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A P P L I C A T I O N N O T E
NanoTemper Technologies devices such as the
Monolith, Dianthus and Dianthus uHTS have
been engineered to measure spectral shis at
low target concentrations (usually 1 – 5 nM) by
monitoring emission intensities at 670 nm and
650 nm (Figure 1) and taking the ratio of the
two wavelengths. In drug discovery, Spectral
Shi technology can be used to identify and
characterize potential drug candidates by
monitoring molecular interactions. The direction
of the emission shi can provide low resolution
information on conformational changes,
contributing to a deeper understanding of
biological processes. When monitoring the
signal as a function of concentration of analyte,
it is possible to directly calculate the dissociation
constant, K
d
.
The use of small molecule fluorescent probes
attached via a number of different chemistries
allows measurement of binding interactions in
solution, and as the signal is not proportional
to mass changes, many target classes and drug
modalities can be characterized (e.g. nucleic
acids [2], small molecule inhibitors [3], covalent
inhibitors [4], antibodies, molecular glues [5],
fragment screening [6], PPI [7]).
Figure 1. Spectral Shift detection in Dianthus uHTS is achieved by monitoring at emission intensity at two distinct
wavelengths, 650 and 670 nm. The NanoTemper small molecule fluorescent dyes demonstrate a blue shift when
moving into more polar environments and a red shift when moving into more hydrophobic environments.
