Biomedical Engineering Reference
In-Depth Information
Fig. 4.4
Transmission
spectra of gold NPs
incubated with BSA
measured by UV-Vis
spectroscopy (Adapted from
[
21
])
4.6 Fluorescence Spectroscopy
Fluorescence spectroscopy (FS) which sometimes is called spectrofluorometry is
based on exciting the electrons in the ground state to higher energy levels, called
excited states. The return of excited electrons to their ground state can be radiative
(a photon is emitted) or non-radiative (a phonon is created). The photon emission in
the radiative recombination is called the fluorescence. Therefore, in simple words,
an exciting wavelength is irradiated on the sample and the emitted fluorescence is
measured.
Some amino acid groups such as tyrosine, tryptophan, and phenylalanine have
fluorescent properties and therefore are called fluorophores. In the study of the
NP-protein interactions, either NP or protein or even both could be fluorescent, and
in the case that none of them is fluorescent, a fluorophore should be added to the
system. Since fluorescence labeling can change the conformation or structure of
proteins or change the affinity of proteins for the NP surface,
it could be
challenging.
A derivative of poly(
p
-phenyleneethynylene) (PPE) has been used as a fluores-
cence indicator to study the adsorption of different proteins on the surface of gold
NPs [
25
]. The attachment of PPE on the surface of gold NPs results in the transfer of
excited electrons from the fluorophore to the metallic NP which and hence the
florescence is quenched. The adsorption of other proteins in the environment results
in the detachment of PPE, and therefore, the florescence intensity increases.
Fluorescence spectroscopy is sensitive to protein dynamics because the excited
fluorescent state persists for nanoseconds, which is the time scale of many impor-
tant biological processes such as the rotational motion of protein side chains,
molecular binding, and conformational changes [
10
]. NP-protein binding can be
