Heparin-induced Platelet Aggregation Vs Platelet Factor 4 Enzyme-Linked Immunosorbent Assay in the Diagnosis of Heparin-induced Thrombocytopenia-Thrombosis

Background: Previous studies have demonstrated optimized diagnostic accuracy in utilizing higher antiheparin–platelet factor 4 (PF4) enzyme-linked immunosorbent assay (ELISA) optical density (OD) thresholds for diagnosing heparin-induced thrombocytopenia (HIT). We describe the incidence of positive serotonin release assay (SRA) results, as well as performance characteristics, for antiheparin–PF4 ELISA thresholds ≥0.4, ≥0.8, and ≥1.0 OD units in the diagnosis of HIT at our institution. Methods: Following institutional review board approval, we conducted a single-center retrospective chart review on adult inpatients with a differential diagnosis of HIT evaluated by both antiheparin–PF4 ELISA and SRA from 2012 to 2014. The major endpoints were to assess incidence of positive SRA results, sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and accuracy at antiheparin–PF4 ELISA values ≥0.4 OD units when compared to values ≥0.8 and ≥1.0 OD units. Clinical characteristics, including demographics, laboratory values, clinical and safety outcomes, length of stay, and mortality, were collected. Results: A total of 140 patients with 140 antiheparin–PF4 ELISA and SRA values were evaluated, of which 23 patients were SRA positive (16.4%) and 117 patients were SRA negative (83.6%). We identified a sensitivity of 91.3% versus 82.6% and 73.9%, specificity of 61.5% versus 87.2% and 91.5%, PPV of 31.8% versus 55.9% and 63.0%, NPV of 97.3% versus 96.2% and 94.7%, and accuracy of 66.4% versus 86.4% and 88.6% at antiheparin–PF4 ELISA thresholds ≥0.4, ≥0.8, and ≥1.0 OD units, respectively. Conclusion: Our study suggests an increased antiheparin–PF4 ELISA threshold of 0.8 or 1.0 OD units enhances specificity, PPV, and accuracy while maintaining NPV with decreased sensitivity.

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Antibodies to Macromolecular Platelet Factor 4-Heparin Complexes in Heparin-induced Thrombocytopenia: a Study of 44 Cases

Thrombosis and Haemostasis ◽

10.1055/s-0038-1651670 ◽

1995 ◽

Vol 73 (01) ◽

pp. 021-028 ◽

Cited By ~ 189

Author(s):

J Amiral ◽

F Bridey ◽

M Wolf ◽

C Boyer-Neumann ◽

E Fressinaud ◽

...

Keyword(s):

Molecular Weight ◽

Platelet Aggregation ◽

Early Diagnosis ◽

Healthy Subjects ◽

Platelet Factor 4 ◽

Platelet Factor ◽

Heparin Induced Thrombocytopenia ◽

Sulphated Polysaccharides ◽

Major Interest ◽

Heparin Complexes

SummaryAs heparin-PF4 (H-PF4) complexes are the target for antibodies associated to heparin-induced thrombocytopenia (HIT), an ELISA has been developed and optimised for testing antibodies binding to H-PF4. This test was consistently negative in 50 healthy subjects (A492 <0.3) and 35 patients with other causes of thrombocytopenia (A492 <0.5). In contrast, 43 out of 44 HIT patients showed antibodies to H-PF4 (A492 = 1.70 ± 0.81) including 5 patients with a negative platelet aggregation test. In one patient with HIT, antibodies to H-PF4 were already present at day 7, whereas platelet counts dropped ≤ 100 × 109/l only at days 11–12. Surprisingly, among 41 patients under heparin for >7 days, 5 showed antibodies to H-PF4, without HIT. These findings underline the major interest of this ELISA for the early diagnosis of HIT. We also showed that LMWH as well as other sulphated polysaccharides can bind to HIT antibodies in the presence of PF4 and that their reactivity is dependent on the molecular weight and the sulphation grade. The mechanism for HIT involves platelet PF4 receptors which bind the macromolecular H-PF4 complexes formed in the presence of a well defined heparin/PF4 ratio.

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Measurement of platelet factor 4 and β-thromboglobulin by an enzyme-linked immunosorbent assay

Clinica Chimica Acta ◽

10.1016/0009-8981(88)90042-3 ◽

1988 ◽

Vol 175 (1) ◽

pp. 113-114 ◽

Cited By ~ 9

Author(s):

Hoyu Takahashi ◽

Noriko Yoshino ◽

Akira Shibata

Keyword(s):

Immunosorbent Assay ◽

Platelet Factor 4 ◽

Enzyme Linked Immunosorbent Assay ◽

Platelet Factor

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Mesenteric and coeliac occlusions following heparininduced thrombocytopenia after aortobiprofunda bypass surgery

Annals of The Royal College of Surgeons of England ◽

10.1308/147870810x476629 ◽

2010 ◽

Vol 92 (1) ◽

pp. e15-e17

Author(s):

Ewan Bigsby ◽

Arindam Chaudhuri ◽

Alastair Whiteway

Keyword(s):

