Label-free distinction between platelet and erythrocyte microparticles by Raman microspectroscopy

Oral presentation
E. van der Pol, C.M. Hau, C. Otto, A. Sturk, T.G. van Leeuwen, and R. Nieuwland
XXIII Congress of the International Society on Thrombosis and Haemostasis, Kyoto, Japan
July 27, 2011
Awarded with the Young Investigators Award (YIA)


Background: The cellular origin of microparticles is usually established by fluorescent antibody labeling, which is laborious, expensive, and involves practical problems (JTH 2010, 8: 2596–607). Using an ultra-modern Raman microspectroscopy setup, we have distinguished platelet from erythrocyte microparticles without labeling.

Methods: Platelet and erythrocyte microparticles were isolated from blood bank concentrates by differential centrifugation and analyzed by flow cytometry, transmission electron microscopy, and Raman microspectroscopy. For Raman microspectroscopy, a 35 mW krypton ion laser operating at a wavelength of 647 nm was focused to a probe volume smaller than 1 μm3 by an Olympus microscope objective with an NA of 0.95. The Stokes shift from light scattered by microparticles diffusing through the probe volume was measured by a sensitive spectrograph dispersing in the range 646–849 nm.

Results: The figure shows that platelet microparticles have a Raman spectrum containing spectral transitions characteristic of phospholipids. The Raman spectrum of erythrocyte microparticles, however, is entirely different.

Conclusions: For the first time, platelet microparticles are distinguished from erythrocyte microparticles without fluorescent antibody labeling.