The particle gel immunoassay as a rapid test to rule out heparin-induced thrombocytopenia? (original) (raw)
Transfusion, 1994
Background: As clinical diagnosis of heparin-associated thrombocytopenia (HAT) is often difficult, confirmation by sensitive laboratory assays is desirable. Stud Design and Methods: The sensitivity of the heparin-induced platelet activation (LIPA) test and the platelet aggregation test (PAT) was prospectively compared by using the sera of 209 patients with the putative diagnosis of HAT. Both assays were performed concomitantly with platelets of the same four donors using a different combination of donors from day to day. Further, all sera were assessed with a platelet factor 4 (PF4)/heparin enzyme-linked immunosorbent assay (ELISA). Results: Positive results were obtained with 33 percent of sera in the PFWheparin ELISA, with 33.5 percent of sera in the HlPA test, and with 11.5 percent of sera in the PAT. The PF4/heparin ELISA and the HlPA test showed no difference in sensitivity (p = 0.27 by McNemar's test) and were more sensitive than PAT (~4 0 " by McNemar's test). However, they recognized different patient cohorts. Nine HIPAindeterminate and 12 HIPA-negative sera were positive in the PFWheparin ELISA. Eight of the nine indeterminate sera caused platelet activation with high he arin concentrations in the HlPA test. Eleven of the 12 negative sera contained no LG, but 9 contained I M and 2 contained IgA HAT antibodies. Four sera that were indeterminate in the bF4,heparin ELISA and 18 sera that were negative were positive in the HlPA test. None of the sera that were positive in the PAT was missed in the HlPA test, but two of those were negative in the PFUheparin ELISA. All sera were assessed with four low-molecular-weight heparins and a low-molecular-weight heparinoid in the HlPA test with platelets from the same four donors. Low-molecular-weight heparin caused platelet activation with positive sera in 98 percent of tests, and the heparinoid did so in 10 percent; in a further 12.8 percent, crossreactivity to the low-molecular-weight heparinoid could not be excluded. Conclusion: The majority of HAT antibodies react with a PFWheparin complex, but there is stron evidence that other antigens are involved in some patients.The HlPA test and the jF4iheparin ELSA are sensitive for diagnosing HAT, and they complement one another. TRANSFUSION 1994;34:381-385. Abbreviations: ELSA = enzyme-linked lmmunosorbent assay; HAT = heparln-assoclated thrombocytopenia; HIPA = heparln-induced platelet activation (test); LMW = low molecular weight; PAT = platelet aggregation test; PF4 = platelet factor 4; SRA = serotonin-release assay.
Thrombosis Research, 2011
Background: The in vitro demonstration of antibodies against platelet factor-4/heparin (PF4/hep) complexes is an important contribution to the diagnosis of heparin-induced thrombocytopenia (HIT). The use of PF4/hep IgG-specific immunoassays enhances the specificity of HIT-investigations without any impairment of the sensitivity. Several IgG-specific immunoassays with different origin and structure of the target antigencomplex are commercially available. Methods: Using a retrospective cohort consisting of 459 patients suspected to have HIT, we compared the performance characteristics of two commercially available IgG-specific immunoassays, GTI-(Genetic Testing Institute) and HIA-IgG-ELISA (Hyphen Biomed Research). Results: PF4/hep antibodies were detected in 85 and 81 sera using GTI-and HIA-IgG-ELISA, respectively. OD values and clinical likelihood of patients who tested positive in one assay only were significantly lower than in those who tested positive in both immunoassays. Both IgG-specific assays showed high negative predictive values (100%) and similar but unsatisfactory positive predictive values, determined by a minimum clinical score of 5 and a positive HIPA result (41% and 43%, respectively). The implementation of a confirmatory step using excessive heparin increased the PPV of both assays, but results in a reduction of NPV in HIA-IgG-ELISA. Conclusions: The detection of IgG antibodies alone improves the clinical usefulness of immunoassays. However, functional assays remain indispensable to avoid the overdiagnosis of HIT caused by the detection of IgG non-platelet activating antibodies. The OD value in IgG immunoassays appears to correlate with the clinical relevance of the antibodies and might be used as a predictive parameter in the assessment of HIT.
