Long-lifetime luminescent nanobioprobes for advanced cytometry biosensing
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- Nanotechnology in Australia: Showcase of Early Career Research, 2011, pp. 317 - 345
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In biological discovery and disease diagnosis, there are increasing demands for rapid detection of trace amounts of cells, organisms, and molecules. one of the most popular methods is labelling the target analytes with fluorescent bioprobes, thus making the targets spectrally distinguishable. However, conventional molecular bioprobes are weak, photobleaching, and often not sufficient to suppress autofluorescence backgrounds by spectral discrimination, since many naturally occurring substances are autofluorescent under ultraviolet (uV) or visible-wavelength excitation. This represents a typical biosensing problem of detecting a needle in a haystack. Here we show long-lifetime luminescent silica nanoparticles as bioprobes to provide excellent opportunities in suppression of autofluorescence backgrounds in the temporal domain. This is due to the exceptionally long lifetime from lanthanide bioprobes in the order of several hundred microseconds, in contrast to the few-nanosecond-lifetime autofluorescence backgrounds. using a covalent-binding nanoencapsulation technique, the as-prepared ~40 nm monodisperse silica nanoparticle effectively protects thousands of lanthanide complex dyes against the quenching environment. This results in both remarkable signal amplification and enhanced photostability. Due to the large difference in the lifetime, the microsecondlived signal luminescence can be detected in a background-free condition against the nanosecond-lived autofluorescence using time-gated detection. Furthermore, the effective simultaneous confinement of multiple lanthanide-element dyes within nanoparticles also provides opportunities to produce multiplexing bioprobes. In this chapter, we showcase our successfully engineered microsecond-lifetime nanobioprobes for immunobioassays of human prostate-specific antigen (psA) and bioimaging applications of an environmental pathogen Giardia lamblia. our achievements include a radical extension of the excitation wavelength from uV to visible (excitation peak from 330 nm to 406 nm) and a demonstration for multicolour lanthanide nanobioprobes. In addition, our silica nanoparticles are highly reproducible, biocompatible, and non-toxic. These results suggest a new class of ultrasensitive bioprobes to meet the increasing requirements of "ultrasensitivity" and "multiplexing capacity" in advanced biomolecular and cellular assays. © 2011 by Pan Stanford Publishing Pte. Ltd. All rights reserved.
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