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Flow cytometry has been instrumental in rapid analysis of single cells since the 1970s. One of the common approaches is
the immunofluorescence study involving labeling of cells with antibodies conjugated to organic fluorophores. More
recently, as the application of flow cytometry extended from simple cell detection to single-cell proteomic analysis, the
need of determining the actual number of antigens in a single cell has driven the flow cytomery technique towards a
quantitative methodology. However, organic fluorophores are challenging to use as probes for quantitative detection
due to the lack of photostability and of quantitative fluorescence standards. National Institute of Standards and
Technologies (NIST) provides a set of fluorescein isothiocyanate (FITC) labeled beads, RM 8640, which is the only
nationally recognized fluorescent particle standard. On the other hand, optical characteristics of semiconductor
nanocrystals or quantum dots or QDs are superior to traditional dye molecules for the use as tags for biological and
chemical fluorescent sensors and detectors. Compelling advantages of QDs include long photostability, broad spectral
coverage, easy excitation, and suitability for multiplexed sensing. Recently, novel surface coatings have been
developed to render QDs water soluble and bio-conjugation ready, leading to their use as fluorescent tags and sensors for
a variety of biological applications including immunolabeling of cells. Here, we describe our approach of using
fluorescent semiconductor QDs as a novel tool for quantitative flow cytometry detection. Our strategy involves the
development of immuno-labeled QD-conjugated silica beads as "biomimetic cells." In addition to flow cytometry, the
QD-conjugated silica beads were characterized by fluorescence microscopy to quantitate the number of QDs attached to
a single silica bead. Our approach enables flow cytometry analysis to be highly sensitive, quantitative, and encompass
a wide dynamic range of fluorescence detection. Quantitative aspects of the proposed flow cytometery-based approach
for measurement of the QD-based biomimetic samples are discussed.
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