Huynh A, Kelton JG, Arnold DM, Daka M, Nazy I.
Antibody epitopes in vaccine-induced immune thrombotic thrombocytopaenia.
Nature. 2021 Aug;596(7873):565-569. doi: 10.1038/s41586-021-03744-4. Epub 2021 Jul 7.
Abstract/Text
Vaccine-induced immune thrombotic thrombocytopaenia (VITT) is a rare adverse effect of COVID-19 adenoviral vector vaccines1-3. VITT resembles heparin-induced thrombocytopaenia (HIT) in that it is associated with platelet-activating antibodies against platelet factor 4 (PF4)4; however, patients with VITT develop thrombocytopaenia and thrombosis without exposure to heparin. Here we sought to determine the binding site on PF4 of antibodies from patients with VITT. Using alanine-scanning mutagenesis5, we found that the binding of anti-PF4 antibodies from patients with VITT (n = 5) was restricted to eight surface amino acids on PF4, all of which were located within the heparin-binding site, and that the binding was inhibited by heparin. By contrast, antibodies from patients with HIT (n = 10) bound to amino acids that corresponded to two different sites on PF4. Biolayer interferometry experiments also revealed that VITT anti-PF4 antibodies had a stronger binding response to PF4 and PF4-heparin complexes than did HIT anti-PF4 antibodies, albeit with similar dissociation rates. Our data indicate that VITT antibodies can mimic the effect of heparin by binding to a similar site on PF4; this allows PF4 tetramers to cluster and form immune complexes, which in turn causes Fcγ receptor IIa (FcγRIIa; also known as CD32a)-dependent platelet activation. These results provide an explanation for VITT-antibody-induced platelet activation that could contribute to thrombosis.
© 2021. The Author(s), under exclusive licence to Springer Nature Limited.
Hursting MJ, Pai PJ, McCracken JE, Hwang F, Suvarna S, Lokhnygina Y, Bandarenko N, Arepally GM.
Platelet factor 4/heparin antibodies in blood bank donors.
Am J Clin Pathol. 2010 Nov;134(5):774-80. doi: 10.1309/AJCPG0MNR5NGKNFX.
Abstract/Text
Platelet factor 4 (PF4)/heparin antibody, typically associated with heparin therapy, is reported in some heparin-naive people. Seroprevalence in the general population, however, remains unclear. We prospectively evaluated PF4/heparin antibody in approximately 4,000 blood bank donors using a commercial enzyme-linked immunosorbent assay for initial and then repeated (confirmatory) testing. Antibody was detected initially in 249 (6.6%; 95% confidence interval [CI], 5.8%-7.4%) of 3,795 donors and repeatedly in 163 (4.3%; 95% CI, 3.7%-5.0%) of 3,789 evaluable donors. "Unconfirmed" positives were mostly (93%) low positives (optical density [OD] = 0.40-0.59). Of 163 repeatedly positive samples, 116 (71.2%) were low positives, and 124 (76.1%) exhibited heparin-dependent binding. Predominant isotypes of intermediate to high seropositive samples (OD >0.6) were IgG (20/39 [51%]), IgM (9/39 [23%]), and indeterminate (10/39 [26%]). The marked background seroprevalence of PF4/heparin antibody (4.3%-6.6%) with the preponderance of low (and frequently nonreproducible) positives in blood donors suggests the need for further assay calibration, categorization of antibody level, and studies evaluating clinical relevance of "naturally occurring" PF4/heparin antibodies.
Warkentin TE.
Autoimmune Heparin-Induced Thrombocytopenia.
J Clin Med. 2023 Nov 3;12(21). doi: 10.3390/jcm12216921. Epub 2023 Nov 3.
