Search

Laboratory-developed tests (LDTs) are widely used in clinical hemostasis laboratories. The extent to which LDTs are regulated varies greatly around the world, and proposed changes to regulations have raised concerns about the future of LDTs in clinical laboratories. It is increasingly difficult to justify the use of an LDT where a commercially available method with regulatory approval is available. Conversely, where there is no suitable test with regulatory approval and there is a compelling clinical need, using an LDT outweighs the risk associated with not performing the test. We argue that LDTs are still required in specialist clinical laboratories to fulfill unmet clinical needs, and in lower middle-income countries where they are a vital resource., Competing Interests: C.G. reports personal fees from Sysmex UK, outside the submitted work.R.C.G. reports personal fees from Sysmex America Inc, personal fees from Diagnostica Grifols, other from Diagnostica Stago, outside the submitted work., (Thieme. All rights reserved.)

Background: Severe high-molecular-weight kininogen (HK) deficiency is a poorly studied autosomal recessive contact system defect caused by pathogenic, biallelic KNG1 variants., Aim: We performed the first comprehensive analysis of diagnostic, clinical, genetic, and epidemiological aspects of HK deficiency., Methods: We collected clinical information and blood samples from a newly detected HK-deficient individual and from published cases identified by a systematic literature review. Activity and antigen levels of coagulation factors were determined. Genetic analyses of KNG1 and KLKB1 were performed by Sanger sequencing. The frequency of HK deficiency was estimated considering truncating KNG1 variants from GnomAD., Results: We identified 48 cases of severe HK deficiency (41 families), of these 47 have been previously published (n = 19 from gray literature). We genotyped 3 cases and critically appraised 10 studies with genetic data. Ten HK deficiency-causing variants (one new) were identified. All of them were truncating mutations, whereas the only known HK amino acid substitution with a relevant phenotype instead causes hereditary angioedema. Conservative estimates suggest an overall prevalence of severe HK deficiency of approximately one case per 8 million population, slightly higher in Africans. Individuals with HK deficiency appeared asymptomatic and had decreased levels of prekallikrein and factor XI, which could lead to misdiagnosis., Conclusion: HK deficiency is a rare condition with only few known pathogenic variants. It has an apparently good prognosis but is prone to misdiagnosis. Our understanding of its clinical implications is still limited, and an international prekallikrein and HK deficiency registry is being established to fill this knowledge gap., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Before a new method is used for clinical testing, it is essential that it is evaluated for suitability for its intended purpose. This document gives guidance for the performance, verification and implementation processes required by regulatory and accreditation bodies. It covers the planning and verification of specialist haemostatic tests, including factor assays, D-dimers, direct anticoagulants and thrombophilia testing., (© 2021 John Wiley & Sons Ltd.)

Before a new method is used for clinical testing, it is essential that it is evaluated for suitability for its intended purpose. This document gives guidance for the performance of verification, validation and implementation processes required by regulatory and accreditation bodies. It covers the planning and execution of an evaluation of the commonly performed screening tests (prothrombin time, activated partial thromboplastin time, thrombin time and fibrinogen assay), and instrument-specific issues. Advice on selecting an appropriate haemostasis analyser, planning the evaluation, and assessing the reference, interval, precision, accuracy, and comparability of a haemostasis test system are also given. A second companion document will cover specialist haemostasis testing., (© 2020 John Wiley & Sons Ltd.)

Dengue is a major public health problem worldwide with distinct clinical manifestations: an acute presentation (dengue fever, DF) similar to other febrile illnesses (OFI) and a more severe, life-threatening form (severe dengue, SD). Due to nonspecific clinical presentation during the early phase of dengue infection, differentiating DF from OFI has remained a challenge, and current methods to determine severity of dengue remain poor early predictors. We present a prospective clinical cohort study conducted in Caracas, Venezuela from 2001-2005, designed to determine whether clinical and hematological parameters could distinguish DF from OFI, and identify early prognostic biomarkers of SD. From 204 enrolled suspected dengue patients, there were 111 confirmed dengue cases. Piecewise mixed effects regression and nonparametric statistics were used to analyze longitudinal records. Decreased serum albumin and fibrinogen along with increased D-dimer, thrombin-antithrombin complex, activated partial thromboplastin time and thrombin time were prognostic of SD on the day of defervescence. In the febrile phase, the day-to-day rates of change in serum albumin and fibrinogen concentration, along with platelet counts, were significantly decreased in dengue patients compared to OFI, while the day-to-day rates of change of lymphocytes (%) and thrombin time were increased. In dengue patients, the absolute lymphocytes to neutrophils ratio showed specific temporal increase, enabling classification of dengue patients entering the critical phase with an area under the ROC curve of 0.79. Secondary dengue patients had elongation of Thrombin time compared to primary cases while the D-dimer formation (fibrinolysis marker) remained always lower for secondary compared to primary cases. Based on partial analysis of 31 viral complete genomes, a high frequency of C-to-T transitions located at the third codon position was observed, suggesting deamination events with five major hot spots of amino acid polymorphic sites outside in non-structural proteins. No association of severe outcome was statistically significant for any of the five major polymorphic sites found. This study offers an improved understanding of dengue hemostasis and a novel way of approaching dengue diagnosis and disease prognosis using piecewise mixed effect regression modeling. It also suggests that a better discrimination of the day of disease can improve the diagnostic and prognostic classification power of clinical variables using ROC curve analysis. The piecewise mixed effect regression model corroborated key early clinical determinants of disease, and offers a time-series approach for future vaccine and pathogenesis clinical studies., Competing Interests: IB and BBH are co-founders of E25Bio Inc. (www.e25bio.com), a company that develops point-of-care diagnostics for fever-causing infectious agents.

