Quantitative interpretation of optical density measurements using PF4-dependent enzyme-immunoassays


Ted Warkentin, Hamilton Regional Laboratory Medicine Program, Room 1-180A, Hamilton Health Sciences, Hamilton General Site, 237 Barton Street East, Hamilton, ON L8L2X2, Canada.
Tel.: +1 905 527 0271 ext. 46140; fax: +1 905 577 1421.
E-mail: twarken@mcmaster.ca


Summary. Background: Many laboratories test for heparin-induced thrombocytopenia (HIT) using a PF4-dependent enzyme-immunoassay (EIA). An advantage of the EIA is its simplicity; a disadvantage is that it only indirectly detects heparin-dependent, platelet-activating antibodies (‘HIT antibodies’). Objectives: To determine whether the magnitude of a positive EIA result, expressed in optical density (OD) units, predicts risk of HIT antibodies, defined as a strong-positive platelet serotonin-release assay (SRA) result (≥50% serotonin release). Patients/methods: We determined the risk of a strong-positive SRA result for five categories of OD reactivity (<0.40, 0.40–<1.00, 1.00–<1.40, 1.40–<2.00, and ≥2.00 OD units) using two EIAs (commercial anti-PF4/polyanion IgG/A/M and in-house anti-PF4/heparin–IgG). Results: For patient sera investigated for HIT antibodies, a weak-positive result (0.40–<1.00 OD units) in either EIA indicated a low probability (≤5%) of a strong-positive SRA; the risk increased to ∼90% with an OD ≥ 2.00 units. Quantifying the EIA–SRA relationship for 1553 referred patient sera, we found that for every increase of 0.50 OD units in the EIA–IgG, the risk of a strong-positive SRA result increased by OR = 6.39 [95% confidence interval (CI), 5.13, 7.95; P < 0.0001]. For every increase of 1.00 OD units in the EIA–IgG, the risk increased by OR = 40.81 (95% CI, 26.35, 63.20; P < 0.0001). Conclusions: The probability of HIT antibodies (strong-positive SRA result) inferred by a positive PF4-dependent EIA varies considerably in relation to the magnitude of the EIA result, expressed as OD values. In our laboratory, the probability of HIT antibodies being present reached ≥50% only when the OD level was ≥1.40 units.