Biology subjects

Piestun, R.

Publications and source records attributed to Piestun, R..

3 recordsLinked to original sources

bioRxiv2021

Characterizing and correcting camera noises in back-illuminated sCMOS cameras

With promising properties of fast imaging speed, large field-of-view, relative low cost and many others, back-illuminated sCMOS cameras have been receiving intensive attentions for low-light imaging in the past several years. However, due to the pixel-to-pixel difference of camera noises (called noise non-uniformity) in sCMOS cameras, researchers may hesitate to use them in some application fields, and sometimes wonder whether they should optimize the noise non-uniformity of their sCMOS cameras before using them in a specific application scenario. In this paper, we systematically characterize the impact of different types of sCMOS noises on image quality and perform corrections to these sCMOS noises. We verify that it is possible to use appropriate correction methods to push the non-uniformity of major camera noises, including readout noise, offset, and photon response, to a satisfactory level for conventional microscopy and single molecule localization microscopy. We further find out that, after these corrections, global read noise becomes a major concern that limits the imaging performance of back-illuminated sCMOS cameras. We believe this study provides new insights into the understanding of camera noises in back-illuminated sCMOS cameras, and also provides useful information for future development of this promising camera technology.

bioengineering↗

bioRxiv2019

3D super-resolution imaging using a generalized and scalable progressive refinement method on sparse recovery (PRIS)

Within the family of super-resolution (SR) fluorescence microscopy, single-molecule localization microscopies (PALM[1], STORM[2] and their derivatives) afford among the highest spatial resolution (approximately 5 to 10 nm), but often with moderate temporal resolution. The high spatial resolution relies on the adequate accumulation of precise localizations of bright fluorophores, which requires the bright fluorophores to possess a relatively low spatial density. Several methods have demonstrated localization at higher densities in both two dimensions (2D)[3, 4] and three dimensions (3D)[5-7]. Additionally, with further advancements, such as functional super-resolution[8, 9] and point spread function (PSF) engineering with[8-11] or without[12] multi-channel observations, extra information (spectra, dipole orientation) can be encoded and recovered at the single molecule level. However, such advancements are not fully extended for high-density localizations in 3D. In this work, we adopt sparse recovery using simple matrix/vector operations, and propose a systematic progressive refinement method (dubbed as PRIS) for 3D high-density reconstruction. Our method allows for localization reconstruction using experimental PSFs that include the spatial aberrations and fingerprint patterns of the PSFs[13]. We generalized the method for PSF engineering, multi-channel and multi-species observations using different forms of matrix concatenations. Reconstructions with both double-helix and astigmatic PSFs, for both single and biplane settings are demonstrated, together with the recovery capability for a mixture of two different color species.

bioinformatics↗

bioRxiv2017

Deep brain fluorescence imaging with minimally invasive ultra-thin optical fibers

A major open challenge in neuroscience is the ability to measure and perturb neural activity in vivo from well-defined neural sub-populations at cellular resolution anywhere in the brain. However, limitations posed by scattering and absorption prohibit non-invasive (surface) multiphoton approaches1,2 for deep (>2mm) structures, while Gradient Refreactive Index (GRIN) endoscopes2-4 are thick and cause significant damage upon insertion. Here, we demonstrate a novel microendoscope to image neural activity at arbitrary depths via an ultrathin multimode optical fiber (MMF) probe that is 5-10X thinner than commercially available microendoscopes. We demonstrate micron-scale resolution, multispectral and volumetric imaging. In contrast to previous approaches1,5-8 we show that this method has an improved acquisition speed that is sufficient to capture rapid neuronal dynamics in-vivo in rodents expressing a genetically encoded calcium indicator. Our results emphasize the potential of this technology in neuroscience applications and open up possibilities for cellular resolution imaging in previously unreachable brain regions.

neuroscience↗