Download PDF
Thromb Haemost 2012; 107(03): 575-583
DOI: 10.1160/TH11-09-0631
New Technologies, Diagnostic Tools and Drugs
Schattauer GmbH

Validation of whole blood impedance aggregometry as a new diagnostic tool for HIT

Results of a large Australian study
Marie-Christine Morel-Kopp
1   Northern Blood Research Centre, Kolling Institute of Medical Research, University of Sydney, Sydney, Australia
2   Department of Haematology and Transfusion Medicine, Royal North Shore Hospital, Sydney, Australia
,
Chee Wee Tan
1   Northern Blood Research Centre, Kolling Institute of Medical Research, University of Sydney, Sydney, Australia
2   Department of Haematology and Transfusion Medicine, Royal North Shore Hospital, Sydney, Australia
,
Timothy A Brighton
3   SEALS, Prince of Wales Hospital, Sydney, Australia
,
Simon McRae
4   Department of Haematology SA Pathology, Adelaide, Australia
,
Ross Baker
5   Department of Haematology, Royal Perth Hospital, Perth, Australia
,
Huyen Tran
6   Monash Medical Centre, Melbourne, Australia
,
Peter Mollee
7   Department of Haematology, Princess Alexandra Hospital, Brisbane, Australia
,
Geoffrey Kershaw
8   Royal Prince Alfred Hospital, Sydney, Australia
,
Joanne Joseph
9   Haematology Department, St Vincent Hospital, Sydney, Australia
,
Christopher Ward
1   Northern Blood Research Centre, Kolling Institute of Medical Research, University of Sydney, Sydney, Australia
2   Department of Haematology and Transfusion Medicine, Royal North Shore Hospital, Sydney, Australia
,
the ASTH Clinical Trials Group › Author Affiliations
Further Information

Publication History

Received: 13 September 2011

Accepted after major revision: 01 January 2011

Publication Date:
22 November 2017 (online)

Also available at
    Permissions and Reprints

Summary

Heparin-induced thrombocytopenia (HIT) remains a challenge, with diagnosis confirmed only by functional assays. The gold standard 14C-se-rotonin release assay (SRA) is highly sensitive but technically challenging and unsuitable for routine use. We conducted a large study to validate whole blood impedance aggregometry (WBIA) as a suitable diagnostic tool for HIT. WBIA and SRA were used to test 181 samples positive for H-PF4 antibodies by PaGIA or ELISA. Using the same high responder donor, 77 samples were positive by WBIA (aggregation with low-dose but not high-dose heparin). Using the strict definition for SRA positivity, 72 samples were true HIT. In nine samples, serotonin release with high-dose heparin dropped by > 50% but was still >20%; these were retested after a one-half dilution and 8/9 became positive. Ten other samples were discrepant between the two assays: one strongly positive (89% release) and six weakly positive samples by SRA (average release 56%) were WBIA negative. When these samples were retested using a random donor, only two remained SRA positive. Three samples were strongly WBIA positive but SRA negative; two were retested by SRA with 0.5IU/ml heparin and one became positive. Under controlled conditions, using the same selected high-responder donor, WBIA and SRA performed similarly with slightly increased sensitivity of the WBIA when using the strict definition of SRA positivity. WBIA is easy to perform with rapid turn-around time and warrants a multi-laboratory trial to complete its validation as a confirmatory assay for platelet-activating HIT antibodies.