Abstract/Text
Autoimmune thrombocytopenia (aHIT) is a severe subtype of heparin-induced thrombocytopenia (HIT) with atypical clinical features caused by highly pathological IgG antibodies ("aHIT antibodies") that activate platelets even in the absence of heparin. The clinical features of aHIT include: the onset or worsening of thrombocytopenia despite stopping heparin ("delayed-onset HIT"), thrombocytopenia persistence despite stopping heparin ("persisting" or "refractory HIT"), or triggered by small amounts of heparin (heparin "flush" HIT), most cases of fondaparinux-induced HIT, and patients with unusually severe HIT (e.g., multi-site or microvascular thrombosis, overt disseminated intravascular coagulation [DIC]). Special treatment approaches are required. For example, unlike classic HIT, heparin cessation does not result in de-escalation of antibody-induced hemostasis activation, and thus high-dose intravenous immunoglobulin (IVIG) may be indicated to interrupt aHIT-induced platelet activation; therapeutic plasma exchange may be required if high-dose IVIG is ineffective. Also, aHIT patients are at risk for treatment failure with (activated partial thromboplastin time [APTT]-adjusted) direct thrombin inhibitor (DTI) therapy (argatroban, bivalirudin), either because of APTT confounding (where aHIT-associated DIC and resulting APTT prolongation lead to systematic underdosing/interruption of DTI therapy) or because DTI inhibits thrombin-induced protein C activation. Most HIT laboratories do not test for aHIT antibodies, contributing to aHIT under-recognition.
Warkentin TE, Kelton JG.
Temporal aspects of heparin-induced thrombocytopenia.
N Engl J Med. 2001 Apr 26;344(17):1286-92. doi: 10.1056/NEJM200104263441704.
Abstract/Text
BACKGROUND: Heparin-induced thrombocytopenia is a relatively common antibody-mediated drug reaction. We studied the temporal relation between previous or current heparin therapy and the onset of heparin-induced thrombocytopenia.
METHODS: We examined the time between the start of heparin therapy and the onset of thrombocytopenia in 243 patients with serologically confirmed heparin-induced thrombocytopenia. We also investigated the persistence of circulating heparin-dependent antibodies by performing a platelet serotonin-release assay and an assay for antibodies against platelet factor 4. The outcome in seven patients who had previously had an episode of heparin-induced thrombocytopenia and were later treated again with heparin was also examined.
RESULTS: A fall in the platelet count beginning four or more days after the start of heparin therapy occurred in 170 of the 243 patients (70 percent); in these patients, a history of previous heparin treatment did not influence the timing of the onset of thrombocytopenia. In the remaining 73 patients (30 percent), the onset of thrombocytopenia was rapid (median time of onset, 10.5 hours after the start of heparin administration); all these patients had been treated with heparin within the previous 100 days. During recovery from thrombocytopenia, heparin-dependent antibodies in the serum fell to undetectable levels at a median of 50 to 85 days, depending on the assay performed. In the seven patients who were given heparin again after the disappearance of heparin-dependent antibodies, a new episode of heparin-induced thrombocytopenia did not occur.
CONCLUSIONS: Heparin-induced thrombocytopenia can begin rapidly in patients who have received heparin within the previous 100 days. Heparin-dependent antibodies do not invariably reappear with subsequent heparin use.
Warkentin TE, Kelton JG.
Delayed-onset heparin-induced thrombocytopenia and thrombosis.
Ann Intern Med. 2001 Oct 2;135(7):502-6. doi: 10.7326/0003-4819-135-7-200110020-00009.
Abstract/Text
BACKGROUND: Heparin-induced thrombocytopenia is a prothrombotic drug reaction caused by platelet-activating antibodies that recognize complexes of platelet factor 4 and heparin.
OBJECTIVE: To describe a syndrome termed delayed-onset heparin-induced thrombocytopenia, in which thrombocytopenia and thrombotic events begin 5 or more days after withdrawal of heparin.
DESIGN: Case series.
SETTING: Secondary and tertiary care hospitals.
PATIENTS: 12 patients who presented with serologically confirmed, delayed-onset heparin-induced thrombocytopenia, including 6 outpatients presenting after hospital discharge.
MEASUREMENTS: The platelet serotonin-release assay was used to measure IgG-induced heparin-dependent and heparin-independent platelet activation; an enzyme immunoassay that detects IgG against platelet factor 4-heparin complexes was also used.
RESULTS: Patients with delayed-onset heparin-induced thrombocytopenia presented with thrombocytopenia and associated thrombosis a mean of 9.2 days (range, 5 to 19 days) after stopping heparin therapy. Nine patients received additional heparin, with further decrease in platelet counts. Compared with controls, patients with delayed-onset heparin-induced thrombocytopenia had higher titers of IgG antibodies to platelet factor 4-heparin and greater IgG-induced heparin-dependent and heparin-independent platelet activation.
CONCLUSIONS: Delayed-onset heparin-induced thrombocytopenia should be suspected when patients present with thrombocytopenia and thrombosis up to 3 weeks after exposure to heparin. This syndrome could be caused by high titers of platelet-activating IgG induced by heparin.
