Autofluorescence imaging of human RPE cell granules using structured illumination microscopy

• Verfasst von: Ach, Thomas [VerfasserIn]
• Verfasst von: Best, Gerrit [VerfasserIn]
• Verfasst von: Roßberger, Sabrina [VerfasserIn]
• Verfasst von: Cremer, Christoph [VerfasserIn]
• Verfasst von: Dithmar, Stefan [VerfasserIn]
• Titel: Autofluorescence imaging of human RPE cell granules using structured illumination microscopy
• Verf.angabe: Thomas Ach, Gerrit Best, Sabrina Rossberger, Rainer Heintzmann, Christoph Cremer, Stefan Dithmar
• E-Jahr: 2012
• Jahr: July 14, 2012
• Umfang: 4 S.
• Fussnoten: Published online first 3 July 2012 ; Gesehen am 07.09.2018
• Titel Quelle: Enthalten in: British journal of ophthalmology
• Ort Quelle: London : BMJ Publ. Group, 1917
• Jahr Quelle: 2012
• Band/Heft Quelle: 96(2012), 8, Seite 1141-1144
• ISSN Quelle: 1468-2079
• DOI: doi:10.1136/bjophthalmol-2012-301547
• Datenträger: Online-Ressource
• Sprache: eng
• Sach-SW: Autofluorescence
• Sach-SW: lipofuscin
• Sach-SW: melanolipofuscin
• Sach-SW: structured illumination
• K10plus-PPN: 1580766765

Abstract

Background/aims To characterise single autofluorescent (AF) granules in human retinal pigment epithelium (RPE) cells using structured illumination microscopy (SIM). Methods: Morphological characteristics and autofluorescence behaviour of lipofuscin (LF) and melanolipofuscin (MLF) granules of macular RPE cells (66-year-old donor) were examined with SIM using three different laser light excitation wavelengths (488, 568 and 647 nm). High-resolution images were reconstructed and exported to Matlab R2009a (The Mathworks Inc, Natick, MA, USA) to determine accurate size and emission intensities of LF and MLF granules. Results: SIM doubles lateral resolution compared with conventionally used wide-field microscopy and allows visualisation of intracellular structures down to 110 nm lateral resolution. AF patterns were examined in 133 LF and 27 MLF granules. LF granules (968±220 nm) were significantly smaller in diameter than MLF granules (1097±110 nm; p<0.001). LF granules showed an inhomogeneous intragranular pattern, and the average intensity negatively correlated with the size of these granules when excited at 647 nm. The autofluorescence of MLF granules was more homogeneous, but shifted towards higher excitation wavelengths in the centre of the granules. Conclusion: SIM is a useful tool for examining AF signals within single LF and MLF granules in RPE cells. This allows new insights into RPE autofluorescence patterns.