Evaluation of Stromal Riboflavin Penetration Using Slit-Lamp-Based Digital Image Processing in an Ex Vivo...

Area of Science:

• Ophthalmology
• Biomedical Engineering
• Corneal Science

Background:

• Transepithelial corneal cross-linking (CXL) aims to strengthen the cornea.
• Optimizing riboflavin penetration into the corneal stroma is crucial for CXL efficacy.
• Current methods for assessing riboflavin levels can be invasive or lack precision.

Purpose of the Study:

• To investigate how different iontophoresis current intensities affect stromal riboflavin penetration during transepithelial CXL.
• To develop and validate a simple, quantitative digital slit-lamp biomicroscopy workflow for assessing stromal riboflavin.
• To establish slit-lamp fluorescence as a noninvasive surrogate for measuring stromal riboflavin concentration.

Main Methods:

• Ex vivo bovine corneas were treated with iontophoresis at varying currents (1, 3, 5 mA) or topical riboflavin.
• Digital slit-lamp images were captured and analyzed using a Python-OpenCV workflow to generate a fluorescence score (FS).
• Stromal riboflavin concentration was determined spectrophotometrically for validation against the FS.

Main Results:

• Stromal fluorescence score (FS) increased in a current-dependent manner with iontophoresis.
• Higher iontophoresis currents (up to 5 mA) resulted in greater and more homogeneous stromal riboflavin loading.
• A strong linear correlation (R² = 0.96) was found between the FS and spectrophotometrically measured riboflavin concentration.

Conclusions:

• The developed digital slit-lamp analysis is a simple, objective, and reproducible method for quantifying stromal riboflavin fluorescence.
• Slit-lamp-derived fluorescence serves as an accurate, noninvasive surrogate for assessing stromal riboflavin content.
• This approach holds potential for real-time monitoring and optimization of transepithelial CXL protocols following in vivo validation.