Although the function of platelets in the maintenance of hemostasis has been studied in great detail, more recent evidence has highlighted a central role for platelets in the host inflammatory and immune responses. Platelets by virtue of their large numbers and their ability to rapidly release a broad spectrum of immunomodulatory cytokines, chemokines, and other mediators act as circulating sentinels. Upon detection of a pathogen, platelets quickly activate and begin to drive the ensuing inflammatory response. Platelets have the ability to directly modulate the activity of neutrophils (phagocytosis, oxidative burst), endothelium (adhesion molecule and chemokine expression), and lymphocytes. Due to their diverse array of adhesion molecules and preformed chemokines, platelets are able to adhere to leukocytes and facilitate their recruitment to sites of tissue damage or infection. Furthermore, platelets directly participate in the capture and sequestration of pathogens within the vasculature. Plateletneutrophil interactions are known to induce the release of neutrophil extracellular traps (NETs) in response to either bacterial or viral infection, and platelets have been shown to internalize pathogens, sequestering them in engulfment vacuoles. Finally, emerging data indicate that platelets also participate in the host immune response by directly killing infected cells. This review will highlight the central role platelets play in the initiation and modulation of the host inflammatory and immune responses.
IntroductionThe purpose of this study is to compare the effect of increasing concentrations of direct anti-Xa oral anticoagulants (DOAC) apixaban-, edoxaban-, and rivaroxaban-enriched plasma samples on various prothrombin time (PT), activated partial thromboplastin time (APTT), heparin calibrated anti-Xa methods, and other coagulation assays. MethodsApixaban, edoxaban, or rivaroxaban was dissolved in dimethylsulfoxide and added to pooled normal plasma to obtain concentrations ranging from 0 ng/mL to approximately 600 ng/mL. The samples were tested using Innovin((R)), Neoplastine((R)) CI Plus, Recombiplastin 2G, and Thromborel((R)) S for PT testing and Actin, Actin((R)) FS, Actin((R)) FSL, APTT-Automate, and SynthaSIL for APTT. Samples were also tested using four different anti-Xa methods calibrated for unfractionated heparin or low molecular weight heparin. Special coagulation assays included antithrombin activity, lupus anticoagulant assays, and others. For special coagulation assays, the concentration of DOAC that resulted in a >15% change from baseline value was determined. DOAC quantification was performed using liquid chromatography-tandem mass spectrometry. ResultsAll PT and APTT reagents demonstrated a higher sensitivity for edoxaban and rivaroxaban than for apixaban. Anti-Xa methods were able to detect low concentrations of DOAC. DOACs effected special coagulation assays to differing degrees, with lupus anticoagulant testing using dilute viper venom, the most sensitive test to the presence of anti-Xa DOAC. ConclusionNo PT or APTT reagent system effectively detected apixaban. All anti-Xa methods demonstrated sensitivity to low concentrations of DOAC. Dilute viper venom methods are exquisitely sensitive to anti-Xa DOAC, suggesting potential use of this assay for screening or measuring these drugs.
IntroductionHereditary spherocytosis (HS), hereditary elliptocytosis (HE), and hereditary stomatocytosis (HSt) are inherited red cell disorders caused by defects in various membrane proteins. The heterogeneous clinical presentation, biochemical and genetic abnormalities in HS and HE have been well documented. The need to raise the awareness of HSt, albeit its much lower prevalence than HS, is due to the undesirable outcome of splenectomy in these patients. MethodsThe scope of this guideline is to identify the characteristic clinical features, the red cell parameters (including red cell morphology) for these red cell disorders associated, respectively, with defective cytoskeleton (HS and HE) and abnormal cation permeability in the lipid bilayer (HSt) of the red cell. The current screening tests for HS are described, and their limitations are highlighted. ResultsAn appropriate diagnosis can often be made when the screening test result(s) is reviewed together with the patient's clinical/family history, blood count results, reticulocyte count, red cell morphology, and chemistry results. SDS-polyacrylamide gel electrophoresis of erythrocyte membrane proteins, monovalent cation flux measurement, and molecular analysis of membrane protein genes are specialist tests for further investigation. ConclusionSpecialist tests provide additional evidence in supporting the diagnosis and that will facilitate the management of the patient. In the case of a patient's clinical phenotype being more severe than the affected members within the immediate family, molecular testing of all family members is useful for confirming the diagnosis and allows an insight into the molecular basis of the abnormality such as a recessive mode of inheritance or a de novo mutation.
