Background: The use of ultrafast liquid chromatography (UPLC) without a coupled mass spectrometer could allow accurate evaluation of the different forms of vitamin D by combining the advantages of both immunometric methods (rapid response time, high throughput) and traditional HPLC (better accuracy, precision and specificity) without high initial investment costs. In this study we compared a new UPLC method, an immunometric method and a traditional HPLC method for the evaluation of vitamin D levels in plasma and serum. Methods: The vitamin D levels in 40 plasma samples and 50 serum samples were determined using a ClinRep® 25-OHvitamin D2/D3 Recipe kit (BSN Srl, Castelleone, CR, Italia) and a UPLC Nexera Prominence LC20 system (Shimadzu Italia Srl, Milano, Italia). The plasma samples were also analysed using a LIAISON® 25-OH-vitamin D Total kit (DiaSorin SpA, Saluggia, VC, Italia) and a Liaison diagnostic system, and the serum samples were also analysed using a 25-OH-vitamin D3/D2 Bio-Rad Laboratories Srl (Segrate, MI, Italy) HPLC kit and an Agilent Technologies (Santa Clara, CA, USA) series 1200 HPLC system. To verify repeatability and accuracy a plasma pool and low and high concentration controls were used. Results: The UPLC method displayed good precision for both vitamin D forms (5-7 %). Although at low vitamin D levels inaccuracy was low (2 %), at higher concentrations (>30 ng/ml) inaccuracy ranged from 6.5 % to 8.5 %. For analysing plasma, UPLC indicated vitamin D levels twice those obtained with the DiaSorin kit, as confirmed by Passing-Bablok regression (UPLC va lues = DiaSorin values × 2.38), with a mean percentage difference of 74.5 %. For analysing serum, UPLC indicated higher vitamin D levels than HPLC, with a mean percentage difference of 11.6 %. The regression equation from Passing-Bablok analysis was: UPLC va lues = HPLC values × 1.09 + 0.27. The misclassification rate in relation to clinical decision points commonly used for the diagnosis of mild or moderate vitamin D deficiency (100 ng/ml) was lower than 20 %. Conclusions: The UPLC method indicated higher vitamin D levels than those obtained using the DiaSorin kit and did not appear to be interchangeable without reference interval modification. Despite displaying inaccuracy close to or higher than the desirable limits suggested in the recent literature, the UPLC method indicated vitamin D levels compatible with those obtained by traditional HPLC.
|Translated title of the contribution||Vitamin D: Evaluation of an ultrafast liquid chromatography method (UPLC) and its comparison with a traditional immunoassay and high-performance liquid chromatography (HPLC) technique|
|Number of pages||11|
|Journal||Rivista Italiana della Medicina di Laboratorio|
|Publication status||Published - Sep 2012|
ASJC Scopus subject areas
- Biochemistry, medical
- Medical Laboratory Technology