High total protein impairs appropriate gel barrier formation in BD Vacutainer blood collection tubes.

Fig. 1A. The Bland-Altman plot (Fig. 1B) confirmed this random tendency, which was independent of concentration and of sex, and also demonstrated an amplified dispersion at 25-OHD concentrations Ͼ50 nmol/L. Our data (Fig. 1B) suggested that CLIA results tended to be higher than RIA at low and high concentrations , conversely to data previously published (4). With a threshold of 25-OHD Ն30 nmol/L but Ͻ50 nmol/L to define vitamin D insufficiency, in 33% of the 54 individuals with 25-OHD concentrations classified as insufficient by RIA, 25-OHD concentrations were normalized by CLIA (range: 53.0 –109.5 nmol/L). In contrast , 21% of the 62 study participants with 25-OHD concentrations Ն50 nmol/L by RIA had insufficient 25-OHD concentrations by CLIA (range: 17.5– 48.0 nmol/L). For optimal serum 25-OHD concentration defined as Ͼ75 nmol/L in osteopo-rotic patients, 35% of the 23 patents with 25-OHD concentrations above this threshold by RIA had concentrations below it by CLIA. The RIA method used a primary antibody to 25-OHD in a homogenous phase with a 2nd antibody used as precipitating agent, whereas CLIA used the same primary antibody immobilized onto coated magnetic particles. This antibody interacts differently with the first calibrator [i.e., 17.5 nmol/L (CLIA); 12.5 nmol/L (RIA) with an optional calibrator of 6.25 nmol/L (B/B0: 91%) created by diluting 12.5 at 1:2 as suggested by the manufac-turer], indicating different affinity profiles that are probably responsible for these random results. These divergent results may also be attributable to different calibrators, constituted in either human-(RIA) or horse-based serum (CLIA), different incubation times (90 min by RIA vs 30 min by CLIA), or an insufficient quantity of reagents used to dissociate 25-OHD from its binding protein. Overall, these 2 methods did not similarly classify individuals with reference to well-known arbitrary cutoff values. The random tendency observed whatever the concentrations measured did not permit the definition of a clear strategy concerning patient follow-up, particularly for those needing treatment with respect to their vitamin D status. The discrepancy between these 2 methods is consistent with either important negative (4, 5) or positive (our data) intercepts traducing differences in the assay response to the calibrant matrix. Finally, our results are consistent with the poor correlation previously reported between RIA and CLIA (5) and demonstrate that in disagreement with recently published data (4), a 25-OHD value Ͻ50 nmol/L used to define vitamin D insufficiency with the DiaSorin RIA is not suitable for use with …