Dynamical measurements of motion behavior of free fluorescent sphere using the wide field temporal focusing microscopy with astigmatism method (Conference Presentation)

Chi-Hsiang Lien; Chun-Yu Lin; Shean-Jen Chen; Fan-Ching Chien

doi:10.1117/12.2252323

24 April 2017 Dynamical measurements of motion behavior of free fluorescent sphere using the wide field temporal focusing microscopy with astigmatism method (Conference Presentation)

Chi-Hsiang Lien, Chun-Yu Lin, Shean-Jen Chen, Fan-Ching Chien

Author Affiliations +

Proceedings Volume 10069, Multiphoton Microscopy in the Biomedical Sciences XVII; 100691V (2017) https://doi.org/10.1117/12.2252323
Event: SPIE BiOS, 2017, San Francisco, California, United States

Abstract

A three-dimensional (3D) single fluorescent particle tracking strategy based on temporal focusing multiphoton excitation microscopy (TFMPEM) combined with astigmatism imaging is proposed for delivering nanoscale-level axial information that reveals 3D trajectories of single fluorospheres in the axially-resolved multiphoton excitation volume without z-axis scanning. It provides the dynamical ability by measuring the diffusion coefficient of fluorospheres in glycerol solutions with a position standard deviation of 14 nm and 21 nm in the lateral and axial direction and a frame rate of 100 Hz. Moreover, the optical trapping force based on the TFMPEM is minimized to avoid the interference in the tracing measurements compared to that in the spatial focusing MPE approaches. Therefore, we presented a three dimensional single particle tracking strategy to overcome the limitation of the time resolution of the multiphoton imaging using fast frame rate of TFMPEM, and provide three dimensional locations of multiple particles using an astigmatism method.

Conference Presentation

Citation Download Citation

Chi-Hsiang Lien, Chun-Yu Lin, Shean-Jen Chen, and Fan-Ching Chien "Dynamical measurements of motion behavior of free fluorescent sphere using the wide field temporal focusing microscopy with astigmatism method (Conference Presentation)", Proc. SPIE 10069, Multiphoton Microscopy in the Biomedical Sciences XVII, 100691V (24 April 2017); https://doi.org/10.1117/12.2252323

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KEYWORDS

3D image processing

Monochromatic aberrations

Motion measurement

Microscopy

Particles

3D metrology

Multiphoton microscopy

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