Sensitive and quantitative, 10-min immunofluorometric assay for D-Dimer in whole blood

IntroductionNormal concentrations of D-Dimer can be used to exclude venous thromboembolism (VTE). However, methods for sensitive and quantitative D-Dimer measurements at the point-of-care (POC) are still limited.Materials and methodsWe developed a 10-min, non-competitive immunofluorometric assay for D-Dimer in citrated whole blood and plasma using pre-dispensed reagents dried in single assay wells. The simple, automated assay procedure comprises a 1 : 50 sample dilution, one-step incubation, washing, and time-resolved fluorometric measurement directly from the wet well surface.ResultsThe limits of detection (background + 3SD) and quantification (CV < 15%) were 0.05 and 0.2 mg/L D-Dimer, respectively, and the assay was linear up to 400 mg/L. Correlations to Roche TinaQuant (r = 0.726, n = 200) and Biopool Auto·Dimer (r = 0.190, n = 149) were carried out using citrated plasma. Diagnostic sensitivity, specificity, and negative (NPV) and positive (PPV) predictive values were 98.7%, 64.4%, 99.1% and 55.1%, and 92.2%, 81.0%, 95.9% and 68.3%, respectively, using cut-off values of 0.6 and 1.0 mg/L, respectively, in outpatients with deep vein thrombosis (DVT) and/or pulmonary embolism (PE) (n = 77) compared with outpatients with various other diseases (n = 174). The within- and between-run CVs near the cut-off values were ≤ 10% in both whole blood and plasma. The 95th percentile upper range in apparently healthy individuals was 0.68 mg/L of whole blood (n = 101).ConclusionsThe high sensitivity and NPV suggest that the rapid immunofluorometric assay could be valuable for rapid exclusion of VTE in outpatients. With appropriate cut-offs, the assay could potentially be used as a stand-alone test or combined with clinical probability assessment, but further studies are required.