This revision is intended to update the 1994 ICSH guidelines. It is based on those guidelines but is updated to include new methods, such as digital image analysis for blood cells, a flow cytometric method intended to replace the reference manual 400 cell differential, and numerous new cell indices not identified morphologically are introduced. Haematology analysers are becoming increasingly complex and with technological advancements in instrumentation with more and more quantitative parameters are being reported in the complete blood count. It is imperative therefore that before an instrument is used for testing patient samples, it must undergo an evaluation by an organization or laboratory independent of the manufacturer. The evaluation should demonstrate the performance, advantages and limitations of instruments and methods. These evaluations may be performed by an accredited haematology laboratory where the results are published in a peer‐reviewed journal and compared with the validations performed by the manufacturer. A less extensive validation/transference of the equipment or method should be performed by the local laboratory on instruments prior to reporting of results.
These guidelines provide information on how to reliably and consistently report abnormal red blood cells, white blood cells and platelets using manual microscopy. Grading of abnormal cells, nomenclature and a brief description of the cells are provided. It is important that all countries in the world use consistent reporting of blood cells. An international group of morphology experts have decided on these guidelines using consensus opinion. For some red blood cell abnormalities, it was decided that parameters produced by the automated haematology analyser might be more accurate and less subjective than grading using microscopy or automated image analysis and laboratories might like to investigate this further. A link is provided to show examples of many of the cells discussed in this guideline.
Introduction Studies have shown dabigatran to be an effective anticoagulant with an acceptable bleeding profile. None the less, these patients do suffer from bleeding complications. Unfortunately, there are currently no direct reversal agents to dabigatran or established guidelines on the management of bleeding in these circumstances. Methods We examined the effects on thrombin generation parameters, after ex-vivo spiking the plasma of patients on dabigatran (n=8) with FEIBA (R). These parameters were measured using the calibrated automated thrombography (CAT) machine. Results In our study, we showed the ability of FEIBA (R) to improve the abnormal thrombin generation parameters caused by dabigatran in these patients. Conclusion This provides evidence, lacking in the literature, that this agent may be able to provide haemostatic support in situations where dabigatran induced coagulopathy exists.
Recently several parameters have been introduced to the complete blood count such as nucleated red blood cells, immature granulocytes; immature reticulocyte fraction, immature platelet fraction and red cell fragments as well as new parameters for detection of functional iron deficiency. Leucocyte positional parameters, which may diagnose specific diseases (e.g. differentiate between abnormal lymphocytes in leukaemia and viral conditions and may also detect malarial infection) are now available. At this time they are only used for research; however, generally such parameters later become reportable. One manufacturer's routine analyser allows measurement of cells by flow cytometry using monoclonal antibodies. Currently, there are no accredited external quality assessment schemes (EQAS) for these parameters. For a number of parameters, on some instruments, there is no internal quality control, which brings into question whether these parameters should be used for clinical decision making. Other more established parameters, such as mean platelet volume, red cell distribution width and the erythrocyte sedimentation rate do not have EQAS available. The UK National EQAS for General Haematology held a workshop earlier this year in 2008 to discuss these parameters. Participants were asked to provide a consensus opinion on which parameters are the most important for inclusion in future haematology EQAS.
