Evaluating Diagnostic Algorithms for Heparin-Induced Thrombocytopenia using Two Combined Automated Rapid Immunoassays.

Heparin-induced thrombocytopenia (HIT) is an autoimmune disorder caused by antibodies against platelet factor 4 (PF4) and heparin complexes. Rapid immunoassays (IAs) for detection of these antibodies mark a milestone in HIT diagnosis, despite a higher false-positive rate compared with functional platelet-activation assays. However, combining different rapid IAs may help to improve their diagnostic specificity.

The clinical and laboratory diagnosis of vaccine-induced immune thrombotic thrombocytopenia.

Vaccine-induced immune thrombotic thrombocytopenia (VITT) is a rare but serious adverse syndrome occurring 5 to 30 days after adenoviral vector COVID-19 vaccination. Therefore, a practical evaluation of clinical assessments and laboratory testing for VITT is needed to prevent significant adverse outcomes as the global use of adenoviral vector vaccines continues. We received the clinical information and blood samples of 156 patients in Canada with a suspected diagnosis of VITT between April and July 2021.

Characteristics of Anti-SARS-CoV-2 Antibodies in Recovered COVID-19 Subjects.

Coronavirus Disease 2019 (COVID-19) is a global pandemic caused by the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). While detection of SARS-CoV-2 by polymerase chain reaction with reverse transcription (RT-PCR) is currently used to diagnose acute COVID-19 infection, serological assays are needed to study the humoral immune response to SARS-CoV-2. Anti-SARS-CoV-2 immunoglobulin (Ig)G/A/M antibodies against spike (S) protein and its receptor-binding domain (RBD) were characterized in recovered subjects who were RT-PCR-positive ( = 153) and RT-PCR-negative ( = 55) using an enzyme-linked immunosorbent assay (ELISA).

Post-cardiac surgery thrombotic thrombocytopenic purpura: presence of anti-ADAMTS13 autoantibodies at preoperative baseline.

Thrombotic thrombocytopenic purpura (TTP) rarely complicates acute inflammatory conditions such as surgery, including post-cardiac surgery. Review of 32 previously-reported cases of post-cardiac surgery TTP indicates that this disorder often occurs as early as 2-3 days following surgery, which seems too soon to implicate new formation of anti-ADAMTS13 autoantibodies as a consequence of surgery itself. We diagnosed post-cardiac surgery TTP in a 60-year-old female that began approximately 3 days post-coronary artery bypass surgery in which anti-ADAMTS13 autoantibodies were implicated.

Platelet-activating immune complexes identified in critically ill COVID-19 patients suspected of heparin-induced thrombocytopenia.

Background: Thrombocytopenia and thrombosis are prominent in coronavirus disease 2019 (COVID-19), particularly among critically ill patients; however, the mechanism is unclear. Such critically ill COVID-19 patients may be suspected of heparin-induced thrombocytopenia (HIT), given similar clinical features.

Objectives: We investigated the presence of platelet-activating anti-platelet-factor 4 (PF4)/heparin antibodies in critically ill COVID-19 patients suspected of HIT.

Serotonin-release assay-positive but platelet factor 4-dependent enzyme-immunoassay negative: HIT or not HIT?

IgG-specific and polyspecific PF4-dependent enzyme-immunoassays (EIAs) have exceptionally high sensitivity (≥99%) for diagnosis of heparin-induced thrombocytopenia (HIT), a drug reaction caused by platelet-activating antibodies detectable by serotonin-release assay (SRA). The IgG-specific EIAs are recommended for screening, as their high sensitivity is accompanied by relatively high specificity vis-à-vis polyspecific EIAs. We investigated the frequency of SRA-positive/EIA-negative (SRA+/EIA-) HIT, prompted by referral to our reference HIT laboratory of serial blood samples from a patient ("index case") with false-negative IgG-specific EIAs.

Combination of two complementary automated rapid assays for diagnosis of heparin-induced thrombocytopenia (HIT).

Background: HIT diagnosis typically uses complementary diagnostic assays (eg, a PF4-dependent enzyme-immunoassay [EIA] and a platelet activation assay such as the serotonin-release assay [SRA]).

Objectives: To determine whether the combination of two automated assays-a latex immunoturbidimetric assay (LIA) that evaluates competitive inhibition of a HIT-like monoclonal antibody and a chemiluminescence immunoassay (CLIA) for detecting anti-PF4/heparin IgG-optimizes diagnostic sensitivity while also yielding good specificity, particularly at high assay reactivities.

