Single objective light-sheet microscopy for high-speed whole-cell 3D super-resolution

Abstract

Abstract:We have developed a method for performing light-sheet microscopy with a single high numerical aperture lens by integrating reflective side walls into a microfluidic chip. These 45°side walls generate light-sheet illumination by reflecting a vertical light-sheet into the focal plane of the objective. Light-sheet illumination of cells loaded in the channels increases image quality in diffraction limited imaging via reduction of out-of-focus background light. Single molecule super-resolution is also improved by the decreased background resulting in better localization precision and decreased photo-bleaching, leading to more accepted localizations overall and higher quality images. Moreover, 2D and 3D single molecule superresolution data can be acquired faster by taking advantage of the increased illumination intensities as compared to wide field, in the focused light-sheet. 94, 4957-4970 (2008).

Optical Imaging Approaches to Monitor Static and Dynamic Cell‐on‐Chip Platforms: A Tutorial Review

“…Interested readers are referred to another reference for more information . Light sheet microscopy could be employed for 3D imaging of a wide range of biological samples, from cells (Figure d) to small tissues like spheroids …”

“…d) 3D light sheet microscopy image of a collection of HeLa cells labeled in their mitochondria. Reproduced with permission . Copyright 2016, Optical Society of America.…”

Microfluidic Based Optical Microscopes on Chip

“…In particular, micro‐fabricated reflective components have been developed and tailored for light sheet illumination by Galland et al and Zagato et al : in these implementations a laser beam is reflected horizontally on the sample by a 45° mirror and forms the light sheet in the image plane of the detection objective of an upright or inverted microscope. The buffer solution can be further exchanged automatically by a microfluidic system coupled with the chip that incorporates the mirrors . On the basis of this interest for high resolution devices, we foresee that new solutions that fully integrate super‐resolution light sheet microscopy on a chip will be conceived and implemented in the next future.…”

Imaging neural events in zebrafish larvae with linear structured illumination light sheet fluorescence microscopy

“…[7][8][9][10] Various configurations of LSFM have been developed and applied to different sizes of samples. These include single cell imaging, [11][12][13] small multicellular samples, such as C. elegans, D. melanogaster, and zebrafish embryos, 3,14,15 samples which are hundreds of microns in size, such as C. elegans 16 and zebrafish larvae, 7 and even samples up to a few centimeters in size, such as whole mouse embryos 17 and human prostate tissue. 18 Even though LSFM provides a high temporal resolution and excellence in optical sectioning, the resulting images are greatly affected by the thickness of the sample, especially when applied to large and semiopaque samples.…”