STA Liatest Research Articles

Persisting Endothelial Cell Activation and Hypercoagulability after Recovering from COVID-19: The Roadmap-Post COVID-19 Study

Background: In patients with severe COVID-19, activation of blood coagulation and endothelial cells orchestrated with complement activation and cytokine storm leads to disease worsening and death. Hypercoagulable state and endothelial cell activation and fibrinolytic unbalance are common alterations in patients with COVID-19 hospitalized either at the conventional medical ward or at the intensive care unit (ICU). Two systematic reviews highlight that D-dimer values are higher in non survivors as well as in patients with severe COVID-19 than in those with mild disease, showing the association between D-dimer levels and disease severity or death. Nevertheless, there are limited data in small series of patient, on the persistence of hypercoagulability and endothelial cell activation following recovery from COVID-19. Aims: To investigate the persistence of endothelial cell activation and hypercoagulability after recovery from COVID-19. Patients/Methods. COVID-19 survivors (n = 208) and 30 healthy individuals were enrolled in this study. Methods: In ,total, 208 COVID-19 survivors were included in the study; 125 males (60%) and 83 females (40%) with a median age of 50 years (IQR = 20, range 18 to 78 years). Biomarkers of hypercoagulability and endothelial cell activation: The procoagulant phospholipid-dependent clotting time (PPL-ct) was measured using a factor Xa-based clotting assay STA®-Procoag-PPL. Free-TFPI levels were measured with the ELISA Free TFPI, Asserachrom®; D-dimer and fibrin monomers (FM) were measured by STA Liatest D-Di and STA Liatest FM; soluble thrombomodulin (sTM) was measured by Asserachrom® TM. All assays were from Diagnostica Stago, Asnières, France. Heparanase-levels were measured with ELISA kits R&D. Systems (Lille France). Detection of anti-SARS-CoV2 antibodies: IgG and IgA anti-SARS-CoV-2 antibodies were detected in the sera of the survivors using a semi-quantitative commercial ELISA methodology (Euroimmun Medizinische Labordiagnostika AG, Lubeck, Germany) Results: The median interval between symptom onset and screening for SARS-CoV-2 antibodies was 62 days (IQR = 22 days). Antibodies against SARS-CoV-2 were detected in all survivors enrolled in the study. The median level of IgG and IgA antibodies was 5.68 U (IQR = 6.19) and 3.62 U (IQR = 5.36), respectively. In 112 survivors with known blood group, 48 (43%) were group O, 44 (39%) were group A, 14 (13%) were group B, and 6 (5.4%) were group AB. Survivors showed significantly higher levels of D-Dimers, FM, TFPI, and heparanase as compared to that of the control group. Survivors had significantly shorter PPL-ct . One out of four of COVID-19 survivors showed increase at least one biomarker of endothelial cell activation or hypercoagulability (Table1). Elevated D-dimer was associated with older age. Elevated FM was associated with female gender. Elevated heparanase was independently associated with male gender. Decreased Procoag-PPL clotting time was associated with female gender. Univariate and multivariate logistic regression analysis examining predictors of hypercoagulability in study cohort of COVID-19 survivors are reported in table 2 . The interval between COVID-19 diagnosis and assessment was not associated with the changes of the studied biomarkers. Conclusion: The ROADMAP-postCOVID-19 study offers an analysis of a large panel of biomarkers of endothelial cell activation and hypercoagulability in survivors of COVID-19. The study documents that two months after symptoms' onset, activation of endothelial cells, in vivo thrombin generation, and fibrin lysis are frequent phenomena up to two months from COVID-19 symptom onset. The data presented herein allow to propose that evaluation of blood hypercoagulability (using D-dimer for instance) and endothelial cell activation could offer the possibility of prompt identification of COVID-19 survivors at risk of post-COVID-19 vascular complications. The findings of the present study underline the need for a thorough evaluation of a potential correlation between the sustained hypercoagulability and endothelial cell activation with the symptomatology of long-COVID-19 to apply anticipated diagnostic and therapeutic strategies. The clinical relevance of the identified biomarkers of endothelial activation and hypercoagulability in the assessment of the risk of long COVID-19 must be investigated in a prospective study. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Profile of Perioperative Coagulation in Major Liver Resection for Cancer: A Prospective Observational Study