Warkentin TE.
Clinical picture of heparin-induced thrombocytopenia (HIT) and its differentiation from non-HIT thrombocytopenia.
Thromb Haemost. 2016 Oct 28;116(5):813-822. doi: 10.1160/TH16-06-0435. Epub 2016 Sep 22.
Abstract/Text
HIT is an acquired antibody-mediated disorder strongly associated with thrombosis, including microthrombosis secondary to disseminated intravascular dissemination (DIC). The clinical features of HIT are reviewed from the perspective of the 4Ts scoring system for HIT, which emphasises its characteristic timing of onset of thrombocytopenia. HIT antibodies recognize multimolecular complexes of platelet factor 4 (PF4)/heparin. However, a subset of HIT sera recognise PF4 bound to platelet chondroitin sulfate; these antibodies activate platelets in vitro and in vivo even in the absence of heparin, thus explaining: delayed-onset HIT (where HIT begins or worsens after stopping heparin); persisting HIT (where HIT takes several weeks to recover); spontaneous HIT syndrome (a disorder clinically and serologically resembling HIT but without proximate heparin exposure); and fondaparinux-associated HIT (four distinct syndromes featuring thrombocytopenia that begins or worsens during treatment with fondaparinux), with a new patient case presented with ongoing thrombocytopenia (and fatal haemorrhage) during treatment of HIT with fondaparinux, with fondaparinux-dependent platelet activation induced by patient serum ("fondaparinux cross-reactivity"). Ironically, despite existence of fondaparinux-associated HIT, this pentasaccharide anticoagulant is a frequent treatment for HIT (including one used by the author). HIT can be confused with other disorders, including those with a) timing similar to HIT (e. g. abciximab-associated thrombocytopenia of delayed-onset); b) combined thrombocytopenia/thrombosis (e. g. symmetrical peripheral gangrene secondary to acute DIC and shock liver); and c) both timing of onset and thrombosis (e. g. warfarin-associated venous limb gangrene complicating cancer-associated DIC). By understanding clinical and pathophysiological similarities and differences between HIT and non-HIT mimicking disorders, the clinician is better able to make the correct diagnosis.
Mian H, Warkentin TE, Sheppard JI, MacDonald A, Linkins LA, Benger A, Foley R.
Autoimmune HIT due to apheresis catheter heparin flushes for stem cell harvesting before autotransplantation for myeloma.
Blood. 2017 Oct 5;130(14):1679-1682. doi: 10.1182/blood-2017-06-788679. Epub 2017 Aug 22.
Abstract/Text
Warkentin TE.
Heparin-induced thrombocytopenia: pathogenesis and management.
Br J Haematol. 2003 May;121(4):535-55. doi: 10.1046/j.1365-2141.2003.04334.x.
Abstract/Text
Warkentin TE, Sheppard JA, Manheim JC.
HIT complicating fondaparinux prophylaxis: fondaparinux-dependent platelet activation as a marker for fondaparinux-induced HIT.
Thromb Haemost. 2014 Dec;112(6):1319-22. doi: 10.1160/TH14-08-0711. Epub 2014 Oct 16.
Abstract/Text
Warkentin TE, Makris M, Jay RM, Kelton JG.
A spontaneous prothrombotic disorder resembling heparin-induced thrombocytopenia.
Am J Med. 2008 Jul;121(7):632-6. doi: 10.1016/j.amjmed.2008.03.012.
Abstract/Text
BACKGROUND: Antibodies against the "self" protein, platelet factor 4 (PF4), bound to heparin-the cause of immune heparin-induced thrombocytopenia-are believed invariably to be triggered by preceding heparin therapy. We describe a novel syndrome, spontaneous heparin-induced thrombocytopenia, in which clinical and serologic features characteristic of this adverse drug reaction develop in patients despite the absence of preceding heparin therapy.
METHODS: Three patients met the study criteria (clinical and serologic features of heparin-induced thrombocytopenia without preceding heparin exposure), of whom 2 patients were identified among 225 patients (0.89%, 95% confidence interval, 0.11%-3.17%) with serologically confirmed heparin-induced thrombocytopenia recognized during an 18-year period at 1 hospital. The platelet serotonin-release assay was used to detect heparin-dependent immunoglobulin G-induced platelet activation, and 2 enzyme immunoassays were used to detect antibodies against PF4/heparin.
