Analytical and Clinical Performance of a New Point of Care LABGEO IB D-Dimer Test for Diagnosis of Venous Thromboembolism

Transcription

1 254 Available online at Annals of Clinical & Laboratory Science, vol. 44, no. 3, 2014 Analytical and Clinical Performance of a New Point of Care LABGEO IB D-Dimer Test for Diagnosis of Venous Thromboembolism Jaewoo Song 1, Tae Dong Kweon 2, Yeajin Song 1, Eun Young Lee 1, Sue Jung Kim 1, and Rojin Park 3 1 Department of Laboratory Medicine, Yonsei University College of Medicine, Seoul, 2 Department of Anesthesiology and Pain Medicine, Yonsei University College of Medicine, Seoul, and 3 Department of Laboratory Medicine, Soonchunhyang University Seoul Hospital, Seoul, Korea Abstract. LABGEO IB D-dimer Test is a newly developed POC D-dimer assay and the first commercially available POC immunoassay instrument that exploits the disk rotation method for extraction of plasma. Citrate plasma was obtained from 201 apparently healthy subjects and 91 patients suspected for VTE, and their D-dimer level was measured by the LABGEO IB D-Dimer Test (LABGEO D-dimer) and HemosIL D-dimer test as a comparative method. To examine the effect of blood cells and anticoagulant, paired blood samples anticoagulated by heparin and citrate were obtained from various postoperative patients. The overall diagnostic performance of LABGEO IB D-dimer and HemosIL was comparable with similar area under ROC curve (p=0.79). The cut-off levels recommended by manufacturers (LABGEO D-dimer: 0.45 μg/ml fibrinogen equivalent unit (FEU), HemosIL: 0.23 μg/ml D-dimer unit (DDU)) and those yielding highest diagnostic efficiency (LABGEO D-dimer: 1.41 μg/ml FEU; HemosIL: 0.85 μg/ml DDU), were chosen for the evaluation. For LABGEO D-dimer negative predictive value (NPV), positive predictive value (PPV), sensitivity, specificity, and negative likelihood ratio (LR-neg) were %, 67 89%, %, 53 89% and For HemosIL D-dimer, NPV, PPV, sensitivity, specificity and LR-neg were %, 76 95%, %, 70 96% and , all comparable to results for LABGEO D-dimer. LABGEO D-dimer test demonstrated acceptable performance when used for the VTE diagnostic work-up. Keywords: venous thromboembolism, point-of-care, LABGEO IB D-dimer Test Introduction D-dimer is a sensitive marker of deep vein thrombosis (DVT) and pulmonary embolisms (PE) and an effective aid in the diagnosis of these diseases [1, 2]. However, D-dimer is also elevated in a wide variety of non-thrombotic conditions including infection, trauma, atrial fibrillation, and stroke [1, 2]. Therefore, while D-dimer cannot be used to confirm the diagnosis of venous thromboembolism (VTE), its high sensitivity renders it as a useful tool to rule out this condition in symptomatic subjects with a high negative predictive value. The annual incidence of VTE in the general population is 0.1%, with approximately two-thirds of the cases being manifested as DVT alone and Address correspondence to Rojin Park, MD., PhD., Department of Laboratory Medicine, Soonchunhyang University Seoul Hospital, 59 Daesagwan-ro, Yongsan-gu, Seoul , Korea; phone: ; fax: ; e mail: rpark@schmc.ac.kr one-third including PE as a manifestation [3]. A population-based retrospective study revealed that mortality rates within 7 days after a VTE diagnosis were approximately 4% for DVT patients without PE and 29% for patients with PE; the corresponding 1 year mortality rates after incident VTE were 15% and 43%, respectively [4]. Furthermore, approximately half of the patients with incident DVT will suffer long-term complications as a result of the thrombotic episode [5]. PE presents with diverse non-specific symptoms, and its diagnosis mainly depends on diagnostic imaging tests which are laborious and expensive. Only a small proportion of patients with suspected PE are actually proven to have the disease. Hence, a negative D-dimer measurement in combination with a low Wells score can exclude venous thromboembolism in symptomatic outpatients [6], thus reducing the frequency with which confirmatory imaging tests are required, and subsequently lowering overall patient /14/ by the Association of Clinical Scientists, Inc.