Background: The liver plays a key role in the complex and dynamic process of blood coagulation. Consequently, after liver resection the equilibrium between procoagulant and anticoagulant activity is often deranged. Major hepatectomies have been associated with excessive intraoperative blood loss, hemodynamic instability, massive fluid shifts, and coagulopathy. Alterations of the coagulation profile are always present post-operatively even in case of normal pre-operative coagulation function. Cumulative evidence supports the superiority of global coagulation analysis compared with conventional coagulation for clinical decision-making. Aim: The purpose of this study was to assess the perioperative coagulation status of patients undergoing major hepatectomies for primary or metastatic hepatic malignancy. Conventional coagulation tests, plasma levels of coagulation factors, and TGA measurements were used for longitudinal documentation of the coagulation activity. Methods: Patients included in the study were referred for major liver resection which was defined as a resection of three or more liver segments, and were followed up to the 10th post-operative day by serial measurements of conventional coagulation tests, plasma levels of coagulation factors, and thrombin generation assay parameters. Prothrombin time (PT) and activated partial thromboplastin time (aPTT) were determined by STA Neoplastin R, and STA-Cephascreen, Fibrinogen by STA-Liquid Fib, D-dimer STA Liatest D-DI Plus, all from Diagnostica Stago, France. Coagulation factor activity for FII, FV, FVII, FVIII, FIX, FX, was measured by clotting assay using coagulation factor deficient plasma, vWFAg , and vWFAc by vWFAg and Innovance vWFAc, from Siemens Healthcare Diagnostics, Germany. Thrombin generation (TG) was assessed in samples of PPP with the TF 5 pM PPP-Reagent on Calibrated Automated Thrombogram (Stago, France). Results: Changes over time for FII, FV, FVII and FX were statistically significant (p <0.05). Factor VIII level was above normal values at baseline. A steady rise was observed for fibrinogen, FVIII, and vWF antigen throughout the study period, but only vWF antigen exhibited statistically significant changes over time. Levels of coagulation inhibitors, AT, PC and PS, decreased significantly during the first postoperative day and then increased reaching baseline values by POD5 for protein C and S, but AT, remained low by POD10. INR increased after surgery, reaching a peak on the first post-operative day and decreasing toward baseline levels from then on (p <0.001). The aPTT remained within normal range throughout the study period (Table 1). TG conducted at baseline, on POD1, POD5, and POD10 showed elevated of Lag time and time to peak at baseline, whereas the rest thrombin generation parameters were within reference range. ETP decreased significantly over time, reaching lower values by POD5 and sustaining those low values by POD10. Peak height fluctuated in a similar manner, but changes over time did not bear any statistical significance. Lag-time, which was prolonged at baseline, exhibited a steady rise throughout the study period, bearing borderline statistical significance (Table 2) . A statistically significant positive correlation was detected between AT and time to Peak (ttPeak) at baseline, however any other significant difference between coagulation factors, natural coagulation inhibitors, and TGA parameters was observed. Conclusion: This study shows that patients with hepatic malignancy have systematic endothelial activation and hyper-coagulability as indicated by increased FVIII levels and lag time prolongation in thrombin generation. These patients are in a hyper-coagulable state in the early post-operative period, as shown by increased vWF, FVIII, D-dimer, fibrinogen and decreased levels of natural coagulation inhibitors AT, PC, and PS. Measurements of INR, PT, aPTT are not useful in this setting and could be misleading. Global coagulation analysis may be more appropriate for clinical guidance. To the best of our knowledge this is the first report in literature documenting the pre-existing coagulation disturbances in patients with hepatic carcinoma and their impact on the coagulation status after major hepatectomy. However, the clinical implications of our findings deserve further investigation. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Reporting of D-Dimer and performance of analytical systems: Data from participants to the External Quality Assessment Scheme of the Biomedical Research Centre in Italy

Introduction: D-Dimer assessment represents a cornerstone in the diagnostic approach to several thrombotic disorders. Recent literature has highlighted the role of D-Dimer also in the diagnostic pathway of coronavirus infection (COVID-19) and the importance of harmonized reporting [D-dimer unit (DDU) or fibrinogen equivalent unit (FEU);unit of measure;cut-off] in order to guarantee the correct interpretation of the results. Methods: D-Dimer data from 100 EQA participants and the inter-laboratory variability (CV%) of the last 7 years for the most used analytical systems: Werfen, HemosIL HS;Werfen, HemosIL HS-500;Auto D-D, Sclavo;Innovance, Siemens;VIDAS, bioMerieux and STA Liatest, Stago have been evaluated. Results: Concerning the results expression in DDU or FEU, we observed a prevalence of FEU (55.1%) over DDU (44.9%);the value was confirmed in the last 7 years (average FEU = 55.6%), differently from data obtained in the survey conducted in 2014 at a national level. The units used are: ng/mL (67.8%), μg/L (29.0%) and mg/L (3.2%) for D-Dimer DDU;ng/mL (57.9%), μg/mL (21.0%), μg/L (15.8%) and mg/L (5.3%) for D-Dimer FEU. Inter-laboratory variability (mean CV%) calculated on a total of 72 controls is lower for all diagnostic systems at pathological levels than the one observed for concentrations below the cut-off. Discussion: This study demonstrates that the reporting of D-Dimer results does not comply with the 2014 SIBioC consensus document which recommended the use of μg/L FEU, and highlights 8 different types of information. Data reported in this study call for the harmonization of D-Dimer reporting in order to guarantee the correct interpretation of the information, both in the case of COVID-19 and in all the diseases already known where this analyte has a clinical relevance.