…, 1994
Background: As clinical diagnosis of heparin-associated thrombocytopenia (HAT) is often difficult, confirmation by sensitive laboratory assays is desirable. Stud Design and Methods: The sensitivity of the heparin-induced platelet activation (LIPA) test and the platelet aggregation test (PAT) was prospectively compared by using the sera of 209 patients with the putative diagnosis of HAT. Both assays were performed concomitantly with platelets of the same four donors using a different combination of donors from day to day. Further, all sera were assessed with a platelet factor 4 (PF4)/heparin enzyme-linked immunosorbent assay (ELISA). Results: Positive results were obtained with 33 percent of sera in the PFWheparin ELISA, with 33.5 percent of sera in the HlPA test, and with 11.5 percent of sera in the PAT. The PF4/heparin ELISA and the HlPA test showed no difference in sensitivity (p = 0.27 by McNemar's test) and were more sensitive than PAT (~4 0 " by McNemar's test). However, they recognized different patient cohorts. Nine HIPAindeterminate and 12 HIPA-negative sera were positive in the PFWheparin ELISA. Eight of the nine indeterminate sera caused platelet activation with high he arin concentrations in the HlPA test. Eleven of the 12 negative sera contained no LG, but 9 contained I M and 2 contained IgA HAT antibodies. Four sera that were indeterminate in the bF4,heparin ELISA and 18 sera that were negative were positive in the HlPA test. None of the sera that were positive in the PAT was missed in the HlPA test, but two of those were negative in the PFUheparin ELISA. All sera were assessed with four low-molecular-weight heparins and a low-molecular-weight heparinoid in the HlPA test with platelets from the same four donors. Low-molecular-weight heparin caused platelet activation with positive sera in 98 percent of tests, and the heparinoid did so in 10 percent; in a further 12.8 percent, crossreactivity to the low-molecular-weight heparinoid could not be excluded. Conclusion: The majority of HAT antibodies react with a PFWheparin complex, but there is stron evidence that other antigens are involved in some patients.The HlPA test and the jF4iheparin ELSA are sensitive for diagnosing HAT, and they complement one another. TRANSFUSION 1994;34:381-385. Abbreviations: ELSA = enzyme-linked lmmunosorbent assay; HAT = heparln-assoclated thrombocytopenia; HIPA = heparln-induced platelet activation (test); LMW = low molecular weight; PAT = platelet aggregation test; PF4 = platelet factor 4; SRA = serotonin-release assay.
Cureus, 2018
Heparin-induced thrombocytopenia (HIT) is an adverse reaction to the administration of heparin due to the activation of the platelets by the immunoglobulin G (IgG) antibody-platelet factor 4 (PF4)/heparin immune complex. Since the clinical outcome is uncertain (as it could be associated with significant morbidity and sometimes death), an early diagnosis and appropriate treatment are necessary. The 4Ts pretest clinical scoring system and testing for all anti-PF4/heparin antibodies can markedly improve the diagnosis and prompt adequate treatment. Our study was undertaken to retrospectively evaluate the appropriateness of ordering the PF4 enzyme-linked immunosorbent assay (ELISA) test by using the 4Ts scoring system in a tertiary institution. We examined a database of 118 patients who had the PF4 ELISA test and calculated their 4Ts scores retrospectively. A total of 107 patients were evaluated; 95 patients (88.79%) had a negative PF4 ELISA assay and 12 patients tested positive (11.21%). Only one patient tested weakly positive in the low probability group (negative predictive value 98%). In the intermediate group, six patients were strongly positive (optical density (OD) > 1.0). In this latter group, further confirmatory testing using serotonin release assays (SRAs) could have been done. We also evaluated the setting where the tests were performed and found that the majority of patients (63.55%) were tested in the intensive care unit (ICU) where thrombocytopenia is multifactorial. We concluded that the large majority of patients were not appropriately evaluated prior to testing, which incurred unnecessary expense and patient distress. For the proper identification of patients suspected of HIT who should undergo PF4/heparin antibody testing, further education of the ordering physicians is recommended.
