Biomedical Engineering Reference
In-Depth Information
Fig. 4.5
Fluorescence
emission spectra of BSA at
three different pHs.
Curves a
,
b
, and
c
represent pH 3.8, 7.0,
and 9.0, respectively
(Adapted from [
26
])
monitored by steady-state [
26
] or time-resolved fluorescence spectroscopy, fluores-
cence resonance energy transfer (FRET), or stepwise single-molecule
photobleaching. Rocker et al. [
27
] have quantitatively analyzed the adsorption of
human serum albumin onto polymer-coated FePt and CdSe/ZnS NPs by fluores-
cence correlation spectroscopy (FCS). They have shown the formation of a protein
corona monolayer with a thickness of 3.3 nm. FCS can be used to obtain quantita-
tive data on the dynamics of the protein corona in a biological environment which
would be useful for a multitude of protein nano-science applications.
Due to presence of two tryptophan (Trp) residues on BSA, fluorescence emission
of BSA provides information on the conformational change of BSA in different
environment. Shang et al. [
26
] showed (Fig.
4.5
) that the change of pH can change
the conformation and hence the emission spectra of BSA.
4.7 Mass Spectrometry
Mass spectrometry (MS) is now a powerful and essential instrument in life science
and proteomics studies where it is used as analytical technique to identify protein
identities. Usually, the sample is digested in small peptides which are ionized and
fragmented.
MS provides qualitative and quantitative information of the protein mixture. It
has been successfully applied to identify protein coronas using a gel-based meth-
odology which requires a previous sample separation on SDS-PAGE. The band of
interest is cut, followed by in-gel trypsin digestion to extract the peptides and
analyzed by mass spectrometry. A non-gel-based approach required in-solution
trypsin digestion on the proteins adsorbed on the NP surface. A protein denaturation
is necessary; the choice of the denaturing agent has to be carefully done to avoid