2 Performance of LABGEO IB D-dimer Test 255 Table 1. General and clinical characteristics of patients included in evaluation of diagnostic performance of LABGEO D-dimer for VTE. VTE (n=44) Non-VTE (n=47) Age a 66 (1 90) 59 (20 90) Female (%) 22 (50) 18 (38) D-dimer (μg/ml) a LABGEO 3.46 (0.56 > 4.00) 0.43 (< ) HemosIL 2.73 ( ) 0.16 (< ) Diagnosis Deep vein thrombosis (13) Stable coronary artery diseases (10) Deep vein thrombosis, Acute coronary syndrome (9) not involving lower extremities (2) b Acute myocardial infarction (7) Pulmonary thromboembolism (29) Respiratory diseases (5) Others (4) c Chest pain, cause not determined (9) Dyspnea, cause not determined (3) a Median (range) b Superior mesenteric vein, hepatic veno-occlusive disease c Atrial fibrillation, mitral regurgitation, diffuse large B cell lymphoma, spasmodic dysphonia costs. By performing the D-dimer test in point-ofcare (POC) locations such as the emergency department, physicians can rapidly exclude patients with low pre-test probability of VTE based on negative test results and can immediately refer patients with positive test results for confirmatory imaging tests. The Samsung LABGEO IB D-Dimer Test (LABGEO D-dimer) is a new POC immunoassay that can provide quantitative D-dimer test results in both whole blood and plasma specimens in 20 minutes. In this study, we evaluated the LABGEO D-dimer Test s ability to rule out VTE by using frozen plasma specimens collected from consecutively presenting symptomatic patients. We then compared the results with those from a well-established laboratory D-dimer test (HemosIL D-dimer HS Test) which is based on latex enhanced immunoturbidimetry. We also compared LABGEO D-dimer results in fresh whole blood and plasma specimens using different anticoagulants. Materials and Methods Patients and samples. Plasma specimens (using sodium citrate as the anticoagulant) from 201 apparently healthy subjects (100 male and 101 female subjects) were evaluated using both the LABGEO IB D-dimer Test (LABGEO D-dimer) and the comparative assay, HemosIL D-dimer HS (HemosIL D-dimer) on the ACL-TOP coagulometer (Instrument Laboratory, Lexington, MA, USA) in order to verify the reference intervals of both assays. The concentration of D-dimers was expressed in the fibrinogen equivalent unit (μg/ml FEU) for LABGEO D-dimer and D-dimer unit (μg/ml DDU) for HemosIL D-dimer. FEU means fibrinogen mass from which D-dimer is derived, and 2 μg/ml FEU is similar to 1μg/ml DDU in immunoreactivity. Frozen plasma specimens (using sodium citrate as the anticoagulant) were also obtained from subjects with symptoms suggestive of VTE in order to evaluate the ability of each assay to rule out VTE. Of the 91 subjects from whom specimens were collected and tested, 44 had a confirmed diagnosis of VTE. Matching whole blood specimens (using both lithium heparin and sodium citrate as anticoagulants) were also obtained from 49 independent subjects in order to compare D-dimer results in different specimen matrices. Blood was drawn from postoperative surgical patients whose D-dimer levels spanned the analytical measurement range of the LABGEO D-dimer ( μg/ml FEU). Each whole blood specimen was tested in one replicate using LABGEO D-dimer; the whole blood specimens were then processed to obtain plasma, and each plasma specimen was tested in one replicate using LABGEO D-dimer.

