Aberration correction in epi-fluorescence microscopy using unknown speckle illumination

• Published in: Optics express Vol. 32; no. 21; p. 37658
• Main Authors: Premillieu, Evolene, Caravaca-Aguirre, Antonio M., Labouesse, Simon, Irsch, Kristina, Piestun, Rafael
• Format: Journal Article
• Language: English
• Published: 07-10-2024
• ISSN: 1094-4087; 1094-4087
• DOI: 10.1364/OE.526148

Diffraction-limited imaging in epi-fluorescence microscopy remains a challenge when sample aberrations are present or when the region of interest rests deep within an inhomogeneous medium. Adaptive optics is an attractive solution, albeit with limited field of view and requiring relatively complicated systems. Alternatively, reconstruction algorithms have been developed over the years to correct for aberrations. Unfortunately, purely post-processing techniques tend to be ill-posed and provide only incremental improvements in image quality. Here, we report a computational optical approach using unknown speckle illumination and a matched reconstruction algorithm to correct for aberrations and reach or surpass diffraction limited resolution. The data acquisition is performed by shifting an unknown speckle pattern with respect to a fluorescent object. A key advantage is that the speckle statistics are preserved upon propagation through the aberrations, which avoids the double pass of information through the aberrating medium typical of epi-fluorescence microscopy. The method recovers simultaneously a high-resolution image, the point spread function of the system that contains the aberrations, the speckle illumination pattern, and the shift positions.