RESULTS: Two patients presented with thrombocytopenia and multiple arterial thrombosis, and 1 patient presented with anaphylactoid reactions after 2 subcutaneous injections of low-molecular-weight heparin. All 3 patients had high levels of platelet-activating anti-PF4/heparin antibodies of immunoglobulin G class at presentation despite the absence of previous heparin exposure. However, each patient did have a preceding infectious or inflammatory event; 1 patient had concomitant antiphospholipid antibodies.
CONCLUSION: Circumstances other than heparin use can trigger a spontaneous disorder that closely mimics heparin-induced thrombocytopenia, further supporting the autoimmune nature of this adverse drug reaction.
Greinacher A, Thiele T, Warkentin TE, Weisser K, Kyrle PA, Eichinger S.
Thrombotic Thrombocytopenia after ChAdOx1 nCov-19 Vaccination.
N Engl J Med. 2021 Jun 3;384(22):2092-2101. doi: 10.1056/NEJMoa2104840. Epub 2021 Apr 9.
Abstract/Text
BACKGROUND: Several cases of unusual thrombotic events and thrombocytopenia have developed after vaccination with the recombinant adenoviral vector encoding the spike protein antigen of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) (ChAdOx1 nCov-19, AstraZeneca). More data were needed on the pathogenesis of this unusual clotting disorder.
METHODS: We assessed the clinical and laboratory features of 11 patients in Germany and Austria in whom thrombosis or thrombocytopenia had developed after vaccination with ChAdOx1 nCov-19. We used a standard enzyme-linked immunosorbent assay to detect platelet factor 4 (PF4)-heparin antibodies and a modified (PF4-enhanced) platelet-activation test to detect platelet-activating antibodies under various reaction conditions. Included in this testing were samples from patients who had blood samples referred for investigation of vaccine-associated thrombotic events, with 28 testing positive on a screening PF4-heparin immunoassay.
RESULTS: Of the 11 original patients, 9 were women, with a median age of 36 years (range, 22 to 49). Beginning 5 to 16 days after vaccination, the patients presented with one or more thrombotic events, with the exception of 1 patient, who presented with fatal intracranial hemorrhage. Of the patients with one or more thrombotic events, 9 had cerebral venous thrombosis, 3 had splanchnic-vein thrombosis, 3 had pulmonary embolism, and 4 had other thromboses; of these patients, 6 died. Five patients had disseminated intravascular coagulation. None of the patients had received heparin before symptom onset. All 28 patients who tested positive for antibodies against PF4-heparin tested positive on the platelet-activation assay in the presence of PF4 independent of heparin. Platelet activation was inhibited by high levels of heparin, Fc receptor-blocking monoclonal antibody, and immune globulin (10 mg per milliliter). Additional studies with PF4 or PF4-heparin affinity purified antibodies in 2 patients confirmed PF4-dependent platelet activation.
CONCLUSIONS: Vaccination with ChAdOx1 nCov-19 can result in the rare development of immune thrombotic thrombocytopenia mediated by platelet-activating antibodies against PF4, which clinically mimics autoimmune heparin-induced thrombocytopenia. (Funded by the German Research Foundation.).
Copyright © 2021 Massachusetts Medical Society.
Salih F, Kohler S, Schönborn L, Thiele T, Greinacher A, Endres M.
Early recognition and treatment of pre-VITT syndrome after adenoviral vector-based SARS-CoV-2 vaccination may prevent from thrombotic complications: review of published cases and clinical pathway.
Eur Heart J Open. 2022 May;2(3):oeac036. doi: 10.1093/ehjopen/oeac036. Epub 2022 May 16.
Abstract/Text
Vaccine-induced immune thrombotic thrombocytopenia (VITT) is a rare but highly morbid complication after adenoviral vector-based SARS-CoV-2 vaccination. The pre-VITT syndrome is defined as vaccine-induced immune thrombocytopenia without thrombosis typically presenting with new-onset headache. This review aims to identify at-risk patients before complications such as cerebral venous sinus thrombosis occur. We review previously published reports of 19 patients (median age 35 years, range 23-74; 16 females) who met the diagnostic criteria for a pre-VITT syndrome. Seven patients progressed to VITT, 12 patients did not. Patients who experienced VITT received delayed treatment. The median interval between the onset of headache and VITT-treatment (i.e. anticoagulation, immune globulins, or corticosteroids) was 5 days (range 1-8 days) compared with 2 days (0-5 days) in those without subsequent VITT (P = 0.033). The interval from onset of headache to anticoagulation was longer in patients with VITT (median 7 vs. 2 days; range 3-9 vs. 0-7 days; P = 0.01). Anticoagulation was safe in all patients with a pre-VITT syndrome as no haemorrhagic complications occurred after anticoagulation was started despite low platelets. The transient decline of platelet count after admission was significantly more pronounced in patients who progressed to VITT (median 67 vs. 0 × 103/µL; range 0-77 × 103/µL vs. 0-10 × 103/µL; P = 0.005). d-dimers did not differ between groups. Pre-VITT syndrome is a 'red flag' and allows to identify and preemptively treat patients at-risk of further progression to VITT. However, it must be distinguished from post-vaccination immune thrombocytopenia.
© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology.
See I, Lale A, Marquez P, Streiff MB, Wheeler AP, Tepper NK, Woo EJ, Broder KR, Edwards KM, Gallego R, Geller AI, Jackson KA, Sharma S, Talaat KR, Walter EB, Akpan IJ, Ortel TL, Urrutia VC, Walker SC, Yui JC, Shimabukuro TT, Mba-Jonas A, Su JR, Shay DK.
Case Series of Thrombosis With Thrombocytopenia Syndrome After COVID-19 Vaccination-United States, December 2020 to August 2021.
Ann Intern Med. 2022 Apr;175(4):513-522. doi: 10.7326/M21-4502. Epub 2022 Jan 18.
Abstract/Text
BACKGROUND: Thrombosis with thrombocytopenia syndrome (TTS) is a potentially life-threatening condition associated with adenoviral-vectored COVID-19 vaccination. It presents similarly to spontaneous heparin-induced thrombocytopenia. Twelve cases of cerebral venous sinus thrombosis after vaccination with the Ad26.COV2.S COVID-19 vaccine (Janssen/Johnson & Johnson) have previously been described.
OBJECTIVE: To describe surveillance data and reporting rates of all reported TTS cases after COVID-19 vaccination in the United States.
DESIGN: Case series.
SETTING: United States.
PATIENTS: Case patients receiving a COVID-19 vaccine from 14 December 2020 through 31 August 2021 with thrombocytopenia and thrombosis (excluding isolated ischemic stroke or myocardial infarction) reported to the Vaccine Adverse Event Reporting System. If thrombosis was only in an extremity vein or pulmonary embolism, a positive enzyme-linked immunosorbent assay for antiplatelet factor 4 antibodies or functional heparin-induced thrombocytopenia platelet test result was required.
MEASUREMENTS: Reporting rates (cases per million vaccine doses) and descriptive epidemiology.
RESULTS: A total of 57 TTS cases were confirmed after vaccination with Ad26.COV2.S (n = 54) or a messenger RNA (mRNA)-based COVID-19 vaccine (n = 3). Reporting rates for TTS were 3.83 per million vaccine doses (Ad26.COV2.S) and 0.00855 per million vaccine doses (mRNA-based COVID-19 vaccines). The median age of patients with TTS after Ad26.COV2.S vaccination was 44.5 years (range, 18 to 70 years), and 69% of patients were women. Of the TTS cases after mRNA-based COVID-19 vaccination, 2 occurred in men older than 50 years and 1 in a woman aged 50 to 59 years. All cases after Ad26.COV2.S vaccination involved hospitalization, including 36 (67%) with intensive care unit admission. Outcomes of hospitalizations after Ad26.COV2.S vaccination included death (15%), discharge to postacute care (17%), and discharge home (68%).
LIMITATIONS: Underreporting and incomplete case follow-up.
CONCLUSION: Thrombosis with thrombocytopenia syndrome is a rare but serious adverse event associated with Ad26.COV2.S vaccination. The different demographic characteristics of the 3 cases reported after mRNA-based COVID-19 vaccines and the much lower reporting rate suggest that these cases represent a background rate.
PRIMARY FUNDING SOURCE: Centers for Disease Control and Prevention.
Johansen S, Laegreid IJ, Ernstsen SL, Azrakhsh NA, Kittang AO, Lindås R, Gjertsen BT, Vetti N, Mørtberg TV, Sørvoll IH, Holme PA, Ahlen MT, Reikvam H.
Thrombosis and thrombocytopenia after HPV vaccination.
J Thromb Haemost. 2022 Mar;20(3):700-704. doi: 10.1111/jth.15604. Epub 2021 Dec 7.