3 256 Annals of Clinical & Laboratory Science, vol. 44, no. 3, 2014 Figure 1. Correlation between matching LABGEO D-dimer and HemosIL D-dimer.results obtained from 268 subjects with D-dimer levels within the LABGEO analytical measuring range of µg/ml. Figure 2. Receiver operating characteristic (ROC) curves of LABGEO (solid line) and HemosIL (dotted line) for diagnosing VTE. Ninety one consecutive patients suspected for VTE were included. Cut-off values subsequently evaluated in detail are marked with (LABGEO D-dimer) and (HemosIL D-dimer). LABGEO IB D-Dimer Test. Using lithium heparin or sodium citrate as the anticoagulant, the LABGEO D-dimer Test, otherwise known as LABGEO D-dimer, is a rapid point-of-care (POC) immunoassay for the in vitro quantitative determination of cross-linked fibrin degradation products containing D-Dimer in human whole blood or plasma. The D-Dimer Test is intended for use in conjunction with the Samsung LABGEO IB10 Analyzer. After the addition of the blood specimen to the test disc, the entire test is performed within the Samsung LABGEO IB10 Analyzer, which provides control of the temperature of the disc, as well as the sequence of centrifugal flow, mixing, incubation time, final signal measurement, quantitation and reporting of results. The LABGEO D-dimer combines microfluidics with immunochromatography for the preparation of the plasma and the measurement of the analyte. Both modules are embedded in a disc of the same size and shape as a compact disc. The centripetal force generated by the rotation of the disc rapidly prepares cellfree plasma, and the combination of active flow and capillary action force it through the microfluidic channel to the immunochromatography module to rehydrate, solubilize, and mix with freeze-dried immunoconjugates. The immune complex forms a colored band on the chromatography membrane and the density of the band proportional to the concentration of D-dimer in the specimen is quantitatively measured optically. The whole process takes 20 minutes. The test disc includes a positive internal control to ensure that the test has been performed properly. Statistics. D-dimer measurements obtained from the LABGEO and HemosIL D-dimer assays were compared using sodium citrate plasma specimens from 91 consecutively presenting patients with symptoms suggestive of VTE. Receiver operating characteristic (ROC) curves were constructed and the area under the curve (AUC) was calculated. For each assay, the negative predictive value (NPV), positive predictive value (PPV), sensitivity, specificity, negative likelihood ratio (LR-neg), efficiency, and post-test NPV were calculated using the manufacturer s recommended cut-off, along with the corresponding 95% confidence interval (CI) for each measure. Correlation and comparability between LABGEO and HemosIL D-dimer test results were evaluated using a population of 268 subjects (both apparently healthy individuals and subjects with symptoms suggestive of VTE). The correlation was determined using Spearman s rank correlation coefficient, and the method comparison was performed by the Passing-Bablok regression analysis. The D-dimer levels measured in lithium heparin and sodium citrate whole blood and plasma were also compared using Passing- Bablok regression analyses. The analyses were conducted using Analyse-it for Microsoft Excel ver (Analyse-it Software, UK) and MedCalc ver (MedCalc Software, Belgium).

