8.1.3 Bioluminescence Reporter System
The bioluminescence reporters have advantages in high sensitivity particularly
compared with fluorescence-based imaging. Bioluminescence imaging of EVs
employs luciferase enzymes as imaging reporters. In particular, one of the most
commonly used luciferase enzymes, Gaussia luciferase (gLuc), was reported to be
much brighter than the firefly or Renilla luciferases. But the tissue penetration of the
emission photon makes the advantage of gLuc over firefly or Renilla luciferases.
Luciferases in common, they are free from autofluorescence problems due to absent
bioluminescence signals due to absence of any bioluminescence proteins reacting
with substrates we inject to these small animals. Thus, this approach is more useful
than fluorescence-based imaging for whole body imaging as a biodistribution study.
As a reporter being expressed solely in EVs, a group used a fusion protein which
consists of gLuc enzyme combined with transmembrane domain of lactadherin
[10]. This bioluminescence reporter overcomes the issue of recirculating and persistent fluorescence signals. Thus, EVs using this fusion protein, gLuc-lactadherin,
was used for serial whole body imaging to quantitatively monitor the clearance of
intravenously injected EVs [23]. A similar bioluminescence reporter system was
reported by Lai et al. using membrane-bound variant of the gLuc reporter and a
biotin acceptor peptide [9]. This reporter system allowed multimodal imaging of
bioluminescence and fluorescence in vivo as fluorescence combined with streptavidin could provide fluorescence-mediated tomography imaging simultaneously.
Bioluminescence reporter system has several merits in small animal imaging due
to high sensitivity, specificity for EVs as well as enabling multimodal imaging as
aforementioned. However, this system has an inherent shortcomings that luminescence signals are attenuated in deep organs and that no substrates of luciferin or
coelenterazine for these luciferases are allowed for the administration to humans.
Moreover, relatively complicated labeling method compared with lipophilic dyes
limits the high throughput in vivo kinetics study of various EVs extracted from
different cell types.
Furthermore, similarly to fluorescence reporter imaging, the bioluminescent
signal depends on the reporter protein expression [7, 15]. These labeling procedures
are complicated compared to those of fluorescence dyes, which limits the study of
biodistribution and fate of EVs in various cells under different conditions in vivo.
8.1.4 Radionuclide Imaging
Briefly, radionuclide imaging of EVs is sensitive and feasible to be extended to the
human and clinical applications. Details of radiolabeled EVs are introduced in
previous chapter. Recently developed
99m Tc-labeled EVs used
99m
Tc-HMPAO and
99m Tc-tricarbonyl. They used direct labeling of EVs [24, 25]. Another
125 I-labeled
EVs employed a reporter system. EVs with a fusion protein of streptavidin and
156
H. Choi and D. S. Lee
The bioluminescence reporters have advantages in high sensitivity particularly
compared with fluorescence-based imaging. Bioluminescence imaging of EVs
employs luciferase enzymes as imaging reporters. In particular, one of the most
commonly used luciferase enzymes, Gaussia luciferase (gLuc), was reported to be
much brighter than the firefly or Renilla luciferases. But the tissue penetration of the
emission photon makes the advantage of gLuc over firefly or Renilla luciferases.
Luciferases in common, they are free from autofluorescence problems due to absent
bioluminescence signals due to absence of any bioluminescence proteins reacting
with substrates we inject to these small animals. Thus, this approach is more useful
than fluorescence-based imaging for whole body imaging as a biodistribution study.
As a reporter being expressed solely in EVs, a group used a fusion protein which
consists of gLuc enzyme combined with transmembrane domain of lactadherin
[10]. This bioluminescence reporter overcomes the issue of recirculating and persistent fluorescence signals. Thus, EVs using this fusion protein, gLuc-lactadherin,
was used for serial whole body imaging to quantitatively monitor the clearance of
intravenously injected EVs [23]. A similar bioluminescence reporter system was
reported by Lai et al. using membrane-bound variant of the gLuc reporter and a
biotin acceptor peptide [9]. This reporter system allowed multimodal imaging of
bioluminescence and fluorescence in vivo as fluorescence combined with streptavidin could provide fluorescence-mediated tomography imaging simultaneously.
Bioluminescence reporter system has several merits in small animal imaging due
to high sensitivity, specificity for EVs as well as enabling multimodal imaging as
aforementioned. However, this system has an inherent shortcomings that luminescence signals are attenuated in deep organs and that no substrates of luciferin or
coelenterazine for these luciferases are allowed for the administration to humans.
Moreover, relatively complicated labeling method compared with lipophilic dyes
limits the high throughput in vivo kinetics study of various EVs extracted from
different cell types.
Furthermore, similarly to fluorescence reporter imaging, the bioluminescent
signal depends on the reporter protein expression [7, 15]. These labeling procedures
are complicated compared to those of fluorescence dyes, which limits the study of
biodistribution and fate of EVs in various cells under different conditions in vivo.
8.1.4 Radionuclide Imaging
Briefly, radionuclide imaging of EVs is sensitive and feasible to be extended to the
human and clinical applications. Details of radiolabeled EVs are introduced in
previous chapter. Recently developed
99m Tc-labeled EVs used
99m
Tc-HMPAO and
99m Tc-tricarbonyl. They used direct labeling of EVs [24, 25]. Another
125 I-labeled
EVs employed a reporter system. EVs with a fusion protein of streptavidin and
156
H. Choi and D. S. Lee