ABO blood group, glycosyltransferase activity and risk of venous thromboembolism

IntroductionABO blood group influence the risk of venous thromboembolism (VTE) by modifying A and B glycosyltransferases (AGT and BGT) activities that further modulates Factor VIII (FVIII) and von Willebrand Factor (VWF) plasma levels. The aim of this work was to evaluate the association of plasma GTs activities with VWF/FVIII plasma levels and VTE risk in a case-control study. Materials and methods420 cases were matched with 420 controls for age and ABO blood group. GT activities in plasma were measured using the quantitative transfer of tritiated N-acetylgalactosamine or galactose to the 2′-fucosyl-lactose and expressed in disintegration per minute/30 μL of plasma and 2 h of reaction (dpm/30 μL/2H). FVIII and VWF plasma levels were respectively measured using human FVIII-deficient plasma in a 1-stage factor assay and STA LIATEST VWF (Diagnostica Stago). ResultsA and B GT activities were significantly lower in cases than in controls (8119 ± 4027 vs 9682 ± 4177 dpm/30 μL/2H, p = 2.03 × 10−5, and 4931 ± 2305 vs 5524 ± 2096 dpm/30 μL/2H, p=0.043 respectively). This association was observed whatever the ABO blood groups. The ABO A1 blood group was found to explain~80% of AGT activity. After adjusting for ABO blood groups, AGT activity was not correlated to VWF/FVIII plasma levels. Conversely, there was a moderate correlation (ρ ~ 0.30) between BGT activity and VWF/ FVIII plasma levels in B blood group carriers. ConclusionWork showed, for the first time, that GT activities were decreased in VTE patients in comparison to controls with the same ABO blood group. The biological mechanisms responsible for this association remained to be determined.

Factor VIII Levels in Blood Group O and Non-O in Patients with Non-Severe Haemophilia a

Introduction ABO blood group affects levels of von Willebrand Factor (VWF) and Factor (F) VIII, as individuals with blood group non-O have higher levels of VWF and FVIII. To the best of our knowledge there is no data available regarding the association between blood group and FVIII in non-severe haemophilia. Aim We aimed to explore the influence of ABO blood group on VWF and FVIII levels in patients with non-severe haemophilia A (HA). Methods We recruited persons with non-severe HA (FVIII levels of 1-40%). Adult patients (age ≥ 18 years), with platelet count &gt; 105 /µl, without impaired renal and hepatic function, active cancer, surgery within the last 6 weeks, or overt infection within the last 2 weeks, who gave their informed consent were included. A blood sample was collected for laboratory analysis. The diagnosis of haemophilia was confirmed by molecular analysis of the FVIII gene in all patients. None of the patients had inhibitors against FVIII at the time of sample collection. The lowest FVIII level ever measured in each patient's history served as basis for the assessment of severity. Healthy male persons with a median age comparable to the patient group served as controls. We determined the following parameters: ABO blood group, FVIII, VWF activity (VWF:Act), and VWF antigen (VWF:Ag). FVIII was measured with a one-stage assay (Sysmex-CA7000 - Sysmex, Kobe, Japan, in multi-dilution mode with native FVIII-deficient plasma - Technoclone, Vienna, Austria and Aktin FS activator - Siemens, Marburg, Germany), VWF:Act (BC von Willebrand Reagent or Innovance VWF Ac - Siemens Healthcare, Marburg, Germany) and VWF:Ag levels with a latex agglutination assay (STA LIATEST VWF, Diagnostica Stago). Comparison of VWF and FVIII levels between blood group O and non-O was calculated with Mann-Whitney-U test, correlation between FVIII and VWF levels was calculated with Spearman's correlation, association of FVIII and VWF separated for blood group was calculated with univariable linear regression. Association of FVIII with VWF and blood group was calculated using multivariable linear regression. All calculations were performed with SPSS (IBM Version 25.0). Results A total of 89 persons with HA (71 with mild and 18 with moderate haemophilia) and 82 healthy controls were included. Patient characteristics are listed in Table 1. Median levels of VWF were significantly higher both in patients (p=0.002 for both VWF:Ag and VWF:Act) and healthy controls (p&lt;0.001 for both VWF:Ag and VWF:Act) with blood group non-O compared to individuals with blood group O. FVIII levels were not significantly different between blood group non-O and O in HA patients (15% vs. 14.1%, p=0.716) but significantly higher in healthy controls with blood group non-O versus O (150.0% vs. 109.5%, p&lt;0.001; Table 2). In HA there was no correlation between FVIII and VWF:Act (rho=0.180, p=0.095) or between FVIII and VWF:Ag (rho=0.028, p=0.795, Table 3). In univariable linear regression there was no significant association between VWF and FVIII in HA, neither in those with blood group non-O nor in those with blood group O (Figure 1). Also in multivariable linear regression there was no significant association between VWF (p=0.632) and blood group (p=0.929) with FVIII in patients with HA. In healthy controls, there was a strong positive correlation between FVIII and VWF:Act (rho=0.751, p&lt;0.001) and between FVIII and VWF:Ag (rho=0.790, p&lt;0.001) (Table 3). In the healthy controls with blood group non-O 1% elevation in the VWF:Ag was associated with 0.63% of elevation in the FVIII:Act and in blood group O 1% elevation in the VWF:Ag was associated with 0.66% of elevation in the FVIII:Act (Figure 2). Conclusions Neither the blood group nor the VWF had an influence on FVIII levels in non-severe HA patients. We conclude that the impact of the genetic mutation of the FVIII gene by far outweighs the influence of VWF levels and the blood group. Thus, for the diagnosis of HA and for determination of HA severity, the blood group needs not to be taken into account. Disclosures Pabinger: Roche: Honoraria; Sobi: Research Funding; Novo Nordisk: Research Funding; CSL Behring: Research Funding; Pfizer: Honoraria; Shire: Honoraria; Bayer: Honoraria; Sobi: Honoraria.