4 Performance of LABGEO IB D-dimer Test 257 Table 2. Diagnostic performance (NPV, PPV, sensitivity, specificity, LR-neg, efficiency, post-test NPV) of the LABGEO and HemosIL D-dimer assays for VTE according to variable cut-off levels. Cut-off NPV PPV Sensitivity Specificity LR-neg Efficiency Post-test NPV d (μg/ml) (95% CI) (95% CI) (95% CI) (95% CI) (95% CI) (95% CI) 0.45 a 100 (89 100) 67 (54 78) 100 (93 100) 53 (38 68) (96-100) 0.82 b 97 (86 100) 80 (67 89) 98 (88 100) 77 (62 88) 0.03 ( ) (97-100) LABGEO (83 99) 84 (71 93) 95 (85 99) 83 (69 92) 0.05 ( ) (96-100) (FEU) 1.41 c 93 (82 99) 89 (76-96) 93 (81 99) 89 (77 97) 0.08 ( ) (96-100) (80 98) 91 (78-98) 91 (78 98) 91 (80 98) 0.10 ( ) (95-100) (77 97) 91 (78-97) 89 (75 96) 91 (80 98) 0.12 ( ) (95-99) 0.23 a 100 (91 100) 76 (63-86) 100 (93 100) 70 (55 83) (97-100) 0.31 b 100 (92 100) 80 (67-90) 100 (93-100) 77 (62 88) (98-100) HemosIL (83 99) 81 (67-90) 95 (85-99) 79 (64 89) 0.06 ( ) (96-100) (DDU) (84 99) 86 (73-94) 95 (85 99) 85 (72 94) 0.05 ( ) (96-100) (79 98) 89 (76-96) 91 (78 98) 89 (77 97) 0.10 ( ) (95-100) 0.85 c 90 (78 97) 95 (84-99) 89 (75 96) 96 (86 99) 0.12 ( ) (95-99) a Cut-off level recommended by the manufacturer b Cut-off level was chosen such that the specificity was comparable to cut-off of maximum efficiency and LR-neg was as small as possible. c Cut-off level of maximum efficiency (proportion of correctly classified results) d The post-test predictive value was calculated with supposed VTE prevalence of 15.7% [21]. Results Assay characteristics. The precision of the LABGEO D-dimer assay was estimated using sodium citrate plasma specimens at 2 different concentrations of D-dimer over a 15-day period, with 2 runs per day and 4 replicates per run. The within-run coefficients of variation (%CVs) at mean D-dimer test concentrations of 0.45 μg/ml and 0.84 μg/ml were 6.7% and 8.6% respectively, and the total %CVs at these test concentrations were 6.7% and 9.0%, respectively. We compared LABGEO and matching HemosIL D-dimer test results from a total of 292 subjects comprised of both apparently healthy subjects and subjects presenting with symptoms suggestive of VTE. Of these 292 subjects, 268 had both LABGEO and HemosIL D-dimer levels within the analytical measurement range for the LABGEO assay ( μg/ml). The results from both assays were highly correlated, with a Spearman s rank correlation coefficient of 0.83 (95% CI: ). Overall, the relationship between the 2 assay results was not linear. When the analysis was limited to a narrower range (LABGEO D-dimer level up to 2.5 μg/ml) by excluding very high values that are irrelevant to a diagnostic decision, a linear relationship (cusum linearity test p>0.1) was observed between HemosIL and LABGEO D-dimer results. Assay performance in ruling out VTE in a symptomatic population. The reference interval of LABGEO D-dimer was determined from 201 healthy individuals (age range of 23 to 85). The estimate of the 95% upper limit of the reference interval was 0.55 μg/ml Fibrinogen Equivalent

5 258 Annals of Clinical & Laboratory Science, vol. 44, no. 3, 2014 Units (FEU) for LABGEO (95% CI: μg/ml); the corresponding estimate for HemosIL D-dimer was 0.23 μg/ml D-dimer Units (DDU) (95% CI: μg/ml). Both upper limits were not significantly different from those provided by the manufacturers (0.45 μg/ml FEU and 0.24 μg/ml DDU; p>0.05 in each case). LABGEO and HemosIL D-dimer levels were higher in subjects over 50 years of age (p< for each assay) and in female subjects (p< for each assay). Among 91 patients suspected for VTE, 44 patients were diagnosed as VTE by imaging studies (computed tomography or