Abstract/Text
BACKGROUND: Vaccine-induced immune thrombotic thrombocytopenia (VITT) has so far only been reported after adenovirus vector severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccines.
OBJECTIVE: We report findings in a 25-year-old woman who presented with thrombocytopenia, venous thrombosis, elevated D-dimer levels, and high levels of platelet-activating antibodies to platelet factor 4-polyanion complexes 10 days after Gardasil 9 vaccination for human papillomavirus (HPV). The patient exhibited clinical and laboratory features in line with the recently defined VITT syndrome, described after adenoviral vector vaccination to prevent coronavirus disease 2019.
CONCLUSION: We report a case of VITT following HPV vaccination. This should raise awareness of the possibility of VITT also occurring after other vaccines, not exclusively adenoviral vector-based SARS-CoV-2 vaccines.
© 2021 The Authors. Journal of Thrombosis and Haemostasis published by Wiley Periodicals LLC on behalf of International Society on Thrombosis and Haemostasis.
Cuker A, Gimotty PA, Crowther MA, Warkentin TE.
Predictive value of the 4Ts scoring system for heparin-induced thrombocytopenia: a systematic review and meta-analysis.
Blood. 2012 Nov 15;120(20):4160-7. doi: 10.1182/blood-2012-07-443051. Epub 2012 Sep 18.
Abstract/Text
The 4Ts is a pretest clinical scoring system for heparin-induced thrombocytopenia (HIT). Although widely used in clinical practice, its predictive value for HIT in diverse settings and patient populations is unknown. We performed a systematic review and meta-analysis to estimate the predictive value of the 4Ts in patients with suspected HIT. We searched PubMed, Cochrane Database, and ISI Web of Science for studies that included patients with suspected HIT, who were evaluated by both the 4Ts and a reference standard against which the 4Ts could be compared. Quality of eligible studies was assessed by QUADAS-2 criteria. Thirteen studies, collectively involving 3068 patients, fulfilled eligibility criteria. A total of 1712 (55.8%) patients were classified by 4Ts score as having a low probability of HIT. The negative predictive value of a low probability 4Ts score was 0.998 (95% CI, 0.970-1.000) and remained high irrespective of the party responsible for scoring, the prevalence of HIT, or the composition of the study population. The positive predictive value of an intermediate and high probability 4Ts score was 0.14 (0.09-0.22) and 0.64 (0.40-0.82), respectively. A low probability 4Ts score appears to be a robust means of excluding HIT. Patients with intermediate and high probability scores require further evaluation.
Platton S, Bartlett A, MacCallum P, Makris M, McDonald V, Singh D, Scully M, Pavord S.
Evaluation of laboratory assays for anti-platelet factor 4 antibodies after ChAdOx1 nCOV-19 vaccination.
J Thromb Haemost. 2021 Aug;19(8):2007-2013. doi: 10.1111/jth.15362. Epub 2021 Jul 5.
Abstract/Text
INTRODUCTION: Vaccine-induced immune thrombocytopenia and thrombosis (VITT) following ChAdOx1 nCOV-19 vaccine has been described, associated with unusual site thrombosis, thrombocytopenia, raised D-dimer, and high-titer immunoglobulin-G (IgG) class anti-platelet factor 4 (PF4) antibodies. Enzyme-linked immunosorbent assays (ELISA) have been shown to detect anti-PF4 in patients with VITT, but chemiluminescence assays do not reliably detect them. ELISA assays are not widely available in diagnostic laboratories, and, globally, very few laboratories perform platelet activation assays.
METHODS: Assays that are commercially available in the United Kingdom were evaluated for their ability to identify anti-PF4 antibodies in samples from patients with suspected VITT. Four IgG-specific ELISAs, two polyspecific ELISAs, and four rapid assays were performed on samples from 43 patients with suspected VITT from across the United Kingdom. Cases were identified after referral to the UK Expert Haematology Panel multidisciplinary team and categorized into unlikely, possible, or probable VITT.
RESULTS AND DISCUSSION: We demonstrated that the HemosIL AcuStar HIT-IgG, HemosIL HIT-Ab, Diamed PaGIA gel, and STic Expert assays have poor sensitivity for VITT in comparison to ELISA. Where these assays are used for heparin-induced thrombocytopenia (HIT) diagnosis, laboratories should ensure that requests for suspected VITT are clearly identified so that an ELISA is performed. No superiority of IgG-ELISAs over polyspecific ELISAs in sensitivity to VITT could be demonstrated. No single ELISA method detected all possible/probable VITT cases; if a single ELISA test is negative, a second ELISA or a platelet activation assay should be considered where there is strong clinical suspicion.
