Multicenter evaluation of a new quantitative highly sensitive D-dimer assay, the Hemosil® D-dimer HS 500, in patients with clinically suspected venous thromboembolism

Introduction

D-dimer testing is widely used in conjunction with clinical pretest probability (PTP) for venous thromboembolism (VTE) exclusion. We report on a multicenter evaluation of a new, automated, latex enhanced turbidimetric immunoassay [HemosIL® D-Dimer HS 500, Instrumentation Laboratory (IL)].

Materials and Methods

747 consecutive outpatients with suspected proximal deep vein thrombosis (DVT, n = 401) or pulmonary embolism (PE, n = 346) were evaluated at four university hospitals in a management study with a 3 month follow-up. Samples were tested at each center using the new D-dimer assay on an automated coagulation analyzer [ACL TOP (IL)], with clinical cut-off for VTE at 500 ng/mL (FEU).

Results

The sensitivity and negative predictive value (NPV) were 100% for all PTP subgroups (no false negative results); for both sensitivity and NPV the lower limit of the 95% CI in patients with moderate/low PTP was higher than 95%. The overall specificity was 45.1% (95%CI: 41.1-49.3%). Higher specificity value was recorded in the low PTP subgroup [49.2% (95%CI: 41.7-56.7)]. No significant differences were found between patients suspected of having DVT or PE; sensitivity and NPV were 100%. The reproducibility of the assay was good, being the total CVs% less than 10% for D-dimer concentration near the clinical cut-off.

Conclusions

The new, highly sensitive D-dimer assay proved to be accurate when used for VTE diagnostic work-up in outpatients. Based on 100% sensitivity and NPV and lower limit of the 95% CI higher than 95%, the assay can be used as a stand-alone test in patients with non high PTP.     

Development of a lateral-flow immunochromatographic test device for the rapid detection of difloxacin residues

In this study, a rapid competitive immunochromatographic test strip has been developed for specifically testing residues of difloxacin (DIF). 1H₁₀-2B₂, one of the three hybridoma lines that were tested and selected by enzyme linked immunosorbent assay, was used in the test strip. The monoclonal antibody has a good sensitivity with an IC₅₀ of 2.2 ng/mL to DIF, 0.24% cross-reactivity towards danofloxacin and no cross-reactivity towards other related compounds and other drugs. The calibration curve of DIF test strip presented a typical sigmoidal curve. In practice, the lower limit of detection using a strip reader was 0.5 ng/ml while 2 ng/ml by unaided visual assessment. Samples of milk were spiked at 2–8 ng/mL, presenting recoveries between 94.5 and 107%, and the coefficient of variation (CV,%) (1.6–9%). The data above suggests that the strip meets the requirements of high sensitivity, good specificity, simplicity and speed, as well as the characteristics of reproducibility and accuracy.     

Human neutrophil lipocalin (HNL) as a biomarker of acute infections

The early and accurate discrimination between bacterial and viral causes of acute infections is the key to a better use of antibiotics and will help slow down the fast-growing resistance to commonly used antibiotics. This discrimination is in the vast majority of cases possible to achieve by blood assay of the biomarker human neutrophil lipocalin (HNL), which we showed to be uniquely increased in patients suffering from bacterial infections. In serum, sensitivities and specificities of >90% are achieved in both adults and children. In order to eliminate the need to produce serum, a whole-blood assay with an assay time of <10?min was developed in which blood neutrophils are activated to release HNL. The diagnostic accuracy of this assay also showed sensitivities and specificities of >90% in most infectious diseases and was clearly superior to contemporary assays such as blood neutrophil counts, C-reactive protein, procalcitonin, and expression of CD64 on blood neutrophils. This format lends itself to the development of a point-of-care HNL assay and will be a major step forward to accomplish the goal of accurately diagnosing patients with symptoms of acute infections within 10?min at the emergency room or at the doctor’s office.     

Automated nephelometric immunoassays with novel shell/core particles

Nephelometry has become a very reliable and convenient method for immunoassays. Recent innovations in polymer chemistry have led to the design of highly sensitive and reproducible immunoassays for the Behring Nephelometer Analyzer. The principle of particle-enhanced immunoassays has led to the development of new polymer particles. Key advances include the preparation and use of particles with polystyrene cores covered by thin shells with chemically reactive groups. The cores are prepared by emulsion polymerization, giving particles with a well-defined size. The particle size has been optimized to fulfill the requirements for nephelometric analysis. The shells consist of a polystyrene/polymethacrylate/polymethacrylamidoacet-aldehyde dipentylacetal copolymer, which allows a covalent binding of the immunochemicals of interest. Using these particles, a method for the determination of C-reactive protein has been worked out.     