ultrasonography). For the remaining 47 patients, the final diagnoses were determined based on medical records (Table 1). The ROC curves for the two assay methods are displayed in Figure 1. The area under the ROC curve was for LABGEO (95% CI: ) and for HemosIL (95% CI: ); there was no significant difference between the areas under the ROC curves for the two assays (p=0.79). With respect to LABGEO D-dimer results obtained from these 91 patients suspected of VTE, several putative clinical cut-off levels were selected, and the sensitivity, specificity, PPV, NPV, efficiency (i.e. the proportion of correctly classified cases) and negative likelihood ratio (LR-neg, i.e. the ratio between the probability of a negative test result in subjects with confirmed VTE and the probability of a negative test result in subjects without VTE) were computed separately for each putative cut-off. The results are summarized in Table 2. All D-dimer results obtained from patients with confirmed VTE were above the respective assay cut-offs as recommended by the manufacturers (0.45 μg/ml FEU for LABGEO D-dimer and 0.23 μg/ml DDU for HemosIL D-dimer). Using the LABGEO cut-off level of 0.45 μg/ml FEU, the NPV, PPV, sensitivity and specificity were 100% (95% CI: 89% 100%), 67% (95% CI: 54% 78%), 100% (95% CI: 93% 100%) and 53% (95% CI: 38% 68%) respectively, with an LR-neg of For HemosIL D-dimer, the NPV, PPV, sensitivity, and specificity at the cutoff level of 0.23 μg/ml DDU were 100% (95% CI: 91% 100%), 76% (95% CI: 63%-86%), 100% (95% CI: 93% 100%) and 70% (95% CI: 55% 83%) respectively, with an LR-neg of For LABGEO D-dimer, at the cut-off level of 1.41 μg/ml FEU which yielded the highest diagnostic efficiency (91%), the NPV, PPV, sensitivity and specificity were 93% (95% CI: 82% 99%), 89% (95% CI: 76%-96%), 93% (95% CI: 81% 99%) and 89% (95%CI: 77 97) with an LR-neg of For HemosIL D-dimer, at the cut-off level of 0.85 μg/ml DDU which yielded the highest diagnostic efficiency (92%), the NPV, PPV, sensitivity and specificity were 90% (95% CI: 78% 97%), 95% (95% CI: 84%-99%), 89% (95% CI: 75% 96%) and 96% (95% CI: 86% 99%), with an LR-neg of Effect of blood cells and anticoagulants. LABGEO D-dimer results obtained from whole blood specimens correlated well with results obtained from matching plasma specimens (Figure 3). When comparing citrate whole blood to matching citrate plasma measurements, a Passing- Bablok regression analysis yielded a y-intercept of 0.04 μg/ml FEU (95% CI: ) and a slope of 0.99 (95% CI: ). It also yielded a Spearman correlation coefficient of and a bias at the clinical cut-off (0.45 μg/ml FEU) of 7.1%. A comparison of matching heparin whole blood vs. plasma results yielded a y-intercept of 0.00 μg/ml FEU (95% CI: ), a slope of 1.03 (95% CI: ), correlation coefficient of and a bias of 2.5% at the cutoff. For matching heparin plasma vs. citrate plasma results, the y-intercept and slope were 0.03 μg/ml FEU (95% CI: ) and 0.97 (95% CI: ), respectively (correlation coefficient: 0.975, bias at the cut-off: 3.7%). A comparison of matching LABGEO heparin whole blood vs. citrate whole blood results yielded a y- intercept of 0.01 μg/ml FEU (95% CI: ) and a slope of 0.97 (95% CI: ). The correlation coefficient was and the bias at the cut-off was only -0.9% (Figure 3). In contrast, the effect of anticoagulant on HemosIL D-dimer plasma results was substantial; a comparison of matching HemosIL heparin plasma vs. citrate plasma measurements yielded a y-intercept of μg/ml DDU (95% CI: ), a slope of 1.15 (95% CI: ), a Spearman rank correlation coefficient of 0.991, and a bias at the clinical threshold of 0.23 μg/ml DDU of +13.9%.