Concentration of fibrin monomer in pregnancy and during the postpartum period.

Background D-dimer increases during pregnancy and is problematic to use in the diagnosis of venous thromboembolism. Fibrin monomer represents an alternative biomarker for venous thromboembolism. However, to be useful in pregnancy, the fibrin monomer concentration should be stable throughout pregnancy and during postpartum. Methods To describe the course of fibrin monomer concentration during pregnancy and the postpartum period in healthy pregnant women and to compare their within-subject biological variation (CVI) with non-pregnant women. Blood samples were obtained every fourth week during pregnancy and three samples after delivery in 20 healthy women and every fourth week during a 40-week period in 19 healthy non-pregnant women. Fibrin monomer (STA Liatest FM, Stago) was analysed in duplicates for all samples. Concentrations of fibrin monomer in pregnant and non-pregnant women were compared and the CVI for fibrin monomer was calculated. Results The median fibrin monomer concentration in pregnant women was 6.2 mg/L (2.5 and 97.5 percentiles 3.7–10.8 mg/L) and in non-pregnant women 4.8 mg/L (3.6–8.2) ( P &lt; 0.01). The fibrin monomer concentration was relatively stable during pregnancy, although a few unexplained high fibrin monomer concentrations were found during pregnancy/postpartum. Fibrin monomer CVI in pregnancy and postpartum was 20.6% (95% CI 18.3, 23.5) and in non-pregnant 16.1% (13.7, 18.9). Conclusions For clinical purposes, fibrin monomer concentration can be considered stable during pregnancy, although it is slightly higher than in non-pregnant women. Pregnant and non-pregnant women have the same CVI. The suitability of fibrin monomer in venous thromboembolism diagnosis in pregnant women should be validated in further studies.

D-dimer, FVIII and thrombotic burden in the acute phase of deep vein thrombosis in relation to the risk of post-thrombotic syndrome

BackgroundPost-thrombotic syndrome (PTS) is the most common complication of deep vein thrombosis (DVT), but few data are available on the risk factors for PTS. AimsTo assess whether the time-course of D-dimer, FVIII, and thrombotic burden are related to PTS development. MethodsPatients (n=59) with proximal DVT of the lower limbs (age 64; range:20-88years; male 56%) were enrolled on the day of diagnosis (D0) and all received heparin for 5-7days, overlapped and followed by vitamin K antagonists (VKA) for 3months. Whole-leg compression ultrasound examination was conducted on D0 and 7 (D7), 30 (D30), and 90 (D90) days afterwards, when blood samples were also taken for D-dimer (STA Liatest) and FVIII (chromogenic assay) testing. Thrombotic burden was defined at each time point according to a score, which considered thrombosis extent and occlusion degree. Villalta score was evaluated at D30, D90, and D180. ResultsAt D90, 12 patients developed PTS (Villalta score ≥5) and the median Villalta score was 1 (IQR 0.3-3.0) and was not correlated with either D-dimer or FVIIII time course. At D180, 13 patients had PTS and they had similar thrombotic score at D0, D30 to those without PTS, but higher at D90 (7.6±5.1 vs. 3.2±3.6; p=0.011). Thrombotic score at D90 was correlated with Villalta score at D90 (rho=0.374, p=0.009) and at D180 (rho=0.436, p=0.006). ConclusionsThrombotic burden after 90days of VKA is correlated with PTS.