© 2021 International Society on Thrombosis and Haemostasis.
Warkentin TE, Sheppard JI, Smith JW, Arnold DM, Nazy I.
Timeline of heparin-induced thrombocytopenia seroconversion in serial plasma samples tested using an automated latex immunoturbidimetric assay.
Int J Lab Hematol. 2019 Aug;41(4):493-502. doi: 10.1111/ijlh.13031. Epub 2019 May 3.
Abstract/Text
INTRODUCTION: HIT is caused by platelet-activating IgG that recognize multimolecular PF4/heparin complexes. HIT antibodies are generally detectable by PF4-dependent enzyme immunoassay (EIA) and by platelet serotonin-release assay (SRA) at the beginning of the HIT-related platelet count fall. We determined whether an automated immunoassay for HIT, the latex immunoturbidimetric assay (LIA), also detects antibodies early during the course of HIT. The LIA was also used to evaluate a patient with putative SRA-negative HIT.
METHODS: We evaluated the timing and magnitude of LIA reactivity in serial plasma samples obtained from 19 SRA-positive patients (17 with abnormal platelet count changes indicating HIT; two with subclinical seroconversion) and one putative SRA-negative HIT patient, all obtained from patients who participated in a clinical trial of heparin thromboprophylaxis. We determined LIA status at the onset of the HIT-related platelet count fall.
RESULTS: The LIA was positive in all 19 SRA-positive patients (median value, 7.3 U/mL [range, 1.2-35.5]; cutoff, 1.0 U/mL); for all 13 evaluable patients for whom an informative plasma sample was available at (or shortly before) the onset of the HIT-related platelet count fall, LIA reactivity was positive. Heterogeneity in seroconversion using the LIA was observed; some patients exhibited gradual increases in reactivity, whereas other patients showed rapid increase in reactivity over a few days. The single clinical trial patient who met clinical-pathological criteria for "SRA-negative HIT" tested LIA-positive.
CONCLUSION: The LIA detects HIT antibodies at the beginning of the HIT-associated platelet count fall. The LIA was also positive in a patient with SRA-negative HIT.
© 2019 The Authors International Journal of Laboratory Hematology Published by John Wiley & Sons Ltd.
Bankova A, Andres Y, Horn MP, Alberio L, Nagler M.
Rapid immunoassays for diagnosis of heparin-induced thrombocytopenia: Comparison of diagnostic accuracy, reproducibility, and costs in clinical practice.
PLoS One. 2017;12(6):e0178289. doi: 10.1371/journal.pone.0178289. Epub 2017 Jun 8.
Abstract/Text
BACKGROUND: Immunoassays are crucial in the work-up of patients with suspected heparin-induced thrombocytopenia (HIT) and rapid tests have been recently developed. However, comparative data on diagnostic accuracy, reproducibility, and analytical costs of different immunoassays in clinical practice are limited.
METHODS: Samples of 179 consecutive patients evaluated for suspected HIT in clinical practice using a polyspecific enzyme-linked immunoabsorbent assay (GTI diagnostics; ELISA) and a rapid particle gel immunoassay (PaGIA), were additionally analysed with a IgG-specific chemiluminescent immunoassay (AcuStar HIT-IgG). Presence of HIT was defined as a positive functional heparin-induced platelet aggregation test. Diagnostic accuracy was determined for low, intermediate and high thresholds as previously established (ELISA: optical density 0.4, 1.3, and 2.0 respectively; PaGIA: positive/negative, titre of 4, titre of 32; AcuStar HIT-IgG: 1.0 U/ml, 2.8, 9.4) and reproducibility was assessed by repeated measurements. Costs of test determination were calculated taking reagents, controls, and working time of technicians according to Swiss health care system into account.
RESULTS: Data on PaGIA results were available for 171 patients (95.5%), ELISA for 144 patients (80.4%), and AcuStar HIT-IgG for 179 patients (100%). Sensitivity was above 95% for all assays at low and intermediate thresholds. Specificity increased with higher thresholds and was above 90% for all assays with intermediate and high thresholds. Specificity of AcuStar HIT-IgG (92.8%; 95% CI 87.7, 96.2) was significantly higher than PaGIA (83.0%; 95% CI 76.3, 88.5) and higher than ELISA (81.8%, 95% CI 74.2, 88.0) at low threshold (p<0.05). Reproducibility was adequate for all assays. Total costs per test were CHF 51.02 for ELISA, 117.70 for AcuStar HIT-IgG, and 83.13 for PaGIA.
