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Exp Eye Res. 2018 Sep;174:51-58. doi: 10.1016/j.exer.2018.05.018. Epub 2018 May 19.

Two-photon microscopy of fungal keratitis-affected rabbit cornea ex vivo using moxifloxacin as a labeling agent.

Author information

1
Department of Mechanical Engineering, Pohang University of Science and Technology, 77 Cheongam-ro, Nam-gu, Pohang, Gyeoungbuk, 37673, Republic of Korea.
2
Division of Integrative Biosciences and Biotechnology, Pohang University of Science and Technology, 77 Cheongam-ro, Nam-gu, Pohang, Gyeoungbuk, 37673, Republic of Korea.
3
Department of Ophthalmology, Ulsan College of Medicine, Asan Medical Center, 88 Olympic-ro 43-gil, Songpa-gu, Seoul, 05505, Republic of Korea.
4
Department of Chemistry, Pohang University of Science and Technology, 77 Cheongam-ro, Nam-gu, Pohang, Gyeoungbuk, 37673, Republic of Korea.
5
Department of Ophthalmology, Ulsan College of Medicine, Asan Medical Center, 88 Olympic-ro 43-gil, Songpa-gu, Seoul, 05505, Republic of Korea. Electronic address: joon@amc.seoul.kr.
6
Department of Mechanical Engineering, Pohang University of Science and Technology, 77 Cheongam-ro, Nam-gu, Pohang, Gyeoungbuk, 37673, Republic of Korea; Division of Integrative Biosciences and Biotechnology, Pohang University of Science and Technology, 77 Cheongam-ro, Nam-gu, Pohang, Gyeoungbuk, 37673, Republic of Korea. Electronic address: kiheankim@postech.ac.kr.

Abstract

Two-photon microscopy (TPM) is a three dimensional (3D) microscopic technique based on nonlinear two-photon fluorescence, which has been tested as an alternative to reflectance confocal microscopy (RCM) for detecting fungal keratitis via optical imaging. Although TPM provided images with better contrast than RCM for fungal keratitis, its imaging speed was relatively low because of weak intrinsic signal. Moxifloxacin, a Food and Drug Administration (FDA)-approved antibiotic, was recently used as a cell-labeling agent for TPM. In this study, moxifloxacin was used to label fungal cells for TPM imaging of fungal keratitis models. Fungal cell suspensions and ex vivo fungal keratitis-affected rabbit corneas were prepared using two types of fungal pathogens, Aspergillus fumigatus and Candida albicans, and TPM imaging was performed both with and without moxifloxacin treatment. Fungal cells with enhanced fluorescence were clearly visible by TPM of moxifloxacin-treated fungal cell suspensions. TPM of moxifloxacin-treated fungal keratitis rabbit corneas revealed both the infecting fungal cells and corneal cells similar to those observed in TPM without moxifloxacin treatment, albeit with approximately 10-times enhanced fluorescence. Fungal cells were distinguished from corneal cells on the basis of their distinct morphologies. Thus, TPM with moxifloxacin labeling might be useful for the detection of fungal keratitis at the improved imaging speed.

KEYWORDS:

Fungal keratitis; High-resolution 3D imaging; Moxifloxacin; Rabbit cornea; Two-photon microscopy

PMID:
29787746
DOI:
10.1016/j.exer.2018.05.018
[Indexed for MEDLINE]

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