Testing for HIT Antibodies Using cPRP

The following description of the assay taken from Chong and colleagues (1989, 1993a) has the highest reported sensitivity and specificity among c-PRP methods. Blood is obtained from normal blood donors whose platelets respond well to serum or plasma from HIT patients, and c-PRP is prepared. Testing involves addition of 150 mL of patient heat-inactivated c-PPP or serum to 340 mL of c-PRP (final platelet concentration, 250-350 x 109/L) at 37°C. The platelets are monitored for a few minutes to exclude nonspecific platelet aggregation. After addition of 10 mL heparin-saline, aggregation is monitored over the next 15 min or until aggregation has occurred. A positive result is an increase in light transmission of more than 25% above baseline in the presence of therapeutic-dose heparin (0.5 U/ mL) and patient serum or c-PPP and inhibition of aggregation in the presence of patient serum or plasma and supratherapeutic-dose heparin (100U/mL). Use of such a two-point assay reduced the false-positive rate, as serum or plasma from some patients without HIT caused platelet aggregation at all heparin concentrations tested. To ensure that the platelets are functional, platelets are also tested with collagen (2 mg/mL). Details on methodology of c-PRP assays are also given elsewhere (Kapsch and Silver, 1981; Almedia et al., 1998).

Some workers report that platelets from a patient with HIT are very reactive to heparin-dependent activation by their own serum or plasma (Kappa et al., 1987; Chong et al., 1993b). Use of autologous c-PRP can sometimes be limited by the patient's thrombocytopenia, however. Potential explanations for the high sensitivity of autologous platelets include persisting high Fc receptor expression on platelets of patients with acute HIT (Chong et al., 1993b) and baseline platelet activation (Chong et al., 1994), with the potential for higher PF4 availability.

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