Keywords

Heparin - heparin-induced thrombocytopenia - 4T′s score - serotonin release assay - whole blood impedance aggregometry
 

  • References

  • 1 Amiral J, Bridey F, Dreyfus M. et al. Platelet factor 4 complexed to heparin is the target for antibodies generated in heparin-induced thrombocytopenia. Thromb Haemost 1992; 68: 95-96.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 2 Amiral J, Bridey F, Wolf M. et al. Antibodies to macromolecular platelet factor 4-heparin complexes in heparin-induced thrombocytopenia: a study of 44 cases. Thromb Haemost 1995; 73: 21-28.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 3 Warkentin TE, Chong BH, Greinacher A. Heparin-induced thrombocytopenia: towards consensus. Thromb Haemost 1998; 79: 1-7.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 4 Davenport A. Heparin-induced thrombocytopenia during renal replacement therapy. Hemodial Int 2004; 08: 295-303.
    CrossrefPubMedGoogle Scholar
  • 5 Matsuo T, Wanaka K. Management of uremic patients with heparin-induced thrombocytopenia requiring hemodialysis. Clin Appl Thromb Hemost 2008; 14: 459-464.
    CrossrefPubMedGoogle Scholar
  • 6 Morel-Kopp MC, Aboud M, Tan CW. et al. Whole blood impedance aggrego-metry detects heparin-induced thrombocytopenia antibodies. Thromb Res 2010; 125: e234-239.
    CrossrefPubMedGoogle Scholar
  • 7 Warkentin TE. Heparin-induced thrombocytopenia: pathogenesis and management. Br J Haematol 2003; 121: 535-555.
    CrossrefPubMedGoogle Scholar
  • 8 Lo GK, Juhl D, Warkentin TE. et al. Evaluation of pretest clinical score (4 T’s) for the diagnosis of heparin-induced thrombocytopenia in two clinical settings. J Thromb Haemost 2006; 04: 759-765.
    CrossrefPubMedGoogle Scholar
  • 9 Worthington RE, Carroll RC, Boucheix C. Platelet activation by CD9 monoclonal antibodies is mediated by the Fc gamma II recepto.r. Br J Haematol 1990; 74: 216-222.
    CrossrefPubMedGoogle Scholar
  • 10 Morel MC, Lecompte T, Champeix P. et al. PL2-49, a monoclonal antibody against glycoprotein IIb which is a platelet activator. Br J Haematol 1989; 71: 57-63.
    CrossrefPubMedGoogle Scholar
  • 11 Warmerdam PA, van de Winkel JG, Gosselin EJ. et al. Molecular basis for a polymorphism of human Fc gamma receptor II (CD32). J Exp Med 1990; 172: 19-25.
    CrossrefPubMedGoogle Scholar
  • 12 Sheridan D, Carter C, Kelton JG. A diagnostic test for heparin-induced throm-bocytopenia. Blood 1986; 67: 27-30.
    CrossrefPubMedGoogle Scholar
  • 13 Lo GK, Sigouin CS, Warkentin TE. What is the potential for overdiagnosis of he-parin-induced thrombocytopenia?. Am J Hematol 2007; 82: 1037-1043.
    CrossrefPubMedGoogle Scholar
  • 14 Messmore HL, Fabbrini N, Bird ML. et al. Simple Scoring System for the Early Management of Heparin-Induced Thrombocytopenia. Clin Appl Thromb He-most 2011; 17: 197-201.
    CrossrefPubMedGoogle Scholar
  • 15 Pouplard C, Gueret P, Fouassier M. et al. Prospective evaluation of the ‘4Ts’ score and particle gel immunoassay specific to heparin/PF4 for the diagnosis of hepa-rin-induced thrombocytopenia. J Thromb Haemost 2007; 05: 1373-1379.
    CrossrefPubMedGoogle Scholar
  • 16 Bryant A, Low J, Austin S. et al. Timely diagnosis and management of heparin-in-duced thrombocytopenia in a frequent request, low incidence single centre using clinical 4T’s score and particle gel immunoassay. Br J Haematol 2008; 143: 721-726.
    CrossrefPubMedGoogle Scholar
  • 17 Cuker A. Heparin-induced thrombocytopenia: present and future. J Thromb Thrombolysis 2011; 31: 353-366.
    CrossrefPubMedGoogle Scholar
