers: only 2 were outside from the obtained RI.
Conclusions & Discussion
The new RI are appropriate for the intended use. When a modification is made on a ISO 15189 accredited procedure, the change has to be considered. Moreover, all the data obtained have to be recorded and a new validation certificate has to be released.
References & Acknowledgements:
1. Antonelli G, Ceccato F, Artusi C, Marinova M, Plebani M. Salivary cortisol and cortisone by LC-MS/MS: validation, reference intervals and diagnostic accuracy in Cushing's syndrome. Clin Chim Acta. 2015;451(Pt B):247-51
2. Antonelli G, Padoan A, Artusi C, Marinova M, Zaninotto M, Plebani M. Automated saliva processing for LC-MS/MS: Improving laboratory efficiency in cortisol cortisone testing. Clin Biochem. 2016;49(6):518-520
3. International Organization for Standardization (ISO). Medical laboratories-Particular requirements for quality and competence. ISO 15189:2012.
4. Antonelli G, Padoan A, Aita A, Sciacovelli L, Plebani M. Verification or validation, that is the question. J Lab Precis Med 2017;2:58.
5. Clinical and Laboratory Standards Institute (CLSI). Defining, establishing, and verifying reference intervals in the clinical laboratory; approved guideline—third edition, CLSI document EP28-A3c, Clinical and Laboratory Standards Institute, ISBN: 1-56238-682-4, 2010.
IP Royalty: no
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