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Rachel Redekop, Frank Corapi, Laura Emptage, Monika Kitor, Melanie C W Campbell; Auto-fluorescence signals of pure amyloid-β and presumed retinal amyloid deposits. Invest. Ophthalmol. Vis. Sci. 2018;59(9):4320.
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Alzheimer’s disease (AD) is neurodegenerative disease and brain amyloid-β (Aβ) is a biomarker for the disease. We and others have found deposits containing Aβ in the retinas of animal models. In humans, we have shown these retinal deposits are predictive of AD severity. Aβ is known to auto-fluoresce for excitation in ultraviolet (UV) and we have reported two photon auto-fluorescence in retinal deposits. There is controversy as to whether amyloid auto-fluorescence is visible in the living eye. Here, auto-fluorescence of pure Aβ is compared to that of presumed retinal amyloid deposits in a canine model.
Aβ 1-42 was purchased from rPeptide in monomer form. The Aβ was incubated in HEPES buffer at 37°C for 89 hours. Fluorescence intensity of the Aβ and control HEPES solutions, before and after incubation was measured at excitations of 360 nm and 480 nm. Ten 50 μL aliquots were deposited on slides and cover slipped. The remaining 10 samples were stained with Thioflavin-T and deposited in 100 μL aliquots. Two retinas from the canine model of AD (with cognitive impairment) were flat mounted in quarters unstained. Pure Aβ 1-42 deposits and the retinas were imaged using both wavelengths, using a microscope fitted with a polarimeter.
Both pure and retinal amyloid deposits had similar, stronger emission signals when excited with 360 nm than with 480 nm light. The strength of auto-fluorescence of unstained Aβ in UV was comparable to the Thioflavin signals from the stained Aβ. Amyloid auto-fluorescence in retinal deposits was substantially stronger than retinal tissue auto-fluorescence in UV but was not distinguishable from tissue auto-fluorescence in 480 nm. There was 100% agreement between Aβ auto-fluorescence and polarisation signals for the pure Aβ deposits.
Strong auto-fluorescence at 360 nm excitation is consistent with the two photon auto-fluorescence we reported for presumed retinal amyloid deposits. A signal indistinguishable from background tissue is consistent with the lack of visibility of retinal deposits reported by us in the living canine eye at 480 nm excitation. Given the absorption of UV and blue light by anterior ocular structures, two photon techniques for imaging auto-fluorescence of amyloid deposits in the living eye may be needed. Polarization and auto-fluorescence are potentially important as non-invasive imaging techniques for amyloid as an important biomarker of AD.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
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