Summary: This review outlines a number of key issues when performing laboratory testing of homeostasis. The effect pre-analytical variables have on the reliability and consistency of screening tests is often forgotten due to a lack of understanding and awareness. This can be improved through educating healthcare professionals who are involved in taking blood for assessment. Recent advances in coagulation testing have not enabled laboratories to replace the Prothrombin Time (PT) and Activated Partial Thromboplastin Time (APTT) screening tests with more advanced assays and they continue to play an important role with the advantage of being easily automated. However, there are many analysers on the market, each with varying sensitivity to coagulation defects and it is important to keep this in mind when interpreting results.

Therapeutic options for developing countries have to assure an optimum safety and efficacy and low-cost antihaemophilic concentrates. A single blind randomized crossover study was carried out in 12 previously treated HB patients, comparing the pharmacokinetics (PK), thrombogenicity (TG) and safety of two plasma-derived double-inactivated (solvent/detergent heating at 100 degrees C, 30 min) factor IX (FIX) concentrates, UMAN COMPLEX DI (product A) [plasma-derived prothrombin concentrates (PCC)] and a high purity FIX concentrate AIMAFIX DI (product B, HPFIX). In a non-bleeding state, they received one single intravenous dose 50 IU FIX kg(-1) of PCC or HPFIX, and after a wash-out period of 14 days, the other product. We evaluated acute tolerance and determined PK parameters based on FIX levels measured over a 50 h postinfusion period. We studied fibrinogen, platelets, antithrombin, F1 + 2, TAT, D-dimer, over a 360 min postinfusion period. Ten cases remained in on-demand treatment for 6 months, five with PCC and five with HPFIX. PK and anti-FIX inhibitors were repeated at 3 and 6 months. No inhibitors were detected. PK values (PCC vs. HPFIX): clearence (CL; mL h(-1) kg(-1)) 5.2 +/- 1.4 vs. 6.5 +/- 1.4; the volume of distribution at steady state (mL kg(-1)) 154.9 +/- 54.9 vs. 197.5 +/- 72.5; mean residence time (h) 29.7 +/- 8.1 vs. 30.7 +/- 9.2; T(1/2) (h) 22.3 +/- 7 vs. 23.5 +/- 12.3; incremental recovery (IR; U dL(-1) U(-1) kg(-1)) 0.96 +/- 0.17 vs. 0.76 +/- 0.13. HPFIX showed significant lower IR and higher CL. There were no differences in PK at 3 and 6 months. In TG, significant increments in TAT and F1 + 2 at 30 min and 6 h were found with PCC. Product B PK results agrees with reported results for other HPFIX preparations. Use of PCC product A has to consider its thrombogenic activity.

The adsorption of thrombin to fibrin during clotting defines "Antithrombin I" activity. We confirmed that thrombin generation in afibrinogenemic or in Reptilase defibrinated normal plasma was higher than in normal plasma. Repletion of these fibrinogen-deficient plasmas with fibrinogen 1 (gamma A/gamma A), whose fibrin has two "low affinity" non-substrate thrombin binding sites, resulted in moderately reduced thrombin generation by 29-37%. Repletion with fibrinogen 2 (gamma'/gamma A), which in addition to low affinity thrombin-binding sites in fibrin, has a "high affinity" non-substrate thrombin binding site in the carboxy-terminal region of its gamma' chain, was even more effective and reduced thrombin generation by 57-67%. Adding peptides that compete for thrombin binding to fibrin [S-Hir53-64 (hirugen) or gamma'414-427] caused a transient delay in the onset of otherwise robust thrombin generation, indicating that fibrin formation is necessary for full expression of Antithrombin I activity. Considered together, 1) the increased thrombin generation in afibrinogenemic or fibrinogen-depleted normal plasma that is mitigated by fibrinogen replacement; 2) evidence that prothrombin activation is increased in afibrinogenemia and normalized by fibrinogen replacement; 3) the severe thrombophilia that is associated with defective thrombin-binding in dysfibrinogenemias Naples I and New York I, and 4) the association of afibrinogenemia or hypofibrinogenemia with venous or arterial thromboembolism, indicate that Antithrombin I (fibrin) modulates thromboembolic potential by inhibiting thrombin generation in blood.