6 Conclusion and Future Perspectives
The course of evolution of fluorescent proteins from the first known member GFP to
a completely new family of fluorescent proteins spanning across the visible spectra
has emerged in relatively short time span. The rapid growth and applications of the
FP repertoire for in vivo imaging continuously shed light on studying crucial cellular
events which play a vital role in development and progression of diseases. With
the present capabilities of FPs, multicolor labeling of proteins and nucleic acids;
tracking of protein movements, interactions, activities, degradation, organelle motility, and fusion-fission events; and monitoring of promoter activation, as well as
multiparameter imaging of various cellular processes, including changes in concentration of signal molecules, changes of membrane potential and cell state, etc., can be
deduced efficiently. Although extensive variants of biosensors for specific applications have been developed, it is expected that further improvements in brightness,
photostability, maturation rate, pH stability, and performance in fusions will gain
priority in the future. Even though a significant progress has been made in the
demonstration of novel fluorescent protein-based biosensors for in vivo models,
further research is required to establish consistent, reproducible, and reliable imaging
instruments with far-red shifted fluorescent proteins, and animal models for disease
investigations. Although rapid strides have been made in this field, further improvements on deep tissue imaging with higher sensitivity and long-term noninvasive
imaging would open wide range of applications for biosensors. Addressing the
drawback of loss of fluorescent proteins during tissue fixation or subsequent
processing can also improve ex vivo histopathological analysis of tissues from
transgenic animals.
Acknowledgments We would like to thank the Canary Center at Stanford, Department of
Radiology, for providing facility and resources. We also thank SCi3 Small Animal Imaging Service
Center, Stanford University School of Medicine, for providing imaging facilities and data analysis
support. We acknowledge Dr. Sanjiv Sam Gambhir, Chair of the Department of Radiology,
Stanford University, for his constant support and help.
Compliance with Ethical Standards
Conflicts of Interest There are no actual or potential conflicts of interest in regard to this chapter.
Funding This research was supported by NIH R01CA209888 and NIH R21EB022298. This work
was also in part supported by the Center for Cancer Nanotechnology Excellence for Translational
Diagnostics (CCNE-TD) at Stanford University through an award (grant no. U54 CA199075) from
the National Cancer Institute (NCI) of the National Institutes of Health (NIH).
Ethical Approval All applicable international, national, and/or institutional guidelines for the care
and use of animals were followed.
Applications of Fluorescent Protein-Based Sensors in Bioimaging
175
The course of evolution of fluorescent proteins from the first known member GFP to
a completely new family of fluorescent proteins spanning across the visible spectra
has emerged in relatively short time span. The rapid growth and applications of the
FP repertoire for in vivo imaging continuously shed light on studying crucial cellular
events which play a vital role in development and progression of diseases. With
the present capabilities of FPs, multicolor labeling of proteins and nucleic acids;
tracking of protein movements, interactions, activities, degradation, organelle motility, and fusion-fission events; and monitoring of promoter activation, as well as
multiparameter imaging of various cellular processes, including changes in concentration of signal molecules, changes of membrane potential and cell state, etc., can be
deduced efficiently. Although extensive variants of biosensors for specific applications have been developed, it is expected that further improvements in brightness,
photostability, maturation rate, pH stability, and performance in fusions will gain
priority in the future. Even though a significant progress has been made in the
demonstration of novel fluorescent protein-based biosensors for in vivo models,
further research is required to establish consistent, reproducible, and reliable imaging
instruments with far-red shifted fluorescent proteins, and animal models for disease
investigations. Although rapid strides have been made in this field, further improvements on deep tissue imaging with higher sensitivity and long-term noninvasive
imaging would open wide range of applications for biosensors. Addressing the
drawback of loss of fluorescent proteins during tissue fixation or subsequent
processing can also improve ex vivo histopathological analysis of tissues from
transgenic animals.
Acknowledgments We would like to thank the Canary Center at Stanford, Department of
Radiology, for providing facility and resources. We also thank SCi3 Small Animal Imaging Service
Center, Stanford University School of Medicine, for providing imaging facilities and data analysis
support. We acknowledge Dr. Sanjiv Sam Gambhir, Chair of the Department of Radiology,
Stanford University, for his constant support and help.
Compliance with Ethical Standards
Conflicts of Interest There are no actual or potential conflicts of interest in regard to this chapter.
Funding This research was supported by NIH R01CA209888 and NIH R21EB022298. This work
was also in part supported by the Center for Cancer Nanotechnology Excellence for Translational
Diagnostics (CCNE-TD) at Stanford University through an award (grant no. U54 CA199075) from
the National Cancer Institute (NCI) of the National Institutes of Health (NIH).
Ethical Approval All applicable international, national, and/or institutional guidelines for the care
and use of animals were followed.
Applications of Fluorescent Protein-Based Sensors in Bioimaging
175