Light Sheet Microscopy

Frequently Asked Questions

Can't find the answer to your question? Feel free to contact us. We will be happy to connect you with a light-sheet microscopy expert who can assist you with any questions you have. 

General

What is light-sheet fluorescence microscopy?

Light-sheet microscopy (LSFM) is a fluorescence imaging method in which a sample is illuminated from the side with a thin sheet of light, enabling inherent optical sectioning. The fluorescence signal of the entire illuminated 2D plane is detected by a full-frame camera. Therefore, LSFM is fast, enables high temporal and 3D-spatial resolution, and restricts photobleaching/phototoxicity, striping artifacts, and other noise.

Light-sheet fluorescence microscopy can be used to image:

  • Live, fixed, or cleared samples;
  • Large samples, even of whole-body specimens;
  • Fast dynamic biological processes; and
  • Live specimens over long periods.

How does light-sheet microscopy work?

Light-sheet fluorescence microscopy optimizes detection (photon-collection) efficiency using a wide-field fluorescence microscope placed perpendicular to the light-sheet laser source. This decouples the excitation beam path from the detection beam path and provides inherent optical sectioning

A light-sheet microscope illuminates the entire 2D focal plane at once, allowing fluorescence signal collection and full-frame camera imaging of that plane. Users can then collect high-speed 3D images with minimal sample damage, even on very large and whole-body specimens.

Watch part 1 of our on-demand webinar "Bruker Microscopes Fill the Gap Between Traditional Fluorescence and Electron Microscopy" to see this question discussed in greater detail.

What are the advantages of using a light sheet microscope compared to a confocal microscope?

Light-sheet microscopy is 100-1000 times faster and provides significantly gentler (less phototoxic) imaging than conventional laser point-scanning techniques like confocal or multiphoton microscopy.

Light-sheet microscopy can thereby image a wider range of samples and events on live, fixed, or cleared samples. This includes challenging large or whole-body specimens and fast dynamic biological processes that are inaccessible by other techniques.

How can light-sheet microscopy improve 3D image resolution of biological tissues?

In fluorescence microscopy, the axial resolution is often lower than the lateral resolution. In light-sheet fluorescence microscopy, this anisotropic resolution can obstruct data analysis and cause blurriness in the axial imaging plane, especially when imaging biological structures that are smaller than the axial resolution of the microscope (e.g., membranes or subcellular structures).

A dual view light-sheet microscope allows samples to be imaged from different angles, either sequentially, by 90° rotation of the capillary, or simultaneously. Imaging the sample from different angles with multiview fusion and deconvolution algorithms allows for isotropic spatial resolution, meaning the resolution is the same in all three dimensions, with high speed and minimal phototoxicity. Consequently, any of these methods of dual view acquisition can provide isotropic detection of rapid biological processes and exceptionally small, fine samples and target structures.

The QuVi SPIM can be configured to provide dual view acquisition simultaneously or sequentially, while the MuVi SPIM achieves orthogonal dual view by one single sample rotation of 90°. Both microscopes thus achieve isotropic resolution and can do so even more easily when combined with our LuxControl multiview fusion and deconvolution algorithms.



About Luxendo Light-Sheet Microscope Technology

Can I image cleared and live samples using one Luxendo microscope?

Yes, the MuVi SPIM and QuVi SPIM can be configured to support cleared, fixed, and live sample imaging.

Can the Photomanipulation Module be added to all Luxendo SPIM microscopes?

The photomanipulation module can be added to every Luxendo SPIM designed for live imaging (MuVi SPIM, QuVi SPIM, TruLive3D Imager).

Luxendo SPIM systems are designed to provide high modularity and can be easily adapted to suit the requirements of your experiment. Contact us to discuss your specific needs with a light-sheet microscopy expert and to receive recommendations and guidance tailored to your experiment.

How does Luxendo light-sheet technology mitigate striping artifacts?

Because it uses a side-illumination optical arrangement, the light-sheet microscopy technique is prone to striping artifacts ("shadows") when there are obstacles within a sample. With Luxendo SPIM instruments, users can achieve crisper artifact-free images faster than ever.

Even in the presence of inhomogeneities and absorbing obstacles, MuVi SPIM, TruLive3D Imager, and LCS SPIM prevent stripe formation during image acquisition. To do so, they illuminate the sample from different angles using a pivot scanner with a beam rotation much faster than the camera exposure time. This enables these instruments to generate a homogenous, shadow-free illumination profile without compromising acquisition speed.
 

How long can a Luxendo SPIM with environmental control image live samples?

Luxendo light-sheet microscopes have been used for multi-day continuous acquisitions on live samples lasting up to 7 days.

