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applications of QDs for combined spectral and lifetime multiplexing, thereby further
increasing the number of species to be discriminated.
3.6 Strategies for Signal Amplification
Signal enhancement is one of the major challenges not only in the improvement of
luminescent sensors, but also for many luminescence-based methods used for the
analysis of samples available only in very small quantities. This can help to
improve the signal-to-noise ratio and to minimize the influence of background
fluorescence or ambient light. Moreover, it paves the road for increasingly desired
miniaturization and simple readout devices and helps to reduce costs. Fluorescence
amplification strategies include enzymatic amplification, avidin-biotin or antibody-
hapten secondary detection techniques, nucleic acid amplification, controlled
aggregation, chromophore-metal interactions (metal-enhanced fluorescence or
MEF, observed for the metals silver and gold), and multiple-fluorophore labels
(e.g., phycobiliproteins or particle labels including systems with releasable fluor-
ophores, dendrimeric systems, and FRET-based light harvesting systems). Such
amplification strategies have been established for organic dyes and can often be
used only for certain applications, such as fluoroimmunoassays. These approaches
can be transferred to QDs only to certain degrees. For instance, methods involving
the use of a fluorogenic enzyme substrate cannot be transferred to QD technology.
However, enzymatic amplification has been combined with QDs in the past [
130
].
Approaches such as controlled aggregation or the construction of multichromopho-
ric systems like chromophore-doped particle labels are similarly suited for both
organic dyes and QDs. MEF, that exploits the coupling of the chromophore's
transition dipole moment to metal plasmons, can provide emission enhancement
factors of typically ca. 10 up to a few hundred for organic chromophores, depending
on the fluorescence quantum yield of the respective dyes, in conjunction with
reduction in fluorescence lifetime and increased photostability [
131
]. The enhance-
ment factors, however, depend on the type, shape, and size of the metal, on the type
of chromophore, and on geometrical parameters (metal-fluorophore distance, ori-
entation) and thus require sophisticated dye-metal nanoparticle systems or (dye-
doped) core/shell-nanostructures. In the case of QDs, only moderate amplification
effects (e.g., fivefold fluorescence enhancement for a CdTe-Au-system) have been
observed [
132
,
133
]. The potential of this and other signal amplification approaches
to optimize QD properties and to enable new sensor applications still needs to be
thoroughly investigated.
3.7 Reproducibility, Quality Assurance and Limitations
Aside from instrument-specific contributions that can be corrected for, target
quantification from measurements of fluorescence is affected to a nonnegligible
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