Two-Photon Excited Fluorescence Lifetime Imaging of Tetracycline-Labeled Retinal Calcification

Area of Science:

• Ophthalmology
• Biomedical Imaging
• Retinal Diseases

Background:

• Subretinal pigment epithelial (sub-RPE) calcification is implicated in age-related macular degeneration (AMD) and Pseudoxanthoma elasticum (PXE).
• Early diagnosis of AMD is crucial for timely intervention.
• Current imaging methods for sub-RPE calcification have limitations.

Purpose of the Study:

• To investigate the feasibility of using tetracycline fluorescence lifetime imaging for visualizing sub-RPE calcification.
• To demonstrate that multiphoton infrared excitation can be used for this purpose in post-mortem human retinas.

Main Methods:

• Utilized tetracycline antibiotics as fluorescent stains for calcium-containing minerals in post-mortem human retinas.
• Employed multiphoton (infrared) excitation for fluorescence lifetime imaging.
• Analyzed fluorescence intensity and lifetime changes upon tetracycline binding to minerals.

Main Results:

• Tetracycline binding to sub-RPE minerals increased fluorescence intensity and lifetime.
• Fluorescence lifetimes of tetracyclines differed significantly from natural retinal fluorophores.
• Successful imaging of tetracycline-stained drusen was achieved using fluorescence lifetime contrast with infrared multiphoton excitation.

Conclusions:

• Fluorescence lifetime imaging with tetracycline staining is a promising technique for visualizing sub-RPE calcification.
• This approach may aid in the early detection and diagnosis of AMD and PXE.
• Multiphoton infrared excitation overcomes limitations of shorter excitation wavelengths in retinal imaging.