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Multi-photon Fluorescence Micro-spectroscopy of Plant Tissues

Published online by Cambridge University Press:  02 July 2020

F. J. Kao ,
B. L. Lin  and
P. C. Cheng
F. J. Kao
Affiliation:
Dept. of Physics, Natl. Sun Yat-sen Univ., Kaohsiung, Taiwan, 80424Republic of China
B. L. Lin
Affiliation:
Inst of Molecular Biology, Academia Sinic, Taipei, Taiwan, 11529Republic of China
P. C. Cheng
Affiliation:
AMIL, Dept. of Electrical Eng., State Univ. of New York, Buffalo, NY14260 , USA
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Considering its non-linear nature, two-photon excitation may generate very different spectral response in samples when compared with single photon excitation. It is thus necessary to measure the two-photon spectra of samples, so that the two-photon fluorescence microscopic images can be properly interpreted. Fluorescence spectra obtained from bulk samples may not provide useful information for microscopy. For instance, due to the relatively small contribution to the total fluorescence intensity, a small number of fluorescent particles in a generally fluorescing specimen may escape detection when the spectrum of the specimen as a whole is obtained. Under two-photon excitation, the background noise can be greatly reduced due to the naturally limited excitation volume of focused laser beam. In addition, signals resulted from second harmonic generation (SHG) may be mixed with low level broad-band background autofluorescence which is commonly found in biological specimen. Therefore, measuring fluorescence spectrum from a micro-focused volume is essential for the proper interpretation of multi-photon fluorescence images.

Type
Advances in Multi-Photon Imaging
Information
Microscopy and Microanalysis , Volume 6 , Issue S2: Proceedings: Microscopy & Microanalysis 2000, Microscopy Society of America 58th Annual Meeting, Microbeam Analysis Society 34th Annual Meeting, Microscopical Society of Canada/Societe de Microscopie de Canada 27th Annual Meeting, Philadelphia, Pennsylvania August 13-17, 2000 , August 2000 , pp. 808 - 809
Copyright
Copyright © Microscopy Society of America

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References

References:

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Supported by Academia Sinica (BLL), Natl. Science Council, Rep. of China [NSC-88-2311-B-001-087 (FJK), NSC-88-2311-B-001-087 (BLL), (NSC-88-2311-B-001-087 (PCC)] and Mr. Jin-Mu Huang and Mrs. Li-Ling Huang of Aurum Belle Investment Co. (on behalf of the Ge-An Charity, )(PCC).Google Scholar