Extension of dynamic range of sensitive nanoparticle-based immunoassays

Nanoparticles have successfully been employed in immunometric assays that require high sensitivity. Certain analytes, however, require dynamic ranges (DRs) around a predetermined cut-off value. Here, we have studied the effects that antibody orientation and addition of free solid-phase and detection antibodies have on assay sensitivity and DR in traditional sandwich-type immunoassays. D-dimer and cardiac troponin I (cTnI), both routinely used in critical care testing, were applied as model analytes. The assays were performed in microtitration wells with preimmobilized solid-phase antibody. Inherently fluorescent nanoparticles coated with second antibody were used to detect the analyte. The selection of antibody orientation and addition of free solid-phase or detection antibody, with nanoparticles and calibrator, desensitized the assays and extended the DR. With D-dimer the upper limit of the DR was improved from 50 to 10,000ng/ml, and with cTnI from 25 to 1000ng/ml. Regression analysis with the Stago STA Liatest D-dimer assay yielded a slope (95% confidence interval) of 0.09 (0.07–0.11) and a y-intercept of −7.79 (−17.87–2.29)ng/L (n=65, r=0.906). Thus it is concluded that Europium(III)-chelate-doped nanoparticles can also be employed in immunoassays that require wide DRs around a certain cut-off limit.

Age- and sex-dependent reference intervals for D-dimer: Evidence for a marked increase by age

A low D-dimer is commonly used to exclude venous thromboembolism in low risk patients. However, the reference intervals are poorly defined and D-dimer has been shown to increase by patient age. We aimed to establish age- and sex-dependent D-dimer reference intervals and to test the consequence of different cut-off limits. By means of the STA-Lia test D-dimer was measured in 1,352 subjects aged between 23 and 93years from the general population. A further 94 subjects were measured with the HemosIL test. The reference intervals were age-dependent with the upper 95% limit increasing from 0.92 FEU mg/L in the age group <50years to 2.39 FEU mg/L in the group ≥70years. Minor differences in the reference intervals between sexes were found. The commonly used cut-off at 0.5 FEU mg/L resulted in a decrease of negatives from 91% in <50years to 56 % in ≥70years. The age-dependent cut-off value (patient's age*0.01 FEU mg/L in 50+ years) resulted in an increase of negatives by age. The linear increase in D-dimer has a slope at approximately 0.0077*age, meaning that a cut-off at (patient's age*0.0077) FEU mg/L, gives an almost constant number of positives across age groups. The two assays showed a poor correlation and a higher percentage of positives using the STA Lia test. The age-dependent increase in D-dimer seen in patients was confirmed, which questions the clinical value of the present cut-off for venous thromboembolism.

D-dimer testing during anticoagulant therapy should be used to indicate patients who need extended anticoagulant therapy

The prolong study showed that pts with venous thromboembolism (VTE) who had elevated D-dimer level after discontinuation of anticoagulant therapy benefit from resumption of anticoagulant therapy. It is not yet clear should D-dimer testing during anticoagulant therapy be used to indicate VTE pts who need extended anticoagulant therapy. Purpose: To estimate should D-dimer testing during anticoagulant therapy be used to indicate VTE pts who need extended anticoagulant therapy. Methods: One hundred and eleven pts (76 men) in the age 18-76 (mean 54±14) years with deep vein thrombosis (DVT) and/or pulmonary embolism (PE) were included in the study. Pts received unfractionated or low molecular weight heparin for at least 5 days followed by long-term warfarin therapy (international normalized ratio 2,0-3,0). D-dimer testing was performed at baseline and after 1, 3, 6 and 12 months of anticoagulant therapy by a quantitative assay with reagent kits "STA LIATEST D-DI" (Diagnostica Stago) with estimated cut-off level of 0,5 ug/ml. The follow-up period was 12 months. Endpoints were DVT/PE recurrences. Results: At least one episode of elevated D-dimer level during 12 months of anticoagulant therapy was detected in 22% of VTE pts. The frequency of DVT recurrences during 12 months was 11%. There were no PE recurrences. In pts with DVT recurrences, all episodes of elevated D-dimer level preceded DVT recurrences and were detected before DVT recurrences. The frequency of DVT recurrences in pts with elevated D-dimer level during anticoagulant therapy was significantly higher than in pts with normal D-dimer level – 27% vs 7,7% (p=0,022). Cox proportional hazard model showed that any episode of elevated D-dimer level during anticoagulant therapy is associated with a 4-fold increase of the risk of DVT recurrence (HR 4,43; 95% CI 1,42-13,78; p=0,01) compared in pts with normal D-dimer level during anticoagulant therapy. Conclusions: Our pilot study showed that any episode of elevated D-dimer level during 12 months of anticoagulant therapy is associated with a 4-fold increase of the risk of DVT recurrences. We suggest that VTE pts who had at least one episode of elevated D-dimer level during 12 months of anticoagulant therapy need extended anticoagulant therapy. Thus, D-dimer may be one of criteria for determination of duration of anticoagulant therapy. D-dimer testing during anticoagulant therapy also should be used to predict the risk DVT recurrences in VTE pts.

