Detection of Activated Platelets in Whole Blood Using Activation-Dependent Monoclonal Antibodies and Flow Cytometry

Platelets may become activated in a number of clinical disorders and participate in thrombus formation. We developed a direct test for activated platelets in whole blood using flow cytometry. Whole blood was incubated with either biotin-PAC1, a monoclonal antibody specific for the fibrinogen recepto...

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Bibliographic Details
Published in:Blood 1987-07, Vol.70 (1), p.307-315
Main Authors: Shattil, Sanford J., Michael, Cunningham, Hoxie, James A.
Format: Article
Language:English
Subjects:
Adenosine Diphosphate - pharmacology
Antibodies, Monoclonal
Antibody Specificity
Antigens, Surface - analysis
Antigens, Surface - immunology
Binding Sites, Antibody - drug effects
Binding, Competitive
Biological and medical sciences
Biotin - pharmacology
Blood Platelets - cytology
Blood Platelets - immunology
Blood Platelets - ultrastructure
Cell Separation - methods
Chromatography, Gel
Fibrinogen - physiology
Flow Cytometry - methods
Fluorescent Antibody Technique
Fundamental and applied biological sciences. Psychology
Fundamental immunology
Immunohematology
Platelet Aggregation - drug effects
Platelet immunology
Platelet Membrane Glycoproteins - immunology
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Summary:Platelets may become activated in a number of clinical disorders and participate in thrombus formation. We developed a direct test for activated platelets in whole blood using flow cytometry. Whole blood was incubated with either biotin-PAC1, a monoclonal antibody specific for the fibrinogen receptor on activated platelets, or biotin-S12, an antibody specific for an a-granule membrane protein that associates with the platelet surface during secretion. Platelet-bound antibodies were detected with streptavidin conjugated with fluorescein isothiocyanate (FITC) or phycoerythrin (PE). Platelets were differentiated from the larger erythrocytes and WBCs by their light-scatter profile. Alternatively, platelets could be identified with FITC-AP1, an antibody specific for platelet membrane glycoprotein lb, and analyzed further for PAC1 or S12 binding with PE-streptavidin. No centrifugation or washing steps were required. With gel-filtered platelets, there was a direct correlation between ADP-induced biotin-PAC1 binding and binding determined in a conventional 125I-PAC1 binding assay (r =. 99; P
ISSN:0006-4971
1528-0020
DOI:10.1182/blood.V70.1.307.307