Nucleic acid diagnostics is a growing sector in life sciences. Numerous applications
based on the amplification of nucleic acids are utilized to determine the presence,
composition, quality, or quantity of the sequence in question. However, amplification is a
time-consuming, laborious, and error-prone process. Therefore, it would be preferable to
perform tests with a minimized number of amplification cycles or, alternatively, completely
without the amplification.
We present two very sensitive sandwich assay concepts for a measurement of nucleic acids. The
assays are performed either in standard microtitration wells or on microparticles coated with
capture probes. Detection-probe-coated europium(III) nanoparticle labels are used for signal
generation. The detection limits of the microtitration well and microparticle applications were
4.0 × 105
and 6.1 × 104
copies of target sequence, respectively. The reference assay, based on the detection
of europium(III) chelate-labelled detection probes, had a detection limit of
8.5 × 107
copies. Thus, 100–1000-fold improvement in the sensitivity was achieved. The high
sensitivity of the designed assays is based on the long-lifetime fluorescence of nanoparticle
labels, on time-resolved fluorometry, and on signal rather than target amplification. The
presented sandwich assays provide the tool for a development of direct, amplification-free
detection of nucleic acids.