The Wells rule and D‐dimer for the diagnosis of isolated distal deep vein thrombosis

Pretest clinical probability with the Wells rule and D-dimer have been widely investigated for the diagnosis of symptomatic proximal deep vein thrombosis (DVT) of the lower limbs, but they have not been formally tested for symptomatic isolated distal DVT diagnosis. To evaluate the diagnostic accuracy of the Wells rule and D-dimer for isolated distal DVT. This was a single-center, cross-sectional study including 873 consecutive outpatients with suspected DVT, in whom pretest clinical probability determination, D-dimer determination (STA Liatest; cut-off of < 500 ng mL(-1) ) and complete compression ultrasonography of both lower limbs were performed. The isolated distal DVT prevalence was 12.4% (90/725). The sensitivity of the Wells rule for isolated distal DVT was 47% (95% confidence interval [CI] 36-57%), the specificity was 74% (95% CI 70-77%), and the negative and positive predictive values were 91% (95% CI 88-93%) and 20% (95% CI 15-26%), respectively. Patients with isolated distal DVT had higher D-dimer levels than patients without DVT (1759 ± 1576 vs. 862 ± 1079 ng mL(-1) , P = 0.0001). D-dimer was negative in 13 patients with isolated distal DVT. D-dimer sensitivity and specificity for isolated distal DVT were 84% (95% CI 75-91%) and 50% (95% CI 46-54%), respectively, with a negative predictive value of 96% (95% CI 93-98%). In patients with low pretest clinical probability, the D-dimer negative predictive value was 99% (95% CI 95-100%). In clinically suspected DVT with negative proximal compression ultrasonography, pretest clinical probability with the Wells rule has a low diagnostic accuracy for isolated distal DVT. D-dimer has a better negative predictive value, but alone it does not exclude isolated distal DVT. In patients with low pretest clinical probability, D-dimer had a negative predictive value of > 95% for isolated distal DVT.

Age and gender specific cut-off values to improve the performance of d-dimer assays to predict the risk of venous thromboembolism recurrence

The Prolong study shows that continuing vitamin K antagonists (VKA) in patients with abnormal D-dimer (evaluated by a qualitative assay, Clearview Simplify D-dimer) results in a significant reduction of venous thromboembolism (VTE) recurrence. The present study retrospectively analyzes a subgroup of patients enrolled in the Prolong study with a view to calculate cut-off values for six quantitative D-dimer methods to predict the risk of VTE recurrence. We measured D-dimer levels by VIDAS D-dimer Exclusion (bioMerieux), STA Liatest D-dimer (DiagnosticaStago), HemosIL D-dimer and HemosIL D-dimer HS (Instrumentation Laboratory), Innovance D-dimer (Siemens) and AutoDimer (Trinity Biotech) in frozen plasma aliquots sampled 30±10days after VKA cessation in 390 patients enrolled in the Prolong study. During follow-up (562.7years), 28 patients had recurrent VTE (7.2%, 5.0% person-years). Since D-dimer levels are positively correlated with age and significantly lower in men, we calculated method-specific cut-off values according to age and gender. The HRs for VTE recurrence calculated using method-specific cut-off values based on age and gender are higher than those using cut-off values indicated by the manufacturers for VTE exclusion in symptomatic outpatients. These data suggest that method-specific cut-off values calculated according to patient age and gender can be more accurate in identifying patients at a higher risk for VTE recurrence. These method-specific cut-off values are being evaluated in the ongoing prospective management multicenter DULCIS study.

A Randomized, Controlled Trial of Varying D-Dimer Cutpoints According to Clinical Pretest Probability Compared with a Uniform D-Dimer Cutpoint for Diagnosis of Deep Vein Thrombosis

