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Lanthanide-Labeled Polymeric Microspheres for Highly Multiplexed

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					         Lanthanide-Labeled Polymeric Microspheres for Highly Multiplexed Bio-Assays
                                 Utilizing ICP-MS Detection
         Stuart C. Thickett,1,2 Ahmed I. Abdelrahman,1 Olga Ornatsky,1 Dmitry Bandura,1 Vladimir Baranov1 and
                                                   Mitchell A. Winnik1
     1
         Department of Chemistry, The University of Toronto, 80 St George Street Toronto ON M5S 3H6 Canada
              2
                  Current Address: School of Chemistry F11, The University of Sydney, NSW 2006 Australia


    The traditional use of fluorescent chromophores in multiplexed biological assays (such as particle-based
assays) is severely limited by spectral overlap, which reduces the number of unique species that can be resolved
in one experiment. We have focused on utilizing mass spectrometric (ICP-MS) detection in bio-assays through
the development of a unique ‘mass cytometer’1 instrument, coupling individual cytometric delivery with mass
spectrometric detection (see Fig. 1). The resolution and large dynamic range of ICP-MS allows for a
significantly increased multiplexed capability compared to fluorescence. Members of the lanthanide (Ln) series
were chosen as metal ‘tags’ in this work,2 given the large number of Ln isotopes and their low biological
abundance.
    In this work, we present the development of Ln-labeled, functional polymer microspheres that can act as
‘reporter particles’ for bio-assays based on mass cytometric detection.3. Surface-functional polystyrene (PS)
seed latexes were prepared by seeded emulsion polymerization with a mixture of monomer, a pre-formed
hydrophobic Ln complex and a monodisperse PS seed latex to yield polymer particles with encapsulated Ln
labels. Parameters such as final particle size (up to 1 µm diameter), type and concentration of encapsulated Ln
surface chemistry for later bioconjugation could be controlled by this method. Individual analysis of these
particles by mass cytometry demonstrated that the loading efficiency of Ln ions was close to 100 %, and up to
107 Ln ions could be incorporated into each particle. Subsequent covalent attachment of metal-labeled proteins
(such as Neutravidin) to the surface of our particles, followed by mass cytometry analysis, provided quantitative
data regarding the number of bound biomolecules on the surface. These particles could also be used for the
simultaneous detection of labeled antibodies and antigens in a model sandwich assay.




     Figure 1. Schematic representation of mass cytometer, allowing for individual analysis of labeled particles.


1.           Bandura, D. R. et. al. Anal. Chem. 2009, 81, (16), 6813-6822.
2.           Lou, X.; et. al. Angew. Chem. Int. Ed 2007, 119, 1-5.
3.           Thickett, S. C et. al. J. Anal. At. Spectrom. 2010, 25, (2), 269-281.


Stuart Thickett
Dr
School of Chemistry F11, The University of Sydney, NSW 2006 Australia
Phone: +61 2 9351 7596 Fax: +61 2 9351 3329 Email: stuart.thickett@sydney.edu.au
Personal History :
   2007 Doctor of Philosophy in Chemistry, The University of Sydney
   2008 Post-Doctoral Fellow, The University of Queensland
   2008 – 2009: Post-Doctoral Fellow, The University of Toronto
   2009 – : Post-Doctoral Fellow, The University of Sydney
Research Interests: heterogeneous polymerization, stimuli-responsive polymers, polymer surface coatings

				
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