Patients/methods: We determined operating characteristics using combined LIA/CLIA results from a HIT observational trial (n = 430; derivation cohort) and 147 consecutive patients with HIT (n = 147; supplementary derivation cohort).

Autoantibodies to thrombopoietin and the thrombopoietin receptor in patients with immune thrombocytopenia.

Autoantibodies to thrombopoietin (TPO, also termed THPO) or the TPO receptor (cMpl, also termed MPL) could play a pathological role in immune thrombocytopenia (ITP). In this study, we tested for autoantibodies against TPO, cMpl, or the TPO/cMpl complex in ITP and other thrombocytopenic disorders. Using an inhibition step with excess TPO in fluid-phase to improve binding specificity, the prevalence of anti-TPO autoantibodies was: active ITP: 9/32 (28%); remission ITP: 0/14 (0%); non-immune thrombocytopenias: 1/10 (10%); and healthy controls: 1/11 (9%).

Development of a high-yield expression and purification system for platelet factor 4.

Heparin-induced thrombocytopenia (HIT) is an adverse drug reaction characterized by IgG antibodies bound to complexes of platelet factor 4 (PF4) and heparin. The majority of diagnostic tests for HIT rely on an exogenous source of PF4 to identify anti-PF4/heparin antibodies. These include the PF4-dependent enhanced serotonin release assay (PF4-SRA) among others.

Phosphatidylserine externalization and procoagulant activation of erythrocytes induced by Pseudomonas aeruginosa virulence factor pyocyanin.

The opportunistic pathogen Pseudomonas aeruginosa causes a wide range of infections in multiple hosts by releasing an arsenal of virulence factors such as pyocyanin. Despite numerous reports on the pleiotropic cellular targets of pyocyanin toxicity in vivo, its impact on erythrocytes remains elusive. Erythrocytes undergo an apoptosis-like cell death called eryptosis which is characterized by cell shrinkage and phosphatidylserine (PS) externalization; this process confers a procoagulant phenotype on erythrocytes as well as fosters their phagocytosis and subsequent clearance from the circulation.

Producing megakaryocytes from a human peripheral blood source.

Background: Cultured megakaryocytes could prove useful in the study of human diseases, but it is difficult to produce sufficient numbers for study. We describe and evaluate the use of an expansion process to develop mature megakaryocytes from peripheral blood-derived human hematopoietic stem and progenitor cells (HSPCs).

Study Design And Methods: HSPCs (CD34+) were isolated from peripheral blood by positive selection and expanded using an optimal CD34+ expansion supplement.

Pitfalls in the diagnosis of heparin-Induced thrombocytopenia: A 6-year experience from a reference laboratory.

Heparin-induced thrombocytopenia (HIT) is caused by platelet-activating antibodies against complexes of platelet factor 4 (PF4) and heparin. The diagnosis of HIT is contingent on accurate and timely laboratory testing. Recently, alternative anticoagulants for the treatment of HIT have been introduced along with algorithms for better HIT diagnosis.

Congenital thrombotic thrombocytopenic purpura (cTTP) with two novel mutations.

Congenital thrombotic thrombocytopenic purpura (cTTP) is a rare disorder of childhood that has clinical and laboratory similarities to other, more common conditions. Prompt recognition is required as delays in therapy are associated with significant morbidity and failure to treat may lead to death. While the principles of treatment have not changed, enormous progress in the genetic and molecular understanding has taken place.

Studies of the immune response in heparin-induced thrombocytopenia.

Heparin-induced thrombocytopenia (HIT) is caused by platelet-activating antibodies that recognize PF4/heparin complexes. Uncertainties remain regarding HIT immunobiology, including the temporal relation of antibody formation to onset of thrombocytopenia, and whether immunoglobulin class switching occurs. Using serial plasma samples from 2 heparin thromboprophylaxis trials, we determined the time of onset, antibody levels, and immunoglobulin class distributions (IgG, IgA, IgM) for 12 patients with HIT and 36 patients who formed anti-PF4/heparin antibodies, but did not develop HIT ("seropositive non-HIT controls").

Laboratory testing for heparin-induced thrombocytopenia is inconsistent in North America: a survey of North American specialized coagulation laboratories.