AbstractAbstract 5116 Background:D-dimer assays are currently used as a one-size-fits-all diagnostic test for DVT where the same cutpoint is used to define a positive result for all patients. This approach ignores the relationship between the diagnostic properties of the assay and the pretest probability of the disease. Objective:We conducted a randomized, allocation concealed, controlled, assessor-blinded trial to compare the safety and efficiency of two diagnostic strategies for D-dimer testing for first acute DVT of the lower limb: Selective use of D-dimer testing, using different D-dimer cutpoint levels according to clinical pretest probability (C-PTP) and the traditional Uniform approach. Method:1723 consecutive eligible inpatients and outpatients at 5 clinical centres (4 in Ontario, 1 in Montreal) had their C-PTP assessed using the Wells' Score and were randomized to either: Uniform D-dimer testing, in which all patients had a D-dimer test and a level of <0.5 μ g fibrinogen equivalent units (FEU)/mL was classified as negative (the control group); or Selective D-dimer testing, in which D-dimer testing was (a) performed in patients with Low and Moderate C-PTP only, (patients with a High C-PTP had an ultrasound, but no D-dimer testing), and (b) was classified as negative if the D-dimer level was <0.5 μ g FEU/mL with Moderate C-PTP or <1.0 μ g FEU/mL with Low C-PTP (the experimental group). The D-dimer assays used in this study were the MDA D-Dimer (bioMerieux, Inc and Trinity Biotech) and STA Liatest D-Dimer(Diagnostica Stago). All patients with a positive D-dimer had an ultrasound of their leg. Symptomatic venous thromboembolic events (VTE) during 3 months of follow-up were objectively confirmed and independently adjudicated. Results:The incidence of VTE during 3 months of follow-up in patients who were not diagnosed with DVT at initial presentation was 1.1% (9/811) with Uniform testing, and 1.1% (9/814) with Selective testing (difference 0.004%, 95% CI:-1.122, 1.111). The incidence of VTE during 3 months of follow-up in patients in the Low C-PTP Select arm (D-dimer cutpoint 1.0 μ g FEU/mL) was 0% (0/356), and in the Low C-PTP Uniform arm (D-dimer cutpoint 0.5 μ g FEU/mL), it was 0.3% (1/336). Conclusion:This is the first randomized controlled clinical trial to show that Selective use of D-dimer testing, with a higher D-dimer cutpoint in patients with Low C-PTP, is safe and potentially more efficient than current practice. Funded by the Heart & Stroke Foundation of Ontario. Disclosures:Linkins:Trinity Biotech: in-kind donation of D-dimer kits; BioMerieux: in-kind donation of D-dimer kits; Stago Diagnostica: in-kind donation of D-dimer kits. Bates:bioMerieux: in-kind donation of D-dimer kits; Trinity Biotech: Consultancy, in-kind donation of D-dimer kits. Kearon:BioMerieux: in-kind donation of D-dimer kits; Trinity Biotech: in-kind donation of D-dimer kits; Stago Diagnostica: in-kind donation of D-dimer kits.

An age‐adapted approach for the use of D‐dimers in the exclusion of deep venous thrombosis

A normal D-dimer (DD) concentration for the exclusion of deep venous thrombosis (DVT) has a low specificity in older patients and compression ultrasonography is often required. Three D-dimer assays, STA Liatest, Tina-quant, and Innovance, are evaluated in symptomatic outpatients suspected for DVT with emphasis on its performance in older patients by using different cut-off levels. This study includes 466 outpatients suspected for having DVT. The diagnostic accuracy, measured as sensitivity and area under the curve of the receiver operation characteristic curve is good for all DD assays. The specificity of the DD assays combined with a low pretest probability varies from 42.6 to 51.5%. The specificity of the three DD assays in patients > or = 60 years varies, however, between 24.6 and 40.9%. Several cut-off values in different age-subgroups are studied. For patients < 60 years, the most accurate cut-off value is 500 microg/L for all DD assays. For patients > or = 60 years, a threshold of 750 microg/L has the best results with NPV of 100% for all assays and specificity of 48.5% (STA Liatest), 60.6% (Tina-quant), and 49.2% (Innovance), respectively. For the three assays, the number needed to test (NNT) decreases in both subgroups of patients compared to the standard algorithm. A cut-off level of 750 microg/L for patients > or = 60 years improves the clinical performance of DD assays in combination with the PTP score without the loss of NPV. The NNT improves substantially with an age-adapted algorithm.

Reply Re: A Prospective Evaluation of a Quantitative D-dimer Assay in the Evaluation of Acute Pulmonary Embolism

Reply Re: A Prospective Evaluation of a Quantitative D-dimer Assay in the Evaluation of Acute Pulmonary Embolism

Incohérence entre deux méthodes de dosage des D-dimères : un cas d’interférence d’anticorps humain anti-souris

Quantification of D-dimers is the major biometry step in the diagnostic of an episode of the venous thromboembolic disease. The measurement of D-dimers can be performed with ELISA or immunoturbidimetric methods suited to emergency, using a mouse monoclonal antibody as capture and/or revelation antibody. Therefore, the presence in patient's plasma of human antibody mouse (HAMA) that binds the mouse antiglobulin used in immunoassays can lead to false negative or false positive results. In a young woman presenting repetitive thoracic pain suggestive of a pulmonary embolism, a major discrepancy was found between one result of D-dimers above the cut-off with an immunoturbidimetric method (STA Liatest D-DI; Diagnostica Stago) and one result below the cut-off with a sandwich method (Vidas D-Dimer Exclusion; bioMérieux). HAMA, which is known to be responsible for this type of discrepancy, was detected in the patient serum. The false positive result probably impaired with the management of patient. Taking in charge the patient should take into account the possible presence of this antibody. Interference by heterophilic antibodies is not easily detected by the laboratory. Even if their frequency is low, it remains a difficult problem for the biologist. Suspicion generally arises from inconsistency between the clinical data and immunoassay results. A good communication between physician and biologist should avoid to providing false negative or positive results.