During these experiments, high humidity and regulated CO2 / O2 / temperature levels in the acquisition chamber ensure proper and stable growth conditions.

 
How are such vast data sets managed?

Luxendo light-sheet microscopes are compatible with Lux DATA, our comprehensive data processing and storage solution. This data storage and processing unit provides fast data transfer and large-capacity storage as well as remote access and high-performance processing of multidimensional image data, leveraging multi-core and GPU-based computation.

Moreover, Lux DATA is designed to manage not only the vast amount of data generated in light-sheet fluorescence microscopy, but also data from super-resolution or two-photon imaging as well.

LEARN MORE:

What image processing tools are available with Luxendo SPIM microscopes?

Luxendo’s LuxControl software provides a 3D-data viewer, tile stitching, multi-view fusion, and deconvolution as image post-processing tools to deliver whole 3D volume datasets that are ready to use for your image analysis pipeline. 
 

Do I need a specialized room for my Luxendo SPIM microscope?

No, all Luxendo SPIM microscopes are designed to be compact, robust, and vibration-free, and have a fully enclosed optical beam path. They can thus be positioned on any sturdy table (e.g., close to the lab bench) and operated in a daylight room. Additionally, all Luxendo instruments are certified as Laser Safety Class 1 systems, the same safety class as a laser pointer, to allow their unrestricted use.
 

See this question answered in more detail during the audience Q&A session at the end of the light-sheet segment of our on-demand webinar "Bruker Microscopes Fill the Gap Between Traditional Fluorescence and Electron Microscopy" to see this question discussed in greater detail.


 

Choosing A Light-Sheet Microscope

What are some general considerations?

An instrument's utility for any application depends on such factors as sample type and size, resolution requirements, FOV, mounting needs, and imaging speed. The following segments provide some guidelines for choosing the right Luxendo light-sheet microscope for some common applications.

Contact us to discuss your specific measurement requirements with a light-sheet microscopy expert. We will be happy to provide instrument and configuration recommendations tailored to your experiment.

What is the best light-sheet microscope for live samples?

The TruLive 3D and InVi SPIM Lattice Pro support live sample imaging and can be equipped with an environmental control unit for long-term sample cultivation – even up to days of continuous acquisition. Additionally, a photomanipulation module can be added to these instruments for advanced live sample experiments.

The TrueLive 3D Imager is the ideal microscope for multi-positioning experiments (up to 100s of samples and up to six different experimental conditions in a single acquisition period) and imaging delicate live specimens such as small embryos (e.g. mouse), 3D spheroids, organoids, oocytes, and more.

The InVi Lattice Pro includes an advanced illumination module, enabling it to provide high flexibility for sample illumination using static, Gaussian, Bessel beam, or optical lattice illumination. It is thus the ideal microscope when high axial resolution and/or a large depth of field is required at high speed.

What is the best light-sheet microscope for cleared samples?

The LCS SPIM is a dedicated microscope for cleared sample imaging and optimized for very large specimens up to entire adult cleared mice which facilitates visualization of e.g., complete neuronal or vasculature networks at cellular resolution.

What is the best light-sheet microscope for both live and cleared samples?

Both the MuVi SPIM and QuVi SPIM are designed to support:

  • Pure live sample imaging with environmental control and photomanipulation;
  • Pure cleared sample imaging; and
  • Triple-purpose live, fixed, and cleared sample imaging.

The functional flexibility of these instruments is made possible by Luxendo’s highly adaptable modular concept. Users can quickly and easily exchange the central unit, consisting of the illumination objective, detection objective, and the sample mounting chamber, to reconfigure the instrument according to the measurement requirements.

More specifically:

The QuVi SPIM is a highly versatile upright, dual view and dual color light-sheet microscope for 3D imaging of living, fixed, and cleared samples. It is optimized for long-term, high-throughput imaging at subcellular isotropic resolution.

The horizontal 4-axis arrangement of the MuVi SPIM allows for two illumination and two detection sides to be equipped, enabling high-speed volumetric acquisition. It is consequently optimal for studying large biological live samples (e.g., drosophila or zebrafish embryos) and fast biological processes. With the MuVi SPIM CS configuration, large, cleared samples up to entire mouse brains can be resolved at subcellular resolution.

What is the best light-sheet microscope for large samples/whole-body specimens?

The LCS SPIM is a dedicated microscope for cleared sample imaging and optimized for very large specimens up to entire adult cleared mice. This enables visualization of structures like complete neuronal or vasculature networks at cellular resolution.

The MuVi SPIM is also optimal for studying large live samples (e.g., drosophila or zebrafish embryos) and, with the MuVi SPIM CS configuration, large cleared samples up to entire mouse brains can be resolved at subcellular resolution.



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