6 Performance of LABGEO IB D-dimer Test 259 Figure 3. Comparison between LABGEO D-dimer levels measured from whole blood and plasma anticoagulated by citrate and heparin. Comparability was tested with Passing-Bablok regression (blue solid line represents the fitted regression equation; blue dashed lines represent the 95% confidence bounds; grey line represents the identity line). (A) Comparison between citrate whole blood and citrate plasma. (B) Comparison between heparin whole blood and heparin plasma. (C) Comparison between heparin plasma and citrate plasma. (D) Comparison between heparin whole blood and citrate whole blood. Discussion In our study, the LABGEO D-dimer, a newly developed POC D-dimer assay, was evaluated for its analytical and diagnostic performance. LABGEO, to the extent of our knowledge, is the first commercially available POC immunoassay instrument that exploits the disk rotation method used for the extraction of plasma. The precision of the LABGEO D-dimer, as evaluated at concentrations of 0.45 μg/ ml and 0.84 μg/ml FEU, was comparable to those of other POC D-dimer assays appearing in the literature [7-9]. LABGEO D-dimer measurements were generally higher than matching HemosIL D-dimer measurements, as shown in Figure 1. The difference between LABGEO and HemosIL results can largely be attributable to the fact that LABGEO reports the results in FEU units, whereas HemosIL reports the results in DDU units. The difference can also be related to the specificity of the antibodies and their preference for certain binding conditions such as a high molecular weight fibrin complex or a low molecular weight fibrin degradation product [10]. Regarding calibration, Meijer et al. pointed out the differing response of D-dimer assays to high and

7 260 Annals of Clinical & Laboratory Science, vol. 44, no. 3, 2014 low D-dimer levels as observed in our study. They also introduced a method of transforming assay specific calibration result to that of reference method for the harmonization of quantitative D-dimer assays [11]. This issue can be further addressed to improve the standardization of present and future POC D-dimer assays. The main clinical use of D-dimer in managing VTE is to rule out the condition in suspected patients, a checkpoint before proceeding to further expensive and invasive diagnostic workups [12-15]. Thus, the requirements of high sensitivity and high negative predictive value are of paramount importance for D-dimer assays in the differential diagnosis of conditions with high mortality rates such as pulmonary embolisms. The overall diagnostic performance of the LABGEO D-dimer, as evaluated by the ROC curve analysis, was comparable to that of HemosIL D-dimer in our study; equivalent AUC estimates were obtained for both assays. We evaluated detailed diagnostic characteristics with variable cutoff levels that were chosen following different diagnostic rationales. Using cut-off levels provided by manufacturer, the NPV, PPV, sensitivity, and specificity of the two assays were able to be compared. The performance characteristics were also comparable when the cut-off levels were set to maximize the efficiency or to minimize the LR-neg. The sensitivity of the LABGEO D-dimer was 95% or greater over a range of putative cut-off levels from 0.45 μg/ml and 1.17 μg/ml FEU; however, the NPV was maximized at 100% when applying the manufacturer s recommended cut-off of 0.45 μg/ml FEU. In addition to its diagnostic performance, Geersing et al. also addressed the user friendliness of POC instruments in their evaluation study of 5 POC D-dimer assays [16]. Time spent in warming up the LABGEO D-dimer instrument was about 2 minutes. If whole blood was used, it took about 45 minutes from turning on the instrument to obtaining the sample s first result. During this period, the instrument warmed up, and an electronic quality control disc, quality control material and a patient specimen were run. The analysis time of LABGEO is 20 minutes per specimen, compared