Different cut-off values of quantitative D-dimer methods to predict the risk of venous thromboembolism recurrence: a post-hoc analysis of the PROLONG study

The PROLONG study showed that patients with venous thromboembolism who had qualitatively abnormal results in a D-dimer assay (Clearview Simplify D-dimer) after discontinuation of vitamin K antagonism benefit from resumption of treatment with vitamin K antagonism. The objective of this study was to evaluate the possible advantage of using quantitative D-dimer assays. VIDAS D-dimer Exclusion (bioMerieux), Innovance D-DIMER (Dade Behring), HemosIL D-dimer HS (Instrumentation Laboratory) and STA Liatest D-dimer (Diagnostica Stago) assays were performed in plasma aliquots sampled 30+/-10 days after cessation of vitamin K antagonism in 321 patients enrolled in the PROLONG study. During the follow-up without vitamin K antagonism, 25 patients had recurrent venous thromboembolism. The cut-off levels of the quantitative assays giving results most comparable with those of the qualitative test were: VIDAS = 800 ng/mL; Innovance = 800 ng/mL; HemosIL HS = 300 ng/mL; STA Liatest = 700 ng/mL. When the effect of the patients' age (< or = 70 vs. >70 years) was analyzed, it was found that only in younger patients was the rate of recurrence of venous thromboembolism significantly higher in patients with abnormal D-dimer levels. However, using the quantitative assays and age-specific cut-off levels it was possible to determine statistically significant hazard ratios also in elderly patients (VIDAS = 600 and 1200 ng/mL, Innovance = 500 and 900 ng/mL, HemosIL HS = 250 and 450 ng/mL, STA Liatest = 700 and 1000 ng/mL, in patients aged < or = 70 and >70 years, respectively). Quantitative D-dimer assays may provide information useful for evaluating the individual risk of recurrent venous thromboembolism. They seem particularly advantageous since they allow the selection of different cut-off levels according to the age or other characteristics of the patients.

Rapid Differential Diagnosis between Subarachnoid Hemorrhage and Traumatic Lumbar Puncture by D-Dimer Assay

Subarachnoid hemorrhage (SAH) and traumatic lumbar puncture (TLP) are both associated with blood-tinged cerebrospinal fluid (CSF). It is often difficult if not impossible to distinguish between SAH and TLP (1)(2) by available CSF tests such as ( a ) measurement of xanthochromia of the supernatant after centrifugation, ( b ) counting of erythrocytes in the first and last tubes of collected CSF, or ( c ) searching of CSF microscopically for crenated erythrocytes (3)(4)(5). We investigated the measurement of D-dimer (DD) in CSF and evaluated DD measurement as a means to distinguish between SAH and TLP. We measured DD in CSF with the coagulation analyzer STA Compact ® by the immunoturbidimetric assay, STA Liatest D-DI® (Roche Diagnostics). Erythrocyte counts were performed by optical microscopy with a Fuchs-Rosenthal …

Diagnostic value of D-dimer in outpatients with suspected deep venous thrombosis receiving oral anticoagulation

D-dimer has proved a useful diagnostic tool for the exclusion of deep venous thrombosis (DVT). The objective of this paper was to evaluate the diagnostic performance of a diagnostic algorithm combining clinical probability and D-dimer in outpatients receiving oral anticoagulant treatment (OAT) similar to those regularly applied to nonanticoagulated individuals. We enrolled 70 outpatients on OAT who presented with clinically suspected DVT; a standard diagnostic algorithm including clinical evaluation using the modified Wells score and a quantitative immunoturbidimetric D-dimer assay (STA Liatest D-Di; Diagnostica Stago, Asniéres sur Seine, France) was used. A 3-month follow-up period was applied for those patients in whom DVT was initially excluded. The prevalence of DVT was 18.5% (13/70); four of the diagnoses were made during the 3-month follow-up period. The sensitivity, specificity and negative predictive value of D-dimer were 69.2% (95 confidence interval, 42.4-87.3), 47.4% (95% confidence interval, 35.0-60.1) and 87.1% (95% confidence interval, 71.1-94.9), respectively. In conclusion, D-dimer is of limited value in outpatients on OAT presenting with clinically suspected DVT and should be omitted in such individuals; these patients should always undergo compression venous ultrasound, and repeat ultrasonography within 1 week might be warranted in cases with an initial negative examination.

D-dimer is not useful for the diagnosis of isolated superficial venous thrombosis

D-dimer is not useful for the diagnosis of isolated superficial venous thrombosis