title: Quantifying the backscattering of second harmonic generation in tissues with confocal multiphoton microscopy.
abstract: The backward second harmonic generation (SHG) in mouse tissues is studied with a confocal multiphoton microscopy system. The total backward collected SHG (B-SHG) consists of the backward generated SHG and the backward-scattered forward-generated SHG (BS-SHG), which can be modeled by a Gaussian and a uniform distribution, respectively, at the confocal pinhole plane. By varying the pinhole size with a series of collection fibers, the proportion of the BS-SHG to the B-SHG and the proportion of BS-SHG to the forward generated SHG can be obtained. The approach is first validated by Monte Carlo simulation. It is then applied to two types of mouse tissues: mouse tail tendon and Achilles tendon. It is found that the BS-SHG contributes less to the B-SHG for the tail tendon than Achilles tendon with thicknesses of ~300 μm. With the thickness of the Achilles tendon tissue increased to 1000 μm but the focal plane kept at the same depth, as high as ~10% of the total forward SHG is backscattered and collected. The results indicate that BS-SHG may not be the major source of B-SHG in the tail tendon, but it may be the major source in the Achilles tendon. These methods and results provide a noninvasive method and supporting information for investigating the generation mechanism of SHG and help with optimizing backward SHG microscopy and spectroscopy measurements.
authors.affiliation: University of British Columbia, Department of Electrical and Computer Engineering, Vancouver, BC V6T 1Z4, Canada.
authors.forename: Mengzhe
authors.initials: M
authors.lastname: Shen
authors.forename: Yunxian
authors.initials: Y
authors.lastname: Tian
authors.forename: Shau Poh
authors.initials: SP
authors.lastname: Chong
authors.forename: Jianhua
authors.initials: J
authors.lastname: Zhao
authors.forename: Haishan
authors.initials: H
authors.lastname: Zeng
authors.forename: Shuo
authors.initials: S
authors.lastname: Tang
mesh_terms: D000125:Achilles Tendon / Q000737:chemistry; D000162:Acoustics; D000818:Animals; D008297:Male; D051379:Mice; D008809:Mice, Inbred C3H; D018613:Microscopy, Confocal / Q000379:methods*; D036641:Microscopy, Fluorescence, Multiphoton / Q000379:methods*; D009010:Monte Carlo Method; D012815:Signal Processing, Computer-Assisted; D013623:Tail / Q000737:chemistry; D013710:Tendons / Q000737:chemistry