Related Experiment Video
Updated: Jul 17, 2026
Transpupillary Two-Photon In Vivo Imaging of the Mouse Retina
Published on: February 13, 2021
Two-photon imaging microscopy of non-stained human epiretinal membranes
Juan M Bueno1, Xavier Valldeperas2
1Laboratorio de Óptica, Instituto Universitario de Investigación en Óptica y Nanofísica, Campus de Espinardo (Ed. 34), CEIR Campus Mare Nostrum (CMN), Universidad de Murcia, 30100 Murcia, Spain.
Abstract:
Epiretinal membranes (ERMs) are composed of cellular and extracellular matrix elements. They are divided into idiopathic (iERM), of unknown origin, and non-idiopathic (non-iERM), usually associated with proliferative retinopathies and retinal detachment surgery. ERMs have been visualized with complex and highly time-consuming multi-labelling methods, which characterization has been mainly qualitative. Since ERM components may depend on their genesis, two-photon imaging microscopy might be a proper tool to objectively differentiate iERM from non-iERM. In this study we propose the combination of two-photon excitation fluorescence (TPEF) and second harmonic generation (SHG) signals to explore and quantify differences between both types of non-stained human ERMs. An index computed from pairs of TPEF-SHG images and an analysis of the collagen-based tissue spatial arrangement have been employed for this purpose.
Insights
Two-photon imaging microscopy differentiates epiretinal membranes (ERMs) based on origin. This technique quantifies differences between idiopathic (iERM) and non-idiopathic (non-iERM) ERMs using autofluorescence and collagen signals.
Area of Science:
- Ophthalmology
- Biomedical Imaging
- Cell Biology
Background:
- Epiretinal membranes (ERMs) are cellular and extracellular matrix formations.
- ERMs are classified as idiopathic (iERM) or non-idiopathic (non-iERM), with distinct origins.
- Current visualization methods for ERMs are time-consuming and primarily qualitative.
Purpose of the Study:
- To explore and quantify differences between non-stained human iERM and non-iERM.
- To assess the potential of two-photon imaging microscopy for objective ERM characterization.
- To differentiate ERM types based on their genesis using advanced imaging techniques.
Main Methods:
- Utilized two-photon excitation fluorescence (TPEF) and second harmonic generation (SHG) signals.
- Applied TPEF-SHG imaging to non-stained human ERM samples.
- Developed a quantitative index from TPEF-SHG image pairs and analyzed collagen spatial arrangement.
Main Results:
- Established an index to differentiate between iERM and non-iERM based on TPEF-SHG signals.
- Quantified differences in collagen-based tissue spatial arrangement between ERM types.
- Demonstrated the potential of label-free two-photon imaging for ERM characterization.
Conclusions:
- Two-photon imaging microscopy, combining TPEF and SHG, offers an objective method to differentiate ERM types.
- The developed index and collagen analysis provide quantitative insights into ERM composition and origin.
- This technique may improve the understanding and diagnosis of ERM-related conditions.
