New laboratory test in flow cytometry for the combined analysis of serologic and cellular parameters in the diagnosis of heparin-induced thrombocytopenia.
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.
Department of Anatomy, Human Anatomy Section, University of Parma, Parma, Italy.
Giuliana
G
Gobbi
Prisco
P
Mirandola
Pier Luigi
PL
Tazzari
Enrica
E
Talarico
Luigi
L
Caimi
Giuliana
G
Martini
Stefano
S
Papa
Roberto
R
Conte
Francesco A
FA
Manzoli
Marco
M
Vitale
D000328:Adult; D000368:Aged; D017304:Annexin A5; D000936:Antigen-Antibody Complex / Q000032:analysis / Q000276:immunology; D001792:Blood Platelets / Q000166:cytology / Q000187:drug effects / Q000276:immunology / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005434:Flow Cytometry / Q000379:methods*; D006493:Heparin / Q000009:adverse effects* / Q000276:immunology; D006801:Humans; D008297:Male; D008875:Middle Aged; D010978:Platelet Factor 4 / Q000276:immunology; D012699:Serology / Q000379:methods*; D013921:Thrombocytopenia / Q000097:blood / Q000139:chemically induced* / Q000175:diagnosis* / Q000276:immunology
D017304:Annexin A5; D000936:Antigen-Antibody Complex; D010978:Platelet Factor 4; D006493:Heparin