Multimodal nonlinear imaging of the human cornea

Area of Science:

• Biomedical Optics
• Ophthalmology
• Microscopy

Background:

• The human cornea and trabecular meshwork possess complex microstructures crucial for vision.
• Nonlinear optical microscopy offers label-free visualization of biological tissues.
• Understanding tissue microstructure is vital for diagnosing and managing ocular diseases.

Purpose of the Study:

• To assess the utility of combined third-harmonic generation (THG), second-harmonic generation (SHG), and two-photon excited fluorescence (2PEF) microscopy.
• To visualize the intrinsic nonlinear optical properties of the human cornea and trabecular meshwork.
• To correlate nonlinear optical signals with tissue microstructure and physiological states.

Main Methods:

• Utilized multiphoton microscopy with a titanium-sapphire laser and optical parametric oscillator.
• Employed THG, SHG, and 2PEF imaging modalities.
• Examined fresh human corneal and corneoscleral tissues using forward and backward detection channels.

Main Results:

• Elucidated THG contrast mechanisms in the cornea, revealing stromal collagen lamellae stacking and distribution.
• THG imaging identified cellular and anchoring structures and indicated water presence in edematous tissue.
• SHG visualized fibrillar collagen, while 2PEF highlighted the trabecular meshwork's elastic component and cellular fluorescence.

Conclusions:

• Combined THG, SHG, and 2PEF microscopy provides high-contrast, specific imaging of the anterior segment's microstructure and physiological status.
• This multimodal nonlinear imaging approach is valuable for assessing corneal diseases and glaucoma.
• The intrinsic optical properties of tissues can be leveraged for label-free diagnostic imaging.