with the assay times for other POC assays which ranges from 10 to 38 minutes [16]. When considering the frequency of quality control measurements that are adjustable to local laboratory requirements and the fact that personnel can run the assay with minimal training, the assay itself can be considered userfriendly. Overall, LABGEO D-dimer fulfilled the requirements needed for outpatient or emergency situations such as ease of operation, low betweenoperator variability, and rapid turnaround time. As mentioned above LABGEO adopted a unique method to remove cellular components of blood. Whole blood is applied to the inlet on the disk that contains the sample processing and assay modules together. The blood is centrifuged by the rotation of the disk, and extracted plasma is transferred to the assay module. Haeberle et al. introduced the method and demonstrated that plasma purified in this way was acceptable for subsequent on-disk processing for various assays [17]. As the latest achievement, plasma purity superior to 99.9% was reported when using disk rotation [18]. The method is basically the same as the technique of plasma preparation currently used in clinical laboratories worldwide. Other innovative and miniaturized methods such as microchip formatted sedimentation, microfiltration, and cell deviation have also been introduced but still have critical problems concerning yield and purity of plasma, blood cell clogging, and blood cell contamination [19]. With this background, we examined the accordance of the results obtained from both heparinized and citrated plasma and whole blood across the entire analytical measurement range of LABGEO D-dimer. The whole blood and plasma results showed strong correlation and comparability both for heparin and citrate anticoagulants. The correlation was also acceptable between different anticoagulants. The correlation and comparability of LABGEO results in different matrices improved when the results in the lower two thirds of the measurable range, which were clinically more relevant, were analyzed (data not included). The between-anticoagulant comparability was not achieved to the same extent with the HemosIL assay, which showed a significant proportional bias. Interestingly, both the strongest correlation and the narrowest dispersion around the regression line were achieved between citrated and heparinized whole blood (R 2 : 0.98 vs ). Possibly, there exists a certain difference between plasma prepared by the LABGEO disk and that

8 from a laboratory centrifuge. This was not addressed in our study but can be further investigated in a future study. Fukuda et al. reported that the correction of whole blood D-dimer results for hematocrit improved the correlation of whole blood vs. plasma results in their evaluation of the PATHFAST D-dimer assay [20]. It is not clear if the hematocrit influenced the measurement of D-dimer by LABGEO. In conclusion, the LABGEO D-dimer test demonstrated acceptable performance when used for VTE diagnostic work-up. Our study is subject to limitations of small sample size, an over-representation of coronary heart diseases in the non-vte group, and a short period of following up the non-vte patients to rule out VTE. These advocate the need for continued further investigation from various clinical points of view. Acknowledgements This work was supported by the Soonchunhyang University Research Fund. References 1. Torbicki A, Perrier A, Konstantinides S, Agnelli G, Galie N, Pruszczyk P, Bengel F, Brady AJ, Ferreira D, Janssens U, Klepetko W, Mayer E, Remy-Jardin M, Bassand JP. Guidelines on the diagnosis and management of acute pulmonary embolism: the Task Force for the Diagnosis and Management of Acute Pulmonary Embolism of the European Society of Cardiology (ESC). European Heart Journal. 2008; 29(18): Bates SM, Jaeschke R, Stevens SM, Goodacre S, Wells PS, Stevenson MD, Kearon C, Schunemann HJ, Crowther M, Pauker SG, Makdissi R, Guyatt GH. Diagnosis of DVT: Antithrombotic Therapy and Prevention of Thrombosis, 9th ed: American College of Chest Physicians Evidence-Based Clinical Practice Guidelines. Chest. 