Characterization of two-photon point-spread function in turbid medium by direct measurements, multicolor imaging, and blind deconvolution

Chen-Yuan Dong; Eric A. Bevan; Lily Laiho Hsu; Karsten Koenig; Peter T. C. So

doi:10.1117/12.424538

24 April 2001 Characterization of two-photon point-spread function in turbid medium by direct measurements, multicolor imaging, and blind deconvolution

Chen-Yuan Dong, Eric A. Bevan, Lily Laiho Hsu, Karsten Koenig, Peter T. C. So

Author Affiliations +

Proceedings Volume 4262, Multiphoton Microscopy in the Biomedical Sciences; (2001) https://doi.org/10.1117/12.424538
Event: BiOS 2001 The International Symposium on Biomedical Optics, 2001, San Jose, CA, United States

Abstract

Over the past decade, scanning fluorescence microscopy based on two-photon excitation has become an important branch of microscopic bio-imaging. Compared to traditional scanning techniques, two-photon microscopy offers a number of distinct advantages. First, scanning of the point-like excitation spot used for imaging results in images with excellent axial depth discrimination. In addition, the limited extent of the excitation volume also limits specimen photo-damage to the focal volume. Finally, the long, near-infixed wavelengths used for sample excitation allow in-depth, non-invasive imaging of optically turbid biological samples. For in-depth imaging, the microscopic objective and often optically heterogeneous biological specimen forms a complex system. To optimize imaging quality in two-photon microscopy, an understanding of the point-spread-function (PSF) is essential. In this work, we attempted to characterize the two-photon PSF by two methods: direct imaging of 0.1 ym fluorescent microspheres and multicolor imaging of 2 ym green fluorescent microspheres in a uniform blue fluorescent background. In both measurements, the turbidity of the surrounding medium was varied by changing the concentration of Liposyn III, a scattering component in the specimen. We found that at discrete Liposyn III concentrations between 0 and 2%, the PSF widths were not affected by the amount of scatterers present. However, the imaged contrast continued to degrade as a function of the amount of scatter. This suggests that the broadening of the tail region of the PSF can be the cause of image contrast loss. We will also discuss the possibility of using blind- deconvolution as a method to obtain PSF information in complex biological specimen.

Citation Download Citation

Chen-Yuan Dong, Eric A. Bevan, Lily Laiho Hsu, Karsten Koenig, and Peter T. C. So "Characterization of two-photon point-spread function in turbid medium by direct measurements, multicolor imaging, and blind deconvolution", Proc. SPIE 4262, Multiphoton Microscopy in the Biomedical Sciences, (24 April 2001); https://doi.org/10.1117/12.424538

ACCESS THE FULL ARTICLE

INSTITUTIONAL
Select your institution to access the SPIE Digital Library.

SELECT YOUR INSTITUTION

PERSONAL
Sign in with your SPIE account to access your personal subscriptions or to use specific features such as save to my library, sign up for alerts, save searches, etc.

PERSONAL SIGN IN

No SPIE Account? Create one

PURCHASE THIS CONTENT

SUBSCRIBE TO DIGITAL LIBRARY

50 downloads per 1-year subscription

Members: $195

Non-members: $335 ADD TO CART

25 downloads per 1 - year subscription

Members: $145

Non-members: $250 ADD TO CART

PURCHASE SINGLE ARTICLE

Includes PDF, HTML & Video, when available

Members: $17.00

Non-members: $21.00 ADD TO CART

PROCEEDINGS
9 PAGES

DOWNLOAD PAPER SAVE TO MY LIBRARY

GET CITATION

RIGHTS & PERMISSIONS

Get copyright permission Get copyright permission on Copyright Marketplace

KEYWORDS

Point spread functions

Scattering

Tissues

Two photon excitation microscopy

Deconvolution

Luminescence

Biomedical optics

Show All Keywords

Keywords/Phrases

Search In:

Publication Years