  • 18 Greinacher A, Selleng K. Thrombocytopenia in the intensive care unit patient. Hematology Am Soc Hematol Educ Program 2010; 2010: 135-143.
    CrossrefPubMedGoogle Scholar
  • 19 Price EA, Hayward CP, Moffat KA. et al. Laboratory testing for heparin-induced thrombocytopenia is inconsistent in North America: a survey of North American specialized coagulation laboratories. Thromb Haemost 2007; 98: 1357-1361.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 20 Bidet A, Tardy Poncet B, Desprez D. et al. Clinical characteristics and laboratory testing of patients with suspected HIT: a survey on current practice in 11 university hospitals in France. Thromb Res 2010; 125: e294-299.
    CrossrefPubMedGoogle Scholar
  • 21 Bakchoul T, Giptner A, Najaoui A. et al. Prospective evaluation of PF4/heparin immunoassays for the diagnosis of heparin-induced thrombocytopenia. J Thromb Haemost 2009; 103: 145-150.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 22 Schneiter S, Colucci G, Sulzer I. et al. Variability of anti-PF4/heparin antibody results obtained by the rapid testing system ID-H/PF4-PaGIA. J Thromb Haemost 2009; 07: 1649-1655.
    CrossrefPubMedGoogle Scholar
  • 23 Greinacher A, Kohlmann T, Strobel U. et al. The temporal profile of the anti-PF4/heparin immune response. Blood 2009; 113: 4970-4976.
    CrossrefPubMedGoogle Scholar
  • 24 Pouplard C, Leroux D, Regina S. et al. Effectiveness of a new immuno-assay for the diagnosis of heparin-induced thrombocytopenia and improved specificity when detecting IgG antibodies. Thromb Haemost 2010; 103: 145-150.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 25 Morel-Kopp MC, Aboud M, Ta n CW. et al. Heparin-induced thrombocytopenia: evaluation of IgG and IgGAM ELISA assays. Int J Lab Hematol 2011; 33: 245-250.
    CrossrefPubMedGoogle Scholar
  • 26 Whitlatch NL, Kong DF, Metjian AD. et al. Validation of the high-dose heparin confirmatory step for the diagnosis of heparin-induced thrombocytopenia. Blood 2010; 116: 1761-1766.
    CrossrefPubMedGoogle Scholar
  • 27 Bakchoul T, Giptner A, Bein G. et al. Performance characteristics of two commercially available IgG-specific immunoassays in the assessment of heparin-induced thrombocytopenia (HIT). Thromb Res 2011; 127: 345-348.
    CrossrefPubMedGoogle Scholar
  • 28 Fratantoni JC, Pollet R, Gralnick HR. Heparin-induced thrombocytopenia: confirmation of diagnosis with in vitro methods. Blood 1975; 45: 395-401.
    PubMedGoogle Scholar
  • 29 Chong BH, Grace CS, Rozenberg MC. Heparin-induced thrombocytopenia: effect of heparin platelet antibody on platelets. Br J Haematol 1981; 49: 531-540.
    CrossrefPubMedGoogle Scholar
  • 30 Greinacher A, Michels I, Kiefel V. et al. A rapid and sensitive test for diagnosing he-parin-associated thrombocytopenia. Thromb Haemost 1991; 66: 734-736.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 31 Lee DH, Warkentin TE, Denomme GA. et al. A diagnostic test for heparin-in-duced thrombocytopenia: detection of platelet microparticles using flow cyto-metry. Br J Haematol 1996; 95: 724-731.
    CrossrefPubMedGoogle Scholar
  • 32 Mullier F, Bailly N, Cornet Y. et al. Contribution of platelet microparticles generation assay to the diagnosis of type II heparin-induced thrombocytopenia. Thromb Haemost 2010; 103: 1277-1281.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 33 Warkentin TE, Hayward CP, Boshkov LK. et al. Sera from patients with heparin-induced thrombocytopenia generate platelet-derived microparticles with pro-coagulant activity: an explanation for the thrombotic complications of heparin-induced thrombocytopenia. Blood 1994; 84: 3691-3699.