Journal of Thrombosis and …, 2004
The natural history of heparin-induced thrombocytopenia (HIT) in the absence of thrombosis was previously established using functional assays for confirmation of diagnosis (e.g. 14 C serotonin release assay). An enzyme-linked immunosorbent assay (ELISA) that detects the presence of antibodies directed against the heparin-platelet factor-4 (PF4) complex has largely replaced functional assays in many medical centers. Although the ELISA is highly sensitive for detecting HIT antibodies, its usefulness for predicting thrombotic outcomes has not been clearly established. We performed a retrospective chart review of all hospitalized patients at a university hospital who tested seropositive for HIT by a commercial ELISA during 2001 and 2002. A total of 63 inpatients were identified as HIT positive by ELISA. Forty-eight patients had no apparent HIT-associated thrombosis at the time of HIT seropositivity (i.e. isolated HIT) and only one was treated prophylactically with a direct thrombin inhibitor. The 30-day thrombosis rate for patients with isolated HIT was 17% (eight of 48). Higher ELISA optical density (OD) measurements correlated significantly with thrombosis (1.41 ± 0.87 vs. 0.79 ± 0.46, P < 0.001). Patients with isolated HIT and an OD measurement of ‡ 1.0 demonstrated nearly a 6-fold increased risk of thrombosis compared with those with OD values between 0.4 and 0.99 (odds ratio 5.74, 95% confidence interval 1.73, 19.0; absolute rate of thrombosis, 36% vs. 9%, respectively, P ¼ 0.07). We conclude that in hospitalized patients with isolated HIT, the presence of heparin-PF4 antibodies detected by ELISA was associated with a significant risk of subsequent thrombosis and higher ELISA values were observed among patients suffering thrombotic events.
Journal of Clinical Investigation, 1994
Heparin-induced thrombocytopenia/thrombosis (HITP) is thought to be mediated by immunoglobulins that activate platelets in the presence of pharmacologic concentrations of heparin, but the molecular basis for this relatively common and often serious complication of heparin therapy has not been established. We found that plasma from each of 12 patients with HITP contained high titer (. 1:200) antibodies that reacted with immobilized complexes of heparin and platelet factor 4 (PF4), a heparin-binding protein contained in platelet alphagranules. Recombinant human PF4 behaved similarly to PF4 isolated from platelets in this assay system. Complexes formed at an apparent heparin/PF4 molecular ratio of -1:2 (fresh heparin) and -1:12 (outdated heparin) were most effective in binding antibody. Immune complexes consisting of PF4, heparin, and antibody reacted with resting platelets; this interaction was inhibited by a monoclonal antibody specific for the FcyRII receptor and by excess heparin. Human umbilical vein endothelial cells, known to express heparin-like glycosaminoglycan molecules on their surface, were recognized by antibody in the presence of PF4 alone; this reaction was inhibited by excess heparin, but not by anti-Fc'yRII. Antibodies reactive with heparin/PF4 were not found in normal plasma, but IgG and 1gM antibodies were detected at dilutions of 1:10 (IgG) and 1:50 (IgM) in 3 of 50 patients (6%) with other types of immune thrombocytopenia. These findings indicate that antibodies associated with HITP react with PF4 complexed with heparin in solution or with glycosaminoglycan molecules on the surface of endothelial cells and provide the basis for a new hypothesis to explain the development of thrombocytopenia with thrombosis or disseminated intravascular coagulation in patients sensitive to heparin. (J. Clin. Invest. 1994. 93:81-88.)
