Presenter: Marisa B. Marques, MD
Panelists: Dorothy Adcock-Funk, MD, Kandice Kottke-Marchant, MD, PhD, John D. Olson, MD, PhD
Moderator: George A. Fritsma, MS MT (ASCP)
Precision BioLogic Laboratory Medicine Roundtable, June 20 and 21, 2008, Dartmouth, Nova Scotia
I’d like to thank Wendy Porteous, Steve Duff, Michael Scott, and all the Precision BioLogic Inc. participants who organized our June round-table discussion. I’d especially like to acknowledge researchers Balwant Bhullar and Diane Jette who participated in this discussion.
Heparin-induced thrombocytopenia (HIT) is a menacing prothrombotic syndrome caused by antibodies to platelet factor 4 (PF4)-heparin complexes that develop during exposure to heparin, mostly commonly when it is used intravenously. IgG-isotype antibodies predominate and bind platelet membrane Fc receptors, activating them, causing thrombocytopenia and, in a minority of cases, potentially lethal venous and arterial thrombosis.
Patients receiving heparin are monitored daily for platelet count and evidence of thrombosis. A falling platelet count, skin necrosis, and new thrombotic events are used to generate a pretest HIT probability using the “four Ts” score developed at McMaster University by T. Warkentin MD, and colleagues:
4Ts 2 points 1 point 0 points
PLT count falls > 50%, PLT nadir ≥ 20,000/uL
PLT count falls 30–50% or PLT nadir 10–19,000/uL
PLT count falls < 30%, or PLT nadir < 10,000/uL
Timing of PLT count fall Clear onset between 5–10 days; ≤ 1 day when prior heparin exposure within 30 days Onset within 5–10 days but not clear, onset after day 10 or ≤ 1 day when heparin exposure is 30-100 days before Onset < 4 days without recent heparin exposure
Thrombosis or other sequelae New thrombosis (confirmed), skin necrosis, acute systemic reaction postintravenous UFH bolus Progressive, recurrent or suspected thrombosis, non-necrotizing erythematous skin lesions, None
Other causes for thrombocytopenia None apparent Possible
High: 6–8 Intermediate: 4–6 Low: ≤ 3
Lo GK, Juhl D, Warkentin TE, et al. Evaluation of pretest clinical score (4T’s) for the diagnosis of heparin-induced thrombocytopenia in two clinical settings. J Thromb Haemost 2006:4;759–65.
The reference laboratory method for detection of clinically significant antibodies that can cause HIT is considered to be the 3H-serotonin release assay (SRA), first developed in 1975 by Fratantoni, Pollet and Gralnick. Washed normal platelets are labeled with the isotope, incubated with serum from patients exposed to heparin who have developed thrombocytopenia and the release of radioactive serotonin measured as a marker of platelet activation. The SRA is available from several North American specialty reference laboratories, but is seldom offered by local institutions owing to the radioactive isotope license requirement.
Early HIT assays employed optical aggregometry of fresh normal platelets challenged by a therapeutic unfractionated heparin concentration agonist in patient plasma. These assays depended on platelet donor response to heparin secondary to platelet membrane Fc receptor concentration. Optical aggregometry was insensitive and is largely abandoned as a HIT screen. Other assays include platelet release measured by lumiaggregometry and Aniara’s more generalized ZYMUTEST MP-ACTIVITY® platelet microparticle assay.
Local laboratories may employ a colorimetric enzyme-linked immunosorbent assay (ELISA) coated with solid-phase target PF4/heparin or PF4/polyanion surrogate. PF4/target conjugated kits are available from Genetic Testing Institute (GTI), Inc, Brookfield, WI; Aniara Corp, Mason, OH; or Diagnostica Stago, Inc, Parsippany, NJ.
We compared GTI’s current PF4 IgG/M/A® kit with their recently released GTI PF4 IgG®, the SRA reference method from Quest Diagnostics Inc, Chantilly, VA; and 4T scores of patients who were referred for ELISA testing and had a positive result. We presented our results in the recent meeting of the Academy of Clinical Laboratory Physicians and Scientists and will publish our results in the near future. We used the GTI-recommended positivity threshold of 0.400 AU for both PF4 IgG/M/A and PF4 IgG. We learned…
The PF4 IgG/M/A assay overestimates the incidence of thrombosis compared with the clinical probability (positive ELISA and low 4T score).
The PF4 IgG assay is positive in approximately half of IgG/M/A positive cases.
The PF4 IgG assay is negative in a high percentage of high and intermediate 4T score patients.
Likewise, the SRA is negative in a number of high and intermediate 4T score patients.
Our data led us to conclude that neither the PF4 IgG/M/A, PF4 IgG, nor even the SRA reliably predicts the 4T score. Further, interassay results do not correlate well.
Based on the above findings, we recommend that physicians use their clinical judgment aided by the 4T score when evaluating a patient with possible HIT. If the 4T score is ≥ 4 (intermediate or high), all heparin exposure should be prompted stopped, a hematology consult is suggested, and the PF4 IgG/M/A ELISA should be considered. In addition, the presence of a DVT should be sought. If thrombosis or skin necrosis is confirmed, the patient should be treated with intravenous direct thrombin inhibitors such as argatroban or lepirudin and followed with daily platelet counts. Monitoring of the direct thrombin inhibitor should be done with the partial thromboplastin time assay. In patients without thrombosis, Fondaparinux may be used prophylactically as long as the creatinine clearance is greater than 30 ml/hr.
It appears from our findings and the shared experience of the panel that assays designed to measure IgG anti-heparin/PF4 avidity provide a promising direction. However, at present, we must interpret local screening and reference assays in light of their limitations and base a HIT diagnosis primarily on changes in platelet count and clinical impression