Fig 1.
Representative atropine mass spectrum.
(A) Mass spectra in the cornea, the back of the eye (B), and the back of the lens (C). The molecular formula of atropine is C17H23NO3 and its mass peak ([M+H]+) was observed at m/z 290.17474 (theoretical m/z: 290.17507).
Fig 2.
Atropine distribution in a whole-eye section by MALDI-IMS at low magnification and low spatial resolution (100 μm).
(A) Optical image of hematoxylin-eosin-stained whole-eye section. (B) Atropine distribution image obtained by IMS. We corrected the image contrast to the anterior segment by adjusting the full-intensity threshold of the result filter. (C) Atropine distribution image obtained by IMS. We corrected the image contrast to the posterior segment by adjusting the full-intensity threshold of the result filter. AC: anterior chamber; L: lens; V: vitreous body; C: cornea; S: sclera. Scale bar = 2 mm.
Fig 3.
Atropine distribution in a whole-eye section by MALDI-IMS at high magnification and high spatial resolution (10 μm).
IMS revealed atropine distribution around the corneal apex (A, B) and in the inferior cornea-sclera junction (C, D). This image was acquired after low spatial resolution (100 μm) imaging analysis (Fig 2). Consequently, data were missing where the sample had been previously irradiated with laser. Yellow lines indicate organ borders. AC: anterior chamber; EP: epithelium; ST: stroma; EN: endothelium; CB: ciliary body; C-S: cornea-sclera; R-CH: retina choroid; V: vitreous body. Scale bar = 200 μm.
Fig 4.
Line scan data along the outer perimeter of the eyeball section.
(A) Optical image of a hematoxylin-eosin-stained whole-eye section. Numbers indicate distances (mm) from the zero position. (B) Atropine distribution image obtained by IMS. (C) Magnified segmentation image in the corneal region. The corneal apex (0 mm) was set as the graph origin. (D) Line scan data of the superior (closed square) and inferior (open diamond) halves along the outer perimeter of the eyeball section. Signal intensities were compared between superior and inferior points equidistant from the center. Significantly higher in the superior region: † P < 0.05, †† P < 0.01; significantly higher in the inferior region: * P < 0.05, ** P < 0.01. Mann-Whitney U test. AC: anterior chamber; L: lens; V: vitreous body; C: cornea; S: sclera. Scale bars: (A), (B) = 2 mm, (C) = 100 μm.