The above-mentioned (Sect. 6.4.1) research have some major limitations related
to, i.e., sample preparation and fixation methodology of ex vivo tissues and gene
knockout modifications in animal model systems. Thus, it is necessary to reduce
these obstacles and gain non-falsified information about the system. Nowadays, a
lot of effort is put to develop in vivo Raman diagnostics. Spontaneous Raman
spectroscopy can be combined with optical fibers for endoscopy of hollow organs.
This nondestructive label-free approach enables to gather specific information about
chemical structure of the tissue and its alternation from even very early stages of
disease expansion. However, work with fiber optic probes encounters some problems such as a strong background signal arising from silicon-based material of
probe and weak Raman scattering of biological species. To solve these issues,
various sophisticated schemes and optical elements are designed and integrated into
common fiber probes for medical applications [178]. So far, Raman spectroscopy
via fiber optic probes in in vivo biomolecular characterization and diagnosis was
performed on skin [179], brain [180], components of the digestive system including
esophagus [181], larynx [182], stomach [183] and colon [183], breast [184], arteries
[185], lung [186], liver [187], cervix [188], and bladder [189]. The main aim of
these studies was to identify spectroscopic markers associated with the early
detection of cancerous lesions. Recent advances in in vivo Raman spectroscopy are
also summarized in two reviews [161, 165].
6.4.2 Tissue Imaging With Immuno-SERS Microscopy
Label-free conventional Raman spectroscopy has been proven to be a powerful and
nondestructive tool for investigation even such complex chemical structures like
tissues. However, it requires long data collection time to obtain reliable and high
signal-to-noise spectral information. Advantages of SERS over conventional
Raman technique were introduced to the reader in the previous paragraphs of this
chapter (Sects. 6.1 and 6.2) and include recording a selective signal from a target
molecule. Furthermore, because of enhancement, a standard SERS data collection
allows reducing laser powers and acquisition times what is a key in the diagnostic
field. Selectivity of SERS nanoprobes is achieved by functionalization of metal
nanoparticles with antibodies or aptamers. An antibody-based method has been
developed since 1941 when Albert H. Coons had described a new way of localization of antigens in tissues using a fluorescent-labelled antibody which was called
thereafter as immunohistochemical staining [190]. A similar approach was transferred to design SERS probes and develops immuno-SERS staining (iSERS).
Typical SERS tags employed in an immunoassay are composed of a metallic
nanoparticle, Raman reporter, protective shell which also facilities conjugation of a
targeting ligand, and an antibody as a biorecognition molecule (Fig. 6.10a) [48].
Immuno-SERS microscopy in recent time was proposed as an alternative
immunohistochemistry method. Several obstacles typical for immunohistochemical
fluorescence can be overcome by using iSERS nanotags, e.g., the lack of
186
K. Czamara et al.
to, i.e., sample preparation and fixation methodology of ex vivo tissues and gene
knockout modifications in animal model systems. Thus, it is necessary to reduce
these obstacles and gain non-falsified information about the system. Nowadays, a
lot of effort is put to develop in vivo Raman diagnostics. Spontaneous Raman
spectroscopy can be combined with optical fibers for endoscopy of hollow organs.
This nondestructive label-free approach enables to gather specific information about
chemical structure of the tissue and its alternation from even very early stages of
disease expansion. However, work with fiber optic probes encounters some problems such as a strong background signal arising from silicon-based material of
probe and weak Raman scattering of biological species. To solve these issues,
various sophisticated schemes and optical elements are designed and integrated into
common fiber probes for medical applications [178]. So far, Raman spectroscopy
via fiber optic probes in in vivo biomolecular characterization and diagnosis was
performed on skin [179], brain [180], components of the digestive system including
esophagus [181], larynx [182], stomach [183] and colon [183], breast [184], arteries
[185], lung [186], liver [187], cervix [188], and bladder [189]. The main aim of
these studies was to identify spectroscopic markers associated with the early
detection of cancerous lesions. Recent advances in in vivo Raman spectroscopy are
also summarized in two reviews [161, 165].
6.4.2 Tissue Imaging With Immuno-SERS Microscopy
Label-free conventional Raman spectroscopy has been proven to be a powerful and
nondestructive tool for investigation even such complex chemical structures like
tissues. However, it requires long data collection time to obtain reliable and high
signal-to-noise spectral information. Advantages of SERS over conventional
Raman technique were introduced to the reader in the previous paragraphs of this
chapter (Sects. 6.1 and 6.2) and include recording a selective signal from a target
molecule. Furthermore, because of enhancement, a standard SERS data collection
allows reducing laser powers and acquisition times what is a key in the diagnostic
field. Selectivity of SERS nanoprobes is achieved by functionalization of metal
nanoparticles with antibodies or aptamers. An antibody-based method has been
developed since 1941 when Albert H. Coons had described a new way of localization of antigens in tissues using a fluorescent-labelled antibody which was called
thereafter as immunohistochemical staining [190]. A similar approach was transferred to design SERS probes and develops immuno-SERS staining (iSERS).
Typical SERS tags employed in an immunoassay are composed of a metallic
nanoparticle, Raman reporter, protective shell which also facilities conjugation of a
targeting ligand, and an antibody as a biorecognition molecule (Fig. 6.10a) [48].
Immuno-SERS microscopy in recent time was proposed as an alternative
immunohistochemistry method. Several obstacles typical for immunohistochemical
fluorescence can be overcome by using iSERS nanotags, e.g., the lack of
186
K. Czamara et al.