BackgroundIron is essential for hemoglobin synthesis during terminal erythropoiesis. To supply adequate iron the carrier transferrin is required together with transferrin receptor endosomal cycle and normal mitochondrial iron utilization. Iron and iron protein deficiencies result in different types of anemia. Iron-deficiency anemia is the commonest anemia worldwide due to increased requirements, malnutrition, chronic blood losses and malabsorption. Mutations of transferrin, transferrin receptor cycle proteins, enzymes of the first step of heme synthesis and iron sulfur cluster biogenesis lead to rare anemias, usually accompanied by iron overload. Hepcidin plays an indirect role in erythropoiesis by controlling plasma iron. Inappropriately high hepcidin levels characterize the rare genetic iron-refractory iron-deficiency anemia (IRIDA) and the common anemia of chronic disease. Iron modulates both effective and ineffective erythropoiesis: iron restriction reduces heme and alpha-globin synthesis that may be of benefit in thalassemia. Material and MethodsThis review relies on the analysis of the most recent literature and personal data. ResultsErythropoiesis controls iron homeostasis, by releasing erythroferrone that inhibits hepcidin transcription to increase iron acquisition in iron deficiency, hypoxia and EPO treatment. Erythroferrone, produced by EPO-stimulated erythropoiesis, inhibits hepcidin only when the activity of BMP/SMAD pathway is low, suggesting that EPO somehow modulates the latter signaling. Erythroblasts sense circulating iron through the second transferrin receptor (TFR2) that, in animal models, modulates the sensitivity of the erythroid cells to EPO. DiscussionThe advanced knowledge of the regulation of systemic iron homeostasis and erythropoiesis-mediated hepcidin regulation is leading to the development of targeted therapies for anemias and iron disorders.
Schistocytes are fragments of red blood cells (RBCs) produced by extrinsic mechanical damage within the circulation. The detection of schistocytes is an important morphological clue to the diagnosis of thrombotic microangiopathic anemia (TMA). Reporting criteria between different laboratories, however, are not uniform, owing to variability of shape and nature of fragments, as well as subjectivity and heterogeneity in their morphological assessment. Lack of standardization may lead to inconsistency or misdiagnosis, thereby affecting treatment and clinical outcome. The Schistocyte Working Group of the International Council for Standardization in Haematology (ICSH) has prepared specific recommendations to standardize schistocyte identification, enumeration, and reporting. They deal with the type of smear, method of counting, morphological description based on positive criteria (helmet cells, small, irregular triangular, or crescent‐shaped cells, pointed projections, and lack of central pallor). A schistocyte count has a definite clinical value for the diagnosis of TMA in the absence of additional severe red cell shape abnormalities, with a confident threshold value of 1%. Automated counting of RBC fragments is also recommended by the ICSH Working Group as a useful complement to the microscope, according to the high predictive value of negative results, but worthy of further research and with limits in quantitation.
IntroductionThe Sysmex XN-series is a new automated hematology analyzer designed to improve the accuracy of cell counts and the specificity of the flagging events. MethodsThe basic characteristics and the performance of new measurement channels of the XN were evaluated and compared with the Sysmex XE-2100 and the manual method. Fluorescent platelet count (PLT-F) was compared with the flow cytometric method. The low WBC mode and body fluid mode were also evaluated. For workflow analysis, 1005 samples were analyzed on both the XN and the XE-2100, and manual review rates were compared. ResultsAll parameters measured by the XN correlated well with the XE-2100. PLT-F showed better correlation with the flow cytometric method (r(2)=0.80) compared with optical platelet count (r(2)=0.73) for platelet counts <70x10(9)/L. The low WBC mode reported accurate leukocyte differentials for samples with a WBC count <0.5x10(9)/L. Relatively good correlation was found for WBC counts between the manual method and the body fluid mode (r=0.88). The XN made less flags than the XE-2100, while the sensitivities of both instruments were comparable. ConclusionThe XN provided reliable results on low cell counts, as well as reduced manual blood film reviews, while maintaining a proper level of diagnostic sensitivity.
