Method of detecting platelet activating antibodies that cause heparin-induced thrombocytopenia/thrombosis

Inventors

Aster, Richard H. • BOUGIE, Daniel W. • JONES, Curtis Gerald • PADMANABHAN, Anand

Assignees

Versiti Blood Research Institute Foundation Inc

Abstract

The present invention provides a method of detecting platelet activation in a patient, the method comprising the steps of a) obtaining a blood sample from a patient suspected of having heparin-induced thrombocytopenia (HIT); b) incubating an effective amount of platelet factor 4 (PF4) with a sample of platelets to yield a sample of PF4-treated platelets; c) contacting the patient blood sample with the PF4-treated platelets; and d) measuring the extent of platelet activation, wherein an increase in platelet activation compared with results obtained using a normal blood sample is indicative of the patient having HIT.

Core Innovation

The invention relates to methods and kits for determining the presence of platelet-activating pathogenic heparin-induced thrombocytopenia (HIT) antibodies as opposed to non-activating non-pathogenic HIT antibodies in a whole blood, plasma or serum sample from a patient suspected of having HIT. The approach uses purified platelet factor 4 (PF4) to engage the relevant antibodies and employs an antibody or probe capable of detecting platelet activation or antibody bound to platelets.

The kit includes control samples negative and positive for platelet-activating pathogenic HIT antibodies, and provides instructions for use. Detection of platelet activation or platelet-bound antibody is performed using the PF4 component and the antibody or probe that indicates the presence of the pathogenic platelet-activating HIT antibody population in the suspected patient sample.

In described implementations, increased platelet activation is detected by determining platelet activation markers such as p-selectin (CD62P) expression, or by detecting platelet binding of patient IgG to PF4-treated platelets. High-dose heparin is used as a specificity control that suppresses the signal, and optional components include an inhibitor of platelet activation such as prostaglandin E1 and detection reagents such as annexin V or anti-p-selectin.

Claims Coverage

The independent claim is directed to a HIT diagnostic kit that distinguishes platelet-activating pathogenic HIT antibodies from non-activating non-pathogenic HIT antibodies in a whole blood, plasma or serum sample. The inventive features explicitly include PF4, a detecting antibody or probe for platelet activation or platelet-bound antibody, and both negative and positive control samples for platelet-activating pathogenic HIT antibodies. Dependent claims refine the kit by adding heparin, an inhibitor of platelet activation, and fluorescent labeling options for the detecting reagent.

Overall, the claims cover a PF4-based diagnostic kit that detects platelet-activating pathogenic HIT antibodies by measuring platelet activation or platelet-bound antibody, with explicit positive and negative controls for platelet-activating pathogenic HIT antibodies. Dependent claim refinements specify added heparin, an inhibitor of platelet activation, and fluorescent labeling options for the detecting probe, including fluorescent-labeled annexin V.

Stated Advantages

Documented Applications