Three-Dimensional Imaging of Pulmonary Fibrotic Foci at the Alveolar Scale Using Tissue-Clearing Treatment with Staining Techniques of Extracellular Matrix

Figure 4

Fluorescence labeling of fibrotic lungs using fluorescence-conjugated tomato lectin. (a) Serial lung sections were stained using DyLight 488-conjugated tomato lectin and Masson’s trichrome technique. (b–d) DyLight 488-conjugated tomato lectin (2.5 mg/kg) was intravenously administered to mice with bleomycin-induced pulmonary fibrosis. After Clear^T2 tissue-clearing treatment, the lungs were observed using confocal microscopy. (b) Three-dimensional images were reconstructed from single-plane images. Each 5 μm stack is 20–120 μm below the lung surface. (c) The image of cross-sections indicated by the red lines in (b). (d) Single-plane images at the indicated depth (μm) from the lung surface. Green fluorescence indicates the localization of tomato lectin. Scale bar is 100 μm.

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