Anti-CD36 autoantibodies in thrombotic thrombocytopenic purpura and other thrombotic disorders: identification of an 85 kD form of CD36 as a target antigen

The presence of anti-CD36 antibodies in plasma of patients with thrombotic thrombocytopenic purpura (TTP), idiopathic thrombocytopenic purpura (ITP), and heparin-induced thrombocytopenia without/with thrombosis (HIT/HITT) has been examined by immunoblots, and a monoclonal antibody capture assay, the platelet-associated IgG characterization assay (PAICA). Results with PAICA showed that 73% (8/11) of patients with TTP were positive, and 71% (10/14) by immunoblots. With ITP, 20% (6/30) were positive by PAICA and 19% (3/16) by immunoblots; HIT, 30% (3/10) were positive by PAICA and 60% (6/10) by immunoblot; HITT, 50% (2/4) by PAICA and 100% (4/4) by immunoblot. Purification of CD36 by fast protein liquid chromatography (FPLC) from Triton X-100 extracts of normal platelet membranes resulted in the isolation of two different forms: the classic 88 kD form, and a second, lighter 85 kD form. Our data indicated that the patients' plasma autoantibodies reacted strongly with the 85 kD form. Conventional monoclonal and polyclonal antisera produced to the 88 kD form reacted strongly with the 88 kD form but weakly with the 85 kD form. These results confirm the possible importance of anti-CD36 antibodies in the pathophysiology of TTP and other thrombocytopenias and demonstrate the presence of a previously unrecognized target antigen for these antibodies.     

Analytical performance evaluation of ADVIA Chemistry Carbamazepine_2 assay: minimal cross‐reactivity with carbamazepine 10, 11‐epoxide and none with hydroxyzine or cetirizine

Carbamazepine is an anticonvulsant requiring routine therapeutic drug monitoring. Recently, Siemens Healthcare Diagnostic Division released a new carbamazepine assay: ADVIA Chemistry Carbamazepine_2 (Carbamazepine_2) for application on ADVIA analyzers. We evaluated the analytical performance of this assay as well as its potential cross‐reactivities with carbamazepine 10, 11‐epoxide, hydroxyzine, and cetirizine. The within‐run and between‐run precisions of the Carbamzepine‐2 assay were <6% and limit of detection was 0.5 µg/ml using ADVIA 1800 analyzer. The assay was linear up to a carbamazepine concentration of 20.0 µg/ml. The new method compared well with a widely used carbamazepine EMIT 2000 assay on the Hitachi 917 analyzer. Using 75 patients' specimens (where carbamazepine concentrations varied from 0.5 to 21.7 µg/ml) and carbamazepine EMIT 2000 as the reference method (x‐axis), we observed the following regression equation: y=1.04 x+0.32 (r=0.99). The new carbazepine_2 method was not affected by a hemoglobin concentration of 1,000 mg/dl, conjugated or unconjugated bilirubin concentration of 60 mg/dl, and triglyceride concentration of 1,000 mg/dl. In addition, this assay showed no cross‐reactivity with hydroxyzine or cetirizine and demonstrated minimal cross‐reactivity with carbamazepine 10, 11‐epoxide. We conclude that the ADVIA Chemistry carbamazepine_2 assay has adequate precision and accuracy for routine therapeutic drug monitoring of carbamazepine in clinical laboratories. J. Clin. Lab. Anal. 24:278–282, 2010. © 2010 Wiley‐Liss, Inc.     

Le dosage de l’Insulin-Like Growth Factor-1 : les difficultés de la détermination sérique et de l’interprétation des résultats

The IGF-I assay is now at the crossroads of endocrine pathology, which is essential both for diagnosis and for therapeutic management. However, due to the complexity and the intricacies of the regulatory factors and the existence of technical difficulties, results’ interpretation is far from a mundane procedure. In this work, we recalled the main sources of interference, the particular clinical situations that could compromise the results and the recommendations for each disputed case because we think it is useful for any practitioner to be warned in order to better manage them and to minimize errors ‘risk.     

比较两种自动化免疫分析仪对血清25-羟维生素D的测定

目的 比较Architect I2000和DiaSorin LIAISON两种自动化免疫分析仪对血清25-羟维生素D［25(OH)D］测定的相关性及差异，为临床诊疗提供参考依据。方法 根据美国临床实验室标准化委员会（NCCLS）的EP9-A2文件，分别在两个系统检测三个水平（L1, L2, L3）的25(OH)D质控品，通过Excel 2007软件分析两个系统测定25(OH)D的不精密度。收集266份临床样本进行25(OH)D检测，将样本分成两份在本实验室Architect I2000和DiaSorin LIAISON系统同时进行检测。采用MedCalc软件进行统计学分析，以Passing-Bablok回归和Pearson相关系数分析两种方法测定结果之间的相关性，以Bland-Altman比较它们之间的差异。结果 在不精密度评估中，两个系统（DiaSorin LIAISON vs Architect I2000）三个质控水平（L1, L2, L3）的总不精密度分别为L1: 8.86% vs 7.89%，L2: 5.82% vs 4.67%，L3: 6.66% vs 5.71%。Passing-Bablok和Pearson分析显示Architect I2000和DiaSorin LIAISON间相关系数为r=0.98。Bland-Altman分析结果显示，Architect I2000检测的25(OH)D比DiaSorin LIAISON的平均高7.2ng/ml，随着25(OH)D的增高，两者的差值越来越大，最大差值可达43.3ng/ml。结论 Architect I2000和DiaSorin LIAISON两个系统检测的25(OH)D相关性较好。Architect I2000精密度优于DiaSorin LIAISON，Architect I2000系统得到25(OH)D的结果更为准确，与临床诊断更相符。