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Single-step separation of platelets from whole blood coupled with digital quantification by interfacial platelet cytometry (iPC)
Dublin City University.
Royal College of Surgeons in Ireland, Dublin.ORCID iD: 0000-0002-1920-3962
Royal College of Surgeons in Ireland (RCSI), Dublin.
Royal College of Surgeons in Ireland (RCSI), Dublin.
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2010 (English)In: Langmuir, ISSN 0743-7463, E-ISSN 1520-5827, Vol. 26, no 18, 14700-14706 p.Article in journal (Refereed) Published
Abstract [en]

We report the efficient single-step separation of individual platelets from unprocessed whole blood, enabling digital quantification of platelet function using interfacial platelet cytometry (iPC) on a chip. iPC is accomplished by the precision micropatterning of platelet-specific protein surfaces on solid substrates. By separating platelets from whole blood using specific binding to protein spots of a defined size, iPC implements a simple incubate-and-rinse approach, without sample preparation, that enables (1) the study of platelets in the physiological situation of interaction with a protein surface, (2) the choice of the number of platelets bound on each protein spot, from one to many, (3) control of the platelet-platelet distance, including the possibility to study noninteracting single platelets, (4) digital quantification (counting) of platelet adhesion to selected protein matrices, enabling statistical characterization of platelet subpopulations from meaningfully large numbers of single platelets, (5) the study of platelet receptor expression and spatial distribution, and (6) a detailed study of the morphology of isolated single platelets at activation levels that can be manipulated. To date, we have demonstrated 1-4 of the above list. Platelets were separated from whole blood using iPC with fibrinogen, von Willebrand factor (VWF), and anti-CD42b antibody printed "spots" ranging from a fraction of one to several platelet diameters (2-24 μm). The number of platelets captured per spot depends strongly on the protein matrix and the surface area of the spot, together with the platelet volume, morphology, and activation state. Blood samples from healthy donors, a May-Hegglin-anomaly patient, and a Glanzmann's Thrombasthenia patient were analyzed via iPC to confirm the specificity of the interaction between protein matrices and platelets. For example, the results indicate that platelets interact with fibrinogen spots only through the fibrinogen receptor (αIIbβ3) and, relevant to diagnostic applications, platelet adhesion correlates strongly with normal versus abnormal platelet function. A critical function of platelets is to adhere to regions of damage on blood vessel walls; in contrast to conventional flow cytometry, where platelets are suspended in solution, iPC enables physiologically relevant platelet bioassays based on platelet/protein-matrix interactions on surfaces. This technology should be inexpensive to implement in clinical assay format, is readily integrable into fluidic microdevices, and paves the way for high-throughput platelet assays from microliter volumes of whole blood.

Place, publisher, year, edition, pages
ACS , 2010. Vol. 26, no 18, 14700-14706 p.
National Category
Medical and Health Sciences
URN: urn:nbn:se:liu:diva-71574DOI: 10.1021/la9039682PubMedID: 20108942OAI: diva2:450711
Available from: 2011-10-21 Created: 2011-10-21 Last updated: 2015-03-13

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Ramstrom, Sofia
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