J. Bukowska and P. Piotrowski
48
SERS is also applied to microorganisms such as bacteria. It is difficult to introduce metal nanoparticles into bacterial cells because of their size, which is much
smaller in comparison to mammalian cells. therefore, the studies of bacteria with
SERS are mainly focused on the extracellular experiments. however, Jarvis et al.
reported intracellular SERS of Geobacter sulfurreducens using colloidal metal
nanoparticles prepared from the enzymatic reduction of silver and gold salts by
these bacteria [154]. very interesting and critical review of the problems that are
connected with the interpretation of the SERS spectra of such a complex molecular
systems was published by Efrima and Zeiri [155]. they paid attention to considerable differences between the SERS spectra reported for the same bacterium by
various authors. Efrima and Zeiri suggested that the differences observed are due
to differences in preparation procedures that may result in interactions of nanoparticles with various chemical constituents of the cells. they demonstrated clear dependence of the spectra on localization of the nanoparticles. unusual selectivity and
sensitivity of SERS spectrum toward flavins, an essential coenzyme in cells, was
observed for external (on the cell walls) coating of the bacteria with the metal particles. on the contrary, when the metal colloids were produced inside the cell, flavin
signatures disappeared and the bands corresponding to internal chemical constituents appeared instead. to overcome the limitations of colloidal nanoparticles for
SERS of the cells and microorganisms and to gain more control upon the position of
the sensor, new approaches to intracellular sensing have been proposed. the most
important are: using metal-coated fiber-optic tips [156, 157], SERS-active glass
nanopipettes [145, 152] and tip-enhanced Raman scattering (tERS) [158, 159].
tERS technique, briefly described in Sect. 3, is a perfect tool in biosensing.
Cell walls were successfully investigated with tERS. thanks to sub-optical spatial
resolution of this technique, it is possible to distinguish between protein and lipid
components of the Staphylococcus epidermidis cell wall [158, 160]. Fluctuations
in the spectra collected from one spot of the outer layer of the cell may be ascribed
to the bacterial surface dynamics [161]. Surface of a single tobacco mosaic virus
was also characterized with tERS. Not only protein components from the capsid,
but also some RNA features were seen in the spectrum (Fig. 3.9) [159]. these very
promising results show that fast spectroscopic identification of single virus particles
with tERS is possible.
Biosensors are able to look directly into the cell compartments—which used to
be impossible without complex techniques—without affecting cell viability. however, uncoated nanoparticles suffer from lack of specificity. While features of the
spectra could be assigned to some types of biomolecules, such a simple device
fails to determine the presence of particular molecules. Rich spectra of biomolecules make it difficult to determine all the components accurately—profusion of
the bands, as well as their overlapping, hinder clarity of the spectrum. In order to
increase specificity, it is better to detect molecules in an indirect way. In this case,
SERS signal is collected from a small reporter molecule adsorbed on a nanoparticle.
molecular dyes are frequently used as reporter molecules; as a result, even higher
intensities thanks to SERRS effect are obtained.
48
SERS is also applied to microorganisms such as bacteria. It is difficult to introduce metal nanoparticles into bacterial cells because of their size, which is much
smaller in comparison to mammalian cells. therefore, the studies of bacteria with
SERS are mainly focused on the extracellular experiments. however, Jarvis et al.
reported intracellular SERS of Geobacter sulfurreducens using colloidal metal
nanoparticles prepared from the enzymatic reduction of silver and gold salts by
these bacteria [154]. very interesting and critical review of the problems that are
connected with the interpretation of the SERS spectra of such a complex molecular
systems was published by Efrima and Zeiri [155]. they paid attention to considerable differences between the SERS spectra reported for the same bacterium by
various authors. Efrima and Zeiri suggested that the differences observed are due
to differences in preparation procedures that may result in interactions of nanoparticles with various chemical constituents of the cells. they demonstrated clear dependence of the spectra on localization of the nanoparticles. unusual selectivity and
sensitivity of SERS spectrum toward flavins, an essential coenzyme in cells, was
observed for external (on the cell walls) coating of the bacteria with the metal particles. on the contrary, when the metal colloids were produced inside the cell, flavin
signatures disappeared and the bands corresponding to internal chemical constituents appeared instead. to overcome the limitations of colloidal nanoparticles for
SERS of the cells and microorganisms and to gain more control upon the position of
the sensor, new approaches to intracellular sensing have been proposed. the most
important are: using metal-coated fiber-optic tips [156, 157], SERS-active glass
nanopipettes [145, 152] and tip-enhanced Raman scattering (tERS) [158, 159].
tERS technique, briefly described in Sect. 3, is a perfect tool in biosensing.
Cell walls were successfully investigated with tERS. thanks to sub-optical spatial
resolution of this technique, it is possible to distinguish between protein and lipid
components of the Staphylococcus epidermidis cell wall [158, 160]. Fluctuations
in the spectra collected from one spot of the outer layer of the cell may be ascribed
to the bacterial surface dynamics [161]. Surface of a single tobacco mosaic virus
was also characterized with tERS. Not only protein components from the capsid,
but also some RNA features were seen in the spectrum (Fig. 3.9) [159]. these very
promising results show that fast spectroscopic identification of single virus particles
with tERS is possible.
Biosensors are able to look directly into the cell compartments—which used to
be impossible without complex techniques—without affecting cell viability. however, uncoated nanoparticles suffer from lack of specificity. While features of the
spectra could be assigned to some types of biomolecules, such a simple device
fails to determine the presence of particular molecules. Rich spectra of biomolecules make it difficult to determine all the components accurately—profusion of
the bands, as well as their overlapping, hinder clarity of the spectrum. In order to
increase specificity, it is better to detect molecules in an indirect way. In this case,
SERS signal is collected from a small reporter molecule adsorbed on a nanoparticle.
molecular dyes are frequently used as reporter molecules; as a result, even higher
intensities thanks to SERRS effect are obtained.