Heparin-induced thrombocytopenia (HIT) is a serious complication of heparin therapy. As HIT is considered a clinico-pathologic entity, laboratory practices have an important role in diagnosing or excluding HIT. It was the objective of this study to assess the current status of laboratory testing for HIT in North America.

The prevalence of antibodies to the platelet factor 4 -heparin complex and association with access thrombosis in patients on chronic hemodialysis.

Introduction: Heparin-induced thrombocytopenia is a serious complication that can lead to thrombocytopenia, venous and arterial thrombosis. Patients with this disorder develop antibodies to the platelet factor 4-heparin (PF4-H) complex. Hemodialysis patients are repeatedly exposed to heparin and are at risk for developing PF4-H antibodies.

Laboratory testing for the antibodies that cause heparin-induced thrombocytopenia: how much class do we need?

Heparin-induced thrombocytopenia (HIT) is usually caused by platelet-activating antibodies of immunoglobulin G class that recognize platelet factor-4 (PF4) bound to heparin or certain other polyanions. Commercial enzyme immunoassays (EIAs) for PF4/polyanion-reactive antibodies detect two immunoglobulin classes (IgA and IgM) besides IgG. To investigate whether the additional detection of these antibody classes improves or worsens assay operating characteristics, we compared the sensitivity and specificity of EIAs that detect these 3 immunoglobulin classes individually with that of a commercial EIA (Genetic Testing Institute, GTI), as well as a platelet-activation assay, the serotonin-release assay (SRA).

Anti-platelet factor 4/heparin antibodies in orthopedic surgery patients receiving antithrombotic prophylaxis with fondaparinux or enoxaparin.

Heparin-induced thrombocytopenia (HIT) is caused by platelet-activating IgG antibodies that recognize platelet factor 4 (PF4) bound to heparin. Immunogenicity of heparins differs in that unfractionated heparin (UFH) induces more anti-PF4/heparin antibodies than low-molecular-weight heparin (LMWH) and UFH also causes more HIT. Fondaparinux, a synthetic anticoagulant modeled after the antithrombin-binding pentasaccharide, is believed to be nonimmunogenic.

Dissociation between the level of von Willebrand factor-cleaving protease activity and disease in a patient with congenital thrombotic thrombocytopenic purpura.

Decreased von Willebrand factor (VWF)-cleaving protease activity (<5%) has been implicated in patients with congenital thrombotic thrombocytopenic purpura-hemolytic uremic syndrome (Upshaw-Schulman syndrome) and associated with mutations within the ADAMTS13 gene. In this report, we describe longitudinal studies in a patient with congenital TTP who ultimately developed end-stage renal failure and required plasma therapy from infancy. The patient was deficient in plasma high molecular weight (HMW)-VWF multimers during acute disease but had increased amounts of the HMW-VWF multimers during periods of remission.

Gastrointestinal bleeding, angiodysplasia, cardiovascular disease, and acquired von Willebrand syndrome.

Recent evidence points to isolated deficiency of the largest multimers of von Willebrand factor (VWF)-known as von Willebrand syndrome type 2A (VWS-2A)-as a risk factor for bleeding from gastrointestinal (GI) angiodysplasia. This disorder is not widely recognized, perhaps because most patients do not exhibit generalized hemostatic impairment (bleeding is generally restricted to GI angiodysplasia) and because all but the largest multimers of VWF remain detectable in the plasma (thus, routine screening tests for VWS-2A are usually normal). The "Rosetta stone" for elucidating this syndrome was the enigma of Heyde's syndrome (aortic stenosis plus bleeding GI angiodysplasia), particularly the striking observation that aortic valve replacement generally cures GI bleeding and that preoperative deficiency of the largest VWF multimers undergoes long-term normalization after valve replacement.

The IgG subclasses of platelet-associated autoantibodies directed against platelet glycoproteins IIb/IIIa in patients with idiopathic thrombocytopenic purpura.

The majority of patients with idiopathic thrombocytopenic purpura (ITP) have antiplatelet autoantibodies that are most frequently directed against platelet glycoproteins IIb/IIIa or Ib/IX/V. However, there is some debate whether the immune response is oligoclonal or polyclonal in nature. We investigated the subclass distribution of anti-IIb/IIIa IgG autoantibodies in 59 prospectively studied patients with ITP.