Introduction Oral direct thrombin and Xa inhibitors are worldwide distributed for prevention and treatment of thrombosis. It is important to recognize their effects on lupus anticoagulant (LA) testing. The aim of the study is to describe the rate of false-positive results of LA tests on plasmas of patients with previous negative LA tests results that receive dabigatran etexilate (DAB) 110mg/twice a day, rivaroxaban (RIV) 10mg/day or 15mg/twice a day, or enoxaparin 40mg/day. Methods Blood was taken between 1.5 and 4h post administration. Tests evaluated are as follows: prothrombin time, APTT, dilute Russell viper venom time (DRVVT) screen, APTT, and DRVVT mixing studies, index of circulating anticoagulant (ICA) with normal plasma, screen/confirm normalized ratio (NR) for DRVVT and silica clotting time (SCT). Results Plasmas from patients taking DAB (n=22) presented 100% prolonged APTT and DRVVT with ICA above the cutoff point and 81.8% positive screen/confirm NR, 100% prolonged SCT screen, but 4.5% positive confirmatory NR. All patients receiving RIV at 15mg/twice a day (n=4) presented positive DRVVT screen, mixing, and confirmatory tests, 75% and 100% prolonged APTT and SCT screen, with negative screen/confirm NR. Those taking RIV 10mg/day (n=22) showed 81.8% prolonged DRVVT screen, 82.3% and 76.5% of them with positive mixing and confirmatory studies. Patients receiving enoxaparin also presented high prevalence of APTT and DRVVT false-positive results. Conclusion Dabigatran etexilate, RIV, and enoxaparin affect tests for LA not only in screening and mixing, but also in confirmatory studies. We considered that LA testing should not to be performed when patients are taken these drugs, particularly if blood is collected at peak, in order to avoid false-positive results.
IntroductionRotational Thromboelastometry (ROTEM) is a point of care method used to monitor coagulation during surgery and to guide transfusion strategies in patients presenting with severe bleeding. The aim of our study was to determine the impact of four direct oral anticoagulants (DOACs) on 3 commonly used ROTEM tests. MethodsWhole blood samples from 20 healthy donors were spiked in vitro with apixaban, edoxaban, rivaroxaban or dabigatran at 5 different plasma concentrations (0-1000ng/mL). EXTEM, INTEM and FIBTEM tests were systematically performed. ResultsThere was a linear relationship between the increase in clotting time (CT) and plasma DOAC concentrations in both the EXTEM and INTEM tests. We found that the DOAC concentration required to double EXTEM CT was 1042225ng/mL for apixaban, 134 +/- 38ng/mL for edoxaban, 176 +/- 26ng/mL for rivaroxaban and 284 +/- 73ng/mL for dabigatran. INTEM CT was less sensitive than EXTEM CT whatever the anticoagulant. EXTEM CT was above the normal range for 5 of 5 spiked samples when the plasma concentrations were similar to 1000ng/mL for apixaban, similar to 100ng/mL for edoxaban, similar to 200ng/mL for rivaroxaban and similar to 200ng/mL for dabigatran. Maximum Clot Firmness in EXTEM, INTEM and FIBTEM tests was not affected whatever the DOAC or its concentration. ConclusionThis study found a DOAC dose-dependent increase in ROTEM CTs. ROTEM tests were only poorly impacted by low levels of edoxaban, rivaroxaban or dabigatran. Apixaban had only a low effect even at high concentrations.
The foundation of laboratory hematologic diagnosis is the complete blood count and review of the peripheral smear. In patients with anemia, the peripheral smear permits interpretation of diagnostically significant red blood cell (RBC) findings. These include assessment of RBC shape, size, color, inclusions, and arrangement. Abnormalities of RBC shape and other RBC features can provide key information in establishing a differential diagnosis. In patients with microcytic anemia, RBC morphology can increase or decrease the diagnostic likelihood of thalassemia. In normocytic anemias, morphology can assist in differentiating among blood loss, marrow failure, and hemolysisand in hemolysis, RBC findings can suggest specific etiologies. In macrocytic anemias, RBC morphology can help guide the diagnostic considerations to either megaloblastic or nonmegaloblastic causes. Like all laboratory tests, RBC morphologies must be interpreted with caution, particularly in infants and children. When used properly, RBC morphology can be a key tool for laboratory hematology professionals to recommend appropriate clinical and laboratory follow-up and to select the best tests for definitive diagnosis.