CONCLUSIONS: We observed favourable diagnostic accuracy measures and a high reproducibility for PaGIA and AcuStar HIT-IgG. Implementation into 24-hours-service might improve patient care but the results must be confirmed in other settings and larger populations as well.
Tardy B, Lecompte T, Mullier F, Vayne C, Pouplard C.
Detection of Platelet-Activating Antibodies Associated with Heparin-Induced Thrombocytopenia.
J Clin Med. 2020 Apr 24;9(4). doi: 10.3390/jcm9041226. Epub 2020 Apr 24.
Abstract/Text
Heparin-induced thrombocytopenia (HIT) is a prothrombotic immune drug reaction caused by platelet-activating antibodies that in most instances recognize platelet factor 4 (PF4)/polyanion complexes. Platelet activation assays (i.e., functional assays) are more specific than immunoassays, since they are able to discern clinically relevant heparin-induced antibodies. All functional assays used for HIT diagnosis share the same principle, as they assess the ability of serum/plasma from suspected HIT patients to activate fresh platelets from healthy donors in the presence of several concentrations of heparin. Depending on the assay, donors' platelets are stimulated either in whole blood (WB), platelet-rich plasma (PRP), or in a buffer medium (washed platelets, WP). In addition, the activation endpoint studied varies from one assay to another: platelet aggregation, membrane expression of markers of platelet activation, release of platelet granules. Tests with WP are more sensitive and serotonin release assay (SRA) is considered to be the current gold standard, but functional assays suffer from certain limitations regarding their sensitivity, specificity, complexity, and/or accessibility. However, the strict adherence to adequate preanalytical conditions, the use of selected platelet donors and the inclusion of positive and negative controls in each run are key points that ensure their performances.
Warkentin TE, Kelton JG.
A 14-year study of heparin-induced thrombocytopenia.
Am J Med. 1996 Nov;101(5):502-7. doi: 10.1016/s0002-9343(96)00258-6.
Abstract/Text
PURPOSE: To determine the sites of thromboses (venous versus arterial circulation) that complicate the clinical course of immunemediated heparin-induced thrombocytopenia, and to determine the 30-day risk for thrombosis in patients who are initially recognized with isolated heparin-induced thrombocytopenia.
PATIENTS AND METHODS: We analyzed objectively documented thrombotic events that complicated the clinical course of 127 patients with serologically confirmed heparin-induced thrombocytopenia identified in one medical community over a 14-year period. We classified heparin-induced thrombocytopenia patients into two groups: patients recognized with heparin-induced thrombocytopenia only after a new thrombosis had occurred, and patients initially recognized with isolated heparin-induced thrombocytopenia. We determined the subsequent 30-day risk for thrombosis for the cohort of patients initially recognized with isolated thrombocytopenia.
RESULTS: Heparin-induced thrombocytopenia was associated with the development of new venous thrombotic events and arterial thrombotic events in 78 and 18 patients, respectively (ratio venous/arterial thrombosis = 4:1). Pulmonary embolism was the most common life-threatening thrombotic event, occurring in 25% of all patients. Approximately half of all heparin-induced thrombocytopenia patients were recognized only after they had a complicating thrombotic event. Of the remaining 62-patient cohort initially recognized with isolated thrombocytopenia, the subsequent 30-day risk of thrombosis was 52.8%. The risk of thrombosis did not differ whether the heparin had been discontinued alone or whether warfarin had been substituted for the heparin.
CONCLUSIONS: Venous thrombosis complicates heparin-induced thrombocytopenia more frequently than does arterial thrombosis. The high risk of thrombosis in patients initially recognized with isolated thrombocytopenia suggests that conventional management approaches require reappraisal.
Arachchillage DJ, Thachil J, Anderson JAM, Baker P, Poles A, Kitchen S, Laffan M; BSH Committee.
Diagnosis and management of heparin-induced thrombocytopenia: Third edition.
Br J Haematol. 2024 Feb;204(2):459-475. doi: 10.1111/bjh.19180. Epub 2023 Dec 28.
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