Investigation of 2 fluorophores dissolved in predefined solvents with the goal to perform FRET by observing fluorescence
Miltscho Andreev
Institute of Solid State Physics
11:15 - 12:15 Wednesday 09 May 2018 PH01150 Foerster Resonant Energy Transfer (FRET) is a well-known research method
in fields including medicine, chemistry and biology. Based on a dipole-
dipole coupling between 2 fluorophores, defined as donor (D) and acceptor
(A), the molecule interactions serve as a ”spectroscopical ruler”. It is a
common technique applied for example to monitor distances of chemical
reactions, molecule structural elucidations or in vivo cellular analysis.
The main target of this diploma thesis was to investigate the possibility of
FRET between 2 predefined fluorophores dissolved in Dimethylformamid
(DMF) and distilled water at pH 9. For this purpose, the Foerster distance
and the FRET efficiency were introduced as figures of merit to illustrate the
performance of the set-up. According to preliminary assumptions, such as
the negative impact of DMF on fiber-fiber linkage behavior, distilled water
at pH 9 was prioritized in the course of research. Several concentration
series were prepared in the range of a few micromole per liters up to the
saturation concentration of the pure fluorophore solutions. Additionally,
D-A intermixtures were produced to be measured in a spectrophotometer
and a spectrofluorophotometer. As a result of the assessment process, the
Foerster distance was determined to 4.4 nm and the FRET efficiency was
obtained.
The challenge of the research was the lack of information about the per-
formance of the fluorophore interaction within the chosen environment in
dependence of the concentration. Low concentrations did not yield into
a conclusive FRET efficiency, since donor quenching was recorded, but
no acceptor sensitization. Finally, extra fluorescence measurements were
performed with a self-assembled set-up, avoiding inner filtering effects
at relatively high concentrations (around 0.05 mmol/l with standard quartz
) glass cuvette. On the basis of this work distilled water at pH 9 is not a
recommended solvent for this predefined fluorophore pair.
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Institute of Solid State Physics