    CrossrefPubMedGoogle Scholar
  • 34 Jy W, Mao WW, Horstman LL. et al. A flow cytometric assay of platelet activation marker P-selectin (CD62P) distinguishes heparin-induced thrombocytopenia (HIT) from HIT with thrombosis (HITT). Thromb Haemost 1999; 82: 1255-1259.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 35 Denys B, Stove V, Philippe J. et al. A clinical-laboratory approach contributing to a rapid and reliable diagnosis of heparin-induced thrombocytopenia. Thromb Res 2008; 123: 137-145.
    CrossrefPubMedGoogle Scholar
  • 36 Khairy M, Lasne D, Brohard-Bohn B. et al. A new a pproach in t he st udy of t he molecular and cellular events implicated in heparin-induced thrombocytopenia. Formation of leukocyte-platelet aggregates. Thromb Haemost 2001; 85: 1090-1096.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 37 Tomer A. A sensitive and specific functional flow cytometric assay for the diagnosis of heparin-induced thrombocytopenia. Br J Haematol 1997; 98: 648-656.
    CrossrefPubMedGoogle Scholar
  • 38 Tardy-Poncet B, Piot M, Chapelle C. et al. Thrombin generation and heparin-in-duced thrombocytopenia. J Thromb Haemost 2009; 07: 1474-1481.
    CrossrefPubMedGoogle Scholar
  • 39 Toth O, Calatzis A, Penz S. et al. Multiple electrode aggregometry: a new device to measure platelet aggregation in whole blood. Thromb Haemost 2006; 96: 781-788.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 40 Ockenhouse CF, Magowan C, Chulay JD. Activation of monocytes and platelets by monoclonal antibodies or malaria-infected erythrocytes binding to the CD36 surface receptor in vitro. J Clin Invest 1989; 84: 468-475.
    CrossrefPubMedGoogle Scholar
  • 41 Clark MR, Stuart SG, Kimberly RP. et al. A single amino acid distinguishes the high-responder from the low-responder form of Fc receptor II on human mono-cytes. Eur J Immunol 1991; 21: 1911-1916.
    CrossrefPubMedGoogle Scholar
  • 42 Tomiyama Y, Kunicki TJ, Zipf TF. et al. Response of human platelets to activating monoclonal antibodies: importance of Fc gamma RII (CD32) phenotype and level of expression. Blood 1992; 80: 2261-2268.
    PubMedGoogle Scholar
  • 43 Reilly AF, Norris CF, Surrey S. et al. Genetic diversity in human Fc receptor II for immunoglobulin G: Fc gamma receptor IIA ligand-binding polymorphism. Clin Diagn Lab Immunol 1994; 01: 640-644.
    PubMedGoogle Scholar
  • 44 Eichler P, Budde U, Haas S. et al. First workshop for detection of heparin-induced antibodies: validation of the heparin-induced platelet-activation test (HIPA) in comparison with a PF4/heparin ELISA. Thromb Haemost 1999; 81: 625-629.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 45 Otis SA, Zehnder JL. Heparin-induced thrombocytopenia: current status and diagnostic challenges. Am J Hematol 2010; 85: 700-706.
    CrossrefPubMedGoogle Scholar
  • 46 Pauzner R, Greinacher A, Selleng K. et al. False-positive tests for heparin-induced thrombocytopenia in patients with antiphospholipid syndrome and systemic lupus erythematosus. J Thromb Haemost 2009; 07: 1070-1074.
    CrossrefPubMedGoogle Scholar
  • 47 Warkentin TE. Antiphospholipid and anti-PF4 antibodies: an association affecting anti-PF4/heparin assay analysis. J Thromb Haemost 2009; 07: 1067-1069.
    CrossrefPubMedGoogle Scholar
  • 48 Regnault V, de Maistre E, Carteaux JP. et al. Platelet activation induced by human antibodies to interleukin-8. Blood 2003; 101: 1419-1421.
    CrossrefPubMedGoogle Scholar