2012; 141(2 Suppl): e351s-418s. 3. White RH. The epidemiology of venous thromboembolism. Circulation. 2003; 107(23 Suppl 1): I Heit JA, Silverstein MD, Mohr DN, Petterson TM, O Fallon WM, Melton LJ, 3rd. Predictors of survival after deep vein thrombosis and pulmonary embolism: a population-based, cohort study. Archives of Internal Medicine. 1999; 159(5): Beckman MG, Hooper WC, Critchley SE, Ortel TL. Venous thromboembolism: a public health concern. American Journal of Preventive Medicine. 2010; 38(4 Suppl): S Tripodi A, Chantarangkul V. Performance of quantitative D-dimer methods: results of the Italian external quality assessment scheme. Journal of Thrombosis and Haemostasis. 2007; 5(1): Performance of LABGEO IB D-dimer Test Ghys T, Achtergael W, Verschraegen I, Leus B, Jochmans K. Diagnostic accuracy of the Triage D-dimer test for exclusion of venous thromboembolism in outpatients. Thrombosis Research. 2008; 121(6): Legnani C, Fariselli S, Cini M, Oca G, Abate C, Palareti G. A new rapid bedside assay for quantitative testing of D-Dimer (Cardiac D-Dimer) in the diagnostic work-up for deep vein thrombosis. Thrombosis Research. 2003; 111(3): Sidelmann JJ, Gram J, Larsen A, Overgaard K, Jespersen J. Analytical and clinical validation of a new point-of-care testing system for determination of D-Dimer in human blood. Thrombosis Research. 2010; 126(6): Dempfle CE, Zips S, Ergul H, Heene DL. The Fibrin Assay Comparison Trial (FACT): evaluation of 23 quantitative D-dimer assays as basis for the development of D-dimer calibrators. FACT study group. Thrombosis and Haemostasis. 2001; 85(4): Meijer P, Haverkate F, Kluft C, de Moerloose P, Verbruggen B, Spannagl M. A model for the harmonisation of test results of different quantitative D-dimer methods. Thrombosis and Haemostasis. 2006; 95(3): Kelly J, Hunt BJ. A clinical probability assessment and D-dimer measurement should be the initial step in the investigation of suspected venous thromboembolism. Chest. 2003; 124(3): Wells PS, Anderson DR, Rodger M, Forgie M, Kearon C, Dreyer J, Kovacs G, Mitchell M, Lewandowski B, Kovacs MJ. Evaluation of D-dimer in the diagnosis of suspected deep-vein thrombosis. The New England Journal of Medicine. 2003; 349(13): Perrier A, Roy PM, Aujesky D, Chagnon I, Howarth N, Gourdier AL, Leftheriotis G, Barghouth G, Cornuz J, Hayoz D, Bounameaux H. Diagnosing pulmonary embolism in outpatients with clinical assessment, D-dimer measurement, venous ultrasound, and helical computed tomography: a multicenter management study. The American Journal of Medicine. 2004; 116(5): Geersing GJ, Erkens PM, Lucassen WA, Buller HR, Cate HT, Hoes AW, Moons KG, Prins MH, Oudega R, van Weert HC, Stoffers HE. Safe exclusion of pulmonary embolism using the Wells rule and qualitative D-dimer testing in primary care: prospective cohort study. BMJ (Clinical research ed). 2012; 345: e Geersing GJ, Toll DB, Janssen KJ, Oudega R, Blikman MJ, Wijland R, de Vooght KM, Hoes AW, Moons KG. Diagnostic accuracy and user-friendliness of 5 point-of-care D-dimer tests for the exclusion of deep vein thrombosis. Clinical Chemistry. 2010; 56(11): Haeberle S, Brenner T, Zengerle R, Ducree J. Centrifugal extraction of plasma from whole blood on a rotating disk. Lab on a chip. 2006; 6(6): Amasia M, Madou M. Large-volume centrifugal microfluidic device for blood plasma separation. Bioanalysis. 2010; 2(10): Kersaudy-Kerhoas M, Sollier E. Micro-scale blood plasma separation: from acoustophoresis to egg-beaters. Lab on a chip. 2013; 13(17): Fukuda T, Kasai H, Kusano T, Shimazu C, Kawasugi K, Miyazawa Y. A rapid and quantitative D-Dimer assay in whole blood and plasma on the point-of-care PATHFAST analyzer. Thrombosis Research. 2007; 120(5): Deitelzweig SB, Johnson BH, Lin J, Schulman KL. Prevalence of clinical venous thromboembolism in the USA: current trendsand future projections. AmericanJournal of Hematology. 2011; 86(2):