TY - JOUR
T1 - Scanning two-photon microscopy with upconverting lanthanide nanoparticles via Richardson-Lucy deconvolution
AU - Gainer, Christian F.
AU - Utzinger, Urs
AU - Romanowski, Marek
N1 - Funding Information:
The authors would like to acknowledge Faith Rice for providing tissue samples for these experiments. This work was supported by NIH grants EB000809, CA120350, CA109385, and RR023737 as well as the TRIF imaging program at the University of Arizona.
PY - 2012/7
Y1 - 2012/7
N2 - The use of upconverting lanthanide nanoparticles in fast-scanning microscopy is hindered by a long luminescence decay time, which greatly blurs images acquired in a nondescanned mode. We demonstrate herein an image processing method based on Richardson-Lucy deconvolution that mitigates the detrimental effects of their luminescence lifetime. This technique generates images with lateral resolution on par with the system's performance, ∼1.2 μm, while maintaining an axial resolution of 5 μm or better at a scan rate comparable with traditional two-photon microscopy. Remarkably, this can be accomplished with near infrared excitation power densities of 850 W/cm 2, several orders of magnitude below those used in two-photon imaging with molecular fluorophores. By way of illustration, we introduce the use of lipids to coat and functionalize these nanoparticles, rendering them water dispersible and readily conjugated to biologically relevant ligands, in this case epidermal growth factor receptor antibody. This deconvolution technique combined with the functionalized nanoparticles will enable three-dimensional functional tissue imaging at exceptionally low excitation power densities.
AB - The use of upconverting lanthanide nanoparticles in fast-scanning microscopy is hindered by a long luminescence decay time, which greatly blurs images acquired in a nondescanned mode. We demonstrate herein an image processing method based on Richardson-Lucy deconvolution that mitigates the detrimental effects of their luminescence lifetime. This technique generates images with lateral resolution on par with the system's performance, ∼1.2 μm, while maintaining an axial resolution of 5 μm or better at a scan rate comparable with traditional two-photon microscopy. Remarkably, this can be accomplished with near infrared excitation power densities of 850 W/cm 2, several orders of magnitude below those used in two-photon imaging with molecular fluorophores. By way of illustration, we introduce the use of lipids to coat and functionalize these nanoparticles, rendering them water dispersible and readily conjugated to biologically relevant ligands, in this case epidermal growth factor receptor antibody. This deconvolution technique combined with the functionalized nanoparticles will enable three-dimensional functional tissue imaging at exceptionally low excitation power densities.
KW - Lanthanide
KW - Nanoparticle
KW - Optical sectioning
KW - Two-photon microscopy
KW - Upconversion
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U2 - 10.1117/1.JBO.17.7.076003
DO - 10.1117/1.JBO.17.7.076003
M3 - Article
C2 - 22894486
AN - SCOPUS:84870916161
SN - 1083-3668
VL - 17
JO - Journal of biomedical optics
JF - Journal of biomedical optics
IS - 7
M1 - 076003
ER -