Background: Almost all the automated chemiluminescence analyzers in the market cannot detect total 25-hy-droxyvitamin D (25-OH VD) in fingertip blood sample due to the limited volume. This study aimed to develop a relatively simple pretreatment method to obtain plasma from the fingertip blood samples with limited volume and to establish and evaluate the performance of a chemiluminescence immunoassay (CLIA) for detecting 25-OH VD in plasma.
Methods: Fixed volume of plasma was obtained using a special glass capillary to pretreat the fingertip blood. The coated microwells were inserted and the reagents were dispensed into wells in the test strips which can be continuously tested on a small automated chemiluminescence analyzer. The performance of the developed method was evaluated, and the method comparison studies were conducted.
Results: The limit of detection (LOD) was 1.38 ng/mL. The intra-assay precision of the anticoagulant whole blood samples with low, medium, high concentrations of 25-OH VD were 5.45%, 8.20%, 4.97%, respectively. Different metabolic forms of vitamin D have different degrees of cross-interference with antibodies used in the developed method, especially the active metabolites. The cross reactivity (CR) values of 1,25-dihydroxyvitamin D2 (1,25-(OH)2 VD2) and 1,25-dihydroxyvitamin D3 (1,25-(OH)2 VD3) were 5.83% and 9.13%, respectively. The method comparison results of plasma from the venous blood samples showed that correlation coefficient (R2) is 0.94, with a slight positive bias of +1.1% [95% limits of agreement (± 1.96 SD); -25.8% to 28.1%]. Comparing the results detected by the developed method (plasma from fingertip blood samples) with Roche (plasma from anticoagulant venous blood), good linear relativity was noted, with a slight negative bias of -1.2% [95% limits of agreement (± 1.96 SD); -28.7% to 26.4%].
Conclusions: The developed pretreatment method for fingertip blood and the CLIA method can be used to assess the vitamin D status. The correlation of performance and method with the Roche test was good, and the whole test needs only 30 minutes. The developed method can fully satisfy very busy clinical labs.