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Published March 2014 | Published
Journal Article Open

Integrated photoacoustic, confocal, and two-photon microscope


The invention of green fluorescent protein and other molecular fluorescent probes has promoted applications of confocal and two-photon fluorescence microscopy in biology and medicine. However, exogenous fluorescence contrast agents may affect cellular structure and function, and fluorescence microscopy cannot image nonfluorescent chromophores. We overcome this limitation by integrating optical-resolution photoacoustic microscopy into a modern Olympus IX81 confocal, two-photon, fluorescence microscope setup to provide complementary, label-free, optical absorption contrast. Automatically coregistered images can be generated from the same sample. Imaging applications in ophthalmology, developmental biology, and plant science are demonstrated. For the first time, in a familiar microscopic fluorescence imaging setting, this trimodality microscope provides a platform for future biological and medical discoveries.

Additional Information

© 2014 SPIE. Paper 130894R received Dec. 19, 2013; revised manuscript received Jan. 27, 2014; accepted for publication Jan. 28, 2014; published online Mar. 3, 2014. The authors thank Ms. DeGenova Sarah for her assistance in preparation of zebrafish samples and Professor James Ballard for his technical writing support. We thank Dr. Lijun Ma and Dr. Yu Wang for their help on the LabVIEW program. Institutional support from the Nano Research Facility of Washington University in St. Louis is appreciated. This work was sponsored in part by National Institutes of Health (NIH) grants 1S10RR028864, K99AR062530, DP1 EB016986 (NIH Director's Pioneer Award), and R01 CA159959. L. V. Wang has financial interests in Microphotoacoustics Inc. and Endra Inc., which did not support this work.

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