Introduction: The aim of this study was to show variability in the measurement of the mean platelet volume (MPV) depending on the instrument used. Methods: This prospective analysis was carried out to measure MPV with three instruments, in 30 healthy controls and 113 hospital patients. Results: Firstly, for values in the normal range, the values obtained with the Siemens Advia ® 2120 are lower than those given by the Beckman Coulter LH750 ® (−0.89), which are in turn lower than those obtained with the Sysmex XE‐2100D ® (−1.11), which represents a 20–25% variation in the measurement. These results emphasize the lack of universal external calibration for MPV analysis and thus make any intercentre comparison of MPV impossible unless the automated haematology analyser used is indicated. Secondly, we stress the differences in behaviour of the instruments in the presence of abnormally large platelets, i.e. an underestimate of the platelet count and the MPV may be provided because instruments using impedance technology may fail to take into account these platelets, but they rightly flag them. Conclusion: To harmonize our procedures, we propose definitions of platelet size (normal size, macroplatelets and giant platelets) based on the coordinated interpretation of the MPV, the distribution of platelet volume and the morphological appearance.
IntroductionMean platelet volume (MPV) and platelet (PLT) count are the two major parameters that reflect the functions and activities of PLTs. The associations of MPV and PLT count with the occurrence and prognosis of stroke have not been fully clarified. This study aimed to investigate the association of MPV and PLT count with the development and prognosis of first-ever ischemic and hemorrhagic stroke in order to provide evidence for early diagnosis and treatment of both strokes. MethodsThis study included 281 first-ever ischemic stroke and 164 first-ever hemorrhagic stroke patients between 2010 and 2012. All participants received routine blood tests within 2 h after admission and were categorized into good or poor prognosis group based on the Modified Rankin Scale (mRS) score. MPV and PLT counts were transformed into categorical variables and their association with the occurrence and prognosis of both strokes was evaluated by multivariate logistic regression. ResultsThe risk of ischemic and hemorrhagic stroke in MPV group (>13 fL) was 22.17 and 5.21 times higher compared with normal MPV group. The PLT count was positively correlated with the risk of ischemic stroke, but negatively correlated with the risk of hemorrhagic stroke. MPV and PLT count was not correlated with the prognosis of either stroke. ConclusionsIncreased MPV is an independent risk factor for both strokes. Elevated PLT count increases the risk for ischemic stroke, but decreases the risk for hemorrhagic stroke. However, neither MPV nor PLT count has significant association with the prognosis of either stroke.
Treatment of acute myelogenous leukemia ( AML ) over the past four decades remains mostly unchanged and the prognosis for the majority of patients remains poor. Most of the significant advances that have been observed are in defining cytogenetic abnormalities, as well as the genetic and epigenetic profiles of AML patients. While new cytogenetic and genetic aberrations such as the FLT 3‐ ITD and NPM 1 mutations are able to guide prognosis for the majority of patients with AML , outcomes are still dismal and relapse rates remain high. It is thought that relapse in AML is in part driven by minimal residual disease ( MRD ) that remains in the patient following treatment. Thus, there is a need for sensitive and objective methodology for MRD detection. Methodologies such as multiparameter flow cytometry ( MFC ), quantitative real‐time polymerase chain reaction ( RQ ‐ PCR ), digital PCR ( dPCR ), or next‐generation sequencing ( NGS ) are being employed to evaluate their utility in MRD assessment. In this review, we will provide an overview of AML and the clinical utility of MRD measurement. We will discuss optimal timing to MRD measurement, the different approaches that are available, and efforts in the standardization across laboratories.
Although DNA analysis is needed for characterization of the mutations that cause β‐thalassaemia, measurement of the Hb A 2 is essential for the routine identification of people who are carriers of β‐thalassaemia. The methods of quantitating Hb A 2 are described together with pitfalls in undertaking these laboratory tests with particular emphasis on automated high‐performance liquid chromatography and capillary electrophoresis.
Central venous access devices are used in many branched of medicine where venous access is required for either long‐term or a short‐term care. These guidelines review the types of access devices available and make a number of major recommendations. Their respective advantages and disadvantages in various clinical settings are outlined. Patient care prior to, and immediately following insertion is discussed in the context of possible complications and how these are best avoided. There is a section addressing long‐term care of in‐dwelling devices. Techniques of insertion and removal are reviewed and management of the problems which are most likely to occur following insertion including infection, misplacement and thrombosis are discussed. Care of patients with coagulopathies is addressed and there is a section addressing catheter‐related problems.