Cytometry, 2001
Background: Heparin-induced thrombocytopenia (HIT) is a possible complication of heparin therapy that can evolve with life-threatening thromboembolism, for which early diagnosis is essential. However, the specific laboratory approach to the diagnosis of HIT is still controversial. Methods: Sera from 13 patients with HIT, from 15 patients with non-HIT thrombocytopenia, and from 10 normal subjects were used to compare nonfunctional and functional methods to detect anti-heparin:PF-4 antibodies and platelet activation. We used three enzyme-linked immunosorbent assays (ELISAs) and the particle gel immunoassay as nonfunctional tests, and platelet aggregometry, CD62p (p-selectin) phenotypical expression, and Annexin V binding as functional assays. Results: CD62p expression was positive in 85% of the cases and Annexin V was positive in 40% of the HIT cases examined. Aggregometry gave variable results that depend strongly on the donor. Conclusion: Functional tests for platelet activation are more reliable for HIT diagnosis than the nonfunctional tests. We conclude that the phenotypical expression of p-selectin detected by flow cytometry on activated platelets appears to be a good functional marker for the diagnosis of HIT and its possible thromboembolic complications. Cytometry (Comm. Clin. Cytometry) 46:290–295, 2001. © 2001 Wiley-Liss, Inc.
Clinical and Applied Thrombosis-hemostasis, 2007
Objective: Heparin-induced thrombocytopenia (HIT) is a life threatening complication of heparin therapy, causing thrombosis. The aim of our study was to find out the frequencies of HIT antibody seroconversion and clinical HIT in Turkish medical patients on different forms of heparins. Materials and Methods: Our study included 61 patients who were on unfractionated heparin (UFH) (n: 37) and low molecular weight heparin (LMWH) (n: 24) therapies. The frequency of HIT antibody formation was determined by means of antigenic (ELISA), and functional assays (serotonin release assay-SRA). Results: The seroconversion rates in UFH and LMWH groups were found to be 18.9% and 4.1% (ELISA), and 8.1% and 4.1% (SRA), respectively. One patient (2.1%) on UFH therapy developed deep vein thrombosis. No thromboembolic event was observed in patients taking LMWH. Conclusion: Seroconversion rates by means of antigenic and functional assays and clinical HIT were more common in patients on UFH than patients on LMWH therapy. (Turk J Hematol 2009; 26: 171-5) Özet Amaç: Heparine bağlı trombositopeni (HİT) heparin tedavisinin tromboza neden olan, hayatı tehdit eden bir komplikasyonudur. Çalışmamızın amacı farklı heparin formları kullanan dahili Türk hastalarda HİT antikor serokonversiyonu ve klinik HİT sıklığının saptanmasıydı. Yöntem ve Gereçler: Çalışmamıza anfraksiyone heparin (AFH) (n: 37) ve düşük molekül ağırlıklı heparin (DMAH) (n: 24) tedavisi alan 61 hasta katıldı. HIT antikor oluşumu antijenik (ELISA) ve fonksiyonel (serotonin salınım testi-SRA) testler ile değerlendirildi.
Anticoagulation drugs - the Current State of the Art [Working Title]
Heparin-induced thrombocytopenia (HIT) is the most life-threatening adverse effect of heparin therapy and is provoked by the development of drug-dependent antibodies. It occurs more frequently in patients with cardiac or orthopedic surgery or severe circulatory diseases, and the risk depends on the patient pathological status. As heparin is an anticoagulant used for treating thrombotic events or their risk, this iatrogenic complication has a paradoxal effect as it can induce thromboembolic diseases, frequently associated to severe morbidity or fatal outcomes. Diagnosis involves clinical evaluation of disease probability and laboratory tools for testing the presence of heparin-dependent antibodies with immunoassays or their capability to activate platelets with functional assays. Antibodies developed when stoichiometric complexes of platelet factor 4 (PF4) with heparin are formed during therapy. In few cases non-platelet factor 4 antigens can be involved. Antibodies can remain asymptomatic, but pathogenicity occurs in the presence of high concentrations of IgG isotype antibodies, with high avidity: they target and activate platelets or endothelial cells exposing heparin-PF4 (HPF4) complexes and produce thrombocytopenia and sometimes thrombosis. Risk factors which favor the development of antibodies and their pathological effect are discussed. The present understanding of mechanisms underlying disease development and diagnostic strategies of this heparin adverse effect is presented.