New laboratory test in flow cytometry for the combined analysis of serologic and cellular parameters in the diagnosis of heparin‐induced thrombocytopenia

• Published in: Cytometry. Part B, Clinical cytometry Clinical cytometry, 2004-03, Vol.58B (1), p.32-38
• Main Authors: Gobbi, Giuliana, Mirandola, Prisco, Tazzari, Pier Luigi, Talarico, Enrica, Caimi, Luigi, Martini, Giuliana, Papa, Stefano, Conte, Roberto, Manzoli, Francesco A., Vitale, Marco
• Format: Article
• Language: English
• Subjects: Adult; Aged; Annexin A5; Annexin V; Antigen-Antibody Complex - analysis; Antigen-Antibody Complex - immunology; Blood Platelets - cytology; Blood Platelets - drug effects; Blood Platelets - immunology; Blood Platelets - metabolism; Enzyme-Linked Immunosorbent Assay; Female; flow cytometry; Flow Cytometry - methods; Heparin - adverse effects; Heparin - immunology; heparin‐induced thrombocytopenia; Humans; Male; Middle Aged; particle gel immunoassay; platelet factor 4; Platelet Factor 4 - immunology; Serology - methods; Thrombocytopenia - blood; Thrombocytopenia - chemically induced; Thrombocytopenia - diagnosis; Thrombocytopenia - immunology
• ISSN: 1552-4949; 1552-4957
• DOI: 10.1002/cyto.b.10062

Background Heparin‐induced thrombocytopenia (HIT) is a major complication of heparin therapy. A quick and reliable laboratory assay for the combined determination of pathogenic anti‐heparin and platelet factor 4 (H:PF4) antibodies in the serum and platelet activation is not currently available. Methods We developed a new single‐tube assay in flow cytometry that combines the detection of antibodies in the serum and their activatory properties on platelets. The assay was tested on 13 serum samples from patients with suspected HIT and six samples from normal donors. The presence of anti‐H:PF4 antibody complexes was detected by H:PF4‐coated beads, and donor platelet activation induced by HIT sera was determined by Annexin V binding. All data were compared with the patients' clinical setting, laboratory tests, and standard enzyme‐linked immunosorbent assay detection of anti‐H:PF4 antibodies. Results This flow cytometry assay allowed unequivocal, simultaneous detection of anti‐H:PF4 antibodies in sera and their activatory properties on platelets. All cases for which the diagnosis of HIT was confirmed were detected by the flow assay. Conclusions This assay, combining for the first time functional and nonfunctional testing on anti‐H:PF4 antibodies, is likely to influence the clinical decision for the management of HIT patients. © 2003 Wiley‐Liss, Inc.