Parenteral Nutrition ◽

Total Parenteral Nutrition ◽

Immunosorbent Assay ◽

Arterial Thrombosis ◽

Short Distance ◽

Bypass Surgery ◽

Enzyme Linked Immunosorbent Assay ◽

Bowel Ischaemia ◽

Platelet Factor ◽

Heparin Induced Thrombocytopenia

A 56-year-old man developed mid-gut bowel ischaemia following an elective aortobiprofunda bypass for short-distance claudication. The bowel was resected and he was commenced on lifelong total parenteral nutrition. He was found to have developed heparin-induced thrombocytopenia and thrombosis, confirmed by high levels of heparin-platelet factor 4-antibody on enzyme-linked immunosorbent assay (ELISA). He subsequently had foregut ischaemia with a second bout of thrombocytopenia despite not being on heparin. The case describes the first report of bowel ischaemia as a consequence of heparin-induced thrombocytopenia causing sequential superior mesenteric and coeliac arterial thrombosis in this scenario and highlights the importance of the awareness of the association of these pathological entities and subsequent management.

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The role of platelet factor 4 in platelet aggregation induced by the antibodies implicated in heparin-induced thrombocytopenia

Blood Coagulation & Fibrinolysis ◽

10.1097/00001721-200110000-00002 ◽

2001 ◽

Vol 12 (7) ◽

pp. 511-520 ◽

Cited By ~ 11

Author(s):

G. T. Gerotziafas ◽

I. Elalamy ◽

C. Lecrubier ◽

J. Lebrazi ◽

M. Mirshahi ◽

...

Keyword(s):

Platelet Aggregation ◽

Platelet Factor 4 ◽

Platelet Factor ◽

Heparin Induced Thrombocytopenia

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Heparin-Induced Thrombocytopenia: Prevalence in a Large Cohort of Patients and Confirmed Role of PF4/Heparin Complex as the Main Antigen for Antibodies

Clinical and Applied Thrombosis/Hemostasis ◽

10.1177/107602969700300309 ◽

1997 ◽

Vol 3 (3) ◽

pp. 203-209 ◽

Cited By ~ 3

Author(s):

Fabrizio Fabris ◽

Immacolata Cordiano ◽

Federica Salvan ◽

Leopoldo Saggin ◽

Giuseppe Cella ◽

...

Keyword(s):

Molecular Weight ◽

Platelet Count ◽

Platelet Factor 4 ◽

Enzyme Linked Immunosorbent Assay ◽

Low Molecular Weight ◽

Dependent Manner ◽

Platelet Factor ◽

Heparin Induced Thrombocytopenia ◽

Heparin Treatment ◽

Daily Dose

We performed a retrospective study on the prevalence of heparin-induced thrombocytopenia (HIT) in 233 patients receiving hog mucosa heparin therapy. Of these, 82 patients received s.c. calcium heparin, 130 patient received unfractionated (UF) i.v. heparin, and 21 patients received low molecular weight heparins (LMWH). An additional four patients, referred to our consultation and diagnosed by us as having clinically active type II HIT (HIT-II) were also studied. The mean platelet count of the 233 patients receiving heparin showed a significant decrease after 2 days of heparin treatment and a following significant increase 6 days later (basal: 257 ± 147 x 109 platelets/L; day 2: 239 ± 122, p < 0.0002; day 6: 286 ± 119, p < 0.004). Of the 212 patients receiving UF heparin, 13 (6%) fulfilled the criteria for HIT-II: seven of these had received i.v. heparin (mean daily dose 26,600 ± 4,082 IU ± SD) and six had received s.c. heparin (mean daily dose 21,428:t 6,900 IU). Their mean basal platelet count was 226 ± 100 SD × 109 platelets/L and the nadir during heparin treatment was 78 ± 39 x 10 9 platelets/L. Thrombotic complications occurred in four (30.7%) of the 13 patients with HIT-II. Since the immunological mechanism has been demonstrated for HIT-II and since platelet factor 4 (PF4) was identified as the co-factor for the binding of heparin-related antibodies, we set up our own enzyme-linked immunosorbent assay (ELISA) for testing antibodies against PF4/heparin complex bound through electrostatic bridges to the solid phase. The highest binding capacity of HIT-related IgG to the multimolecular complex was obtained at 20 μg/ml for PF4 and 3 μg/ml for heparin, corresponding to 250 ng of PF4 and 42 ng of heparin in each microtiter well. Such binding was inhibited in a dose-dependent manner by increasing amounts of heparin, protamine hydrochloride, and a monoclonal antibody anti-human PF4 clone 1OB2. We observed that HIT-related antibodies bound also to PF4/LMWH complexes but the optimal PF4/glycosaminoglycan ratio appeared more critical for LMWH (enoxaparin, fraxiparin, and pamaparin) than for UF heparin. Sera from eight patients with HIT-II were tested by PF4/heparin ELISA; six of these had IgG against the complex PF4/heparin and three also had IgM. The persistence of HIT-related antibodies was investigated in three patients: in one such antibodies were still detectable 3 years after the acute episode, while in the other two, they disappeared after 6 months and 1 year, respectively. Key Words: Heparin-related anti body—Platelet factor 4 (PF4)—Heparin—Low molecular weight heparin—Thrombocytopenia—Thrombosis.

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