How does the Tilt Light Sheet Module work?

 

The Tilt Light Sheet is a simple technology that can turn almost any inverted microscope into a Light Sheet microscope. It is easy to use, affordable and combines the benefits of Light Sheet microscopy with high-resolution imaging. Keep reading to learn how the Tilt creates a Light Sheet.

Step 1

A collimated laser beam is first passed through a photomask with 4 slits to create 4 separate beams of light. Why was this particular photomask chosen? Check out this paper by Golub et al., (2015) to learn why.

Step 2

Focus the 4 light beams through a cylindrical lens.

Step 3

The 4 light beams then converge and interfere to create a single Light Sheet that is ~4 microns thick and ~300 microns long.

Why create a Light Sheet this way? What are the benefits?

Traditional Gaussian light sheets (created by focusing a laser beam through a cylindrical lens) can have a narrower waist (depending on the lens used to create the Light Sheet) but this waist diverges much more quickly:

 

The waist of the Tilt Light Sheet is ~4 microns thick over ~300 microns whereas a Gaussian Light Sheet with the same waist thickness diverges more quickly (~4 microns thickness over less than 100 microns). This means that the Tilt Light Sheet is thinner over a longer distance. Some reasons why this is advantageous:

1) It can be easier to focus, i.e. image your sample with the thinnest portion of the Light Sheet.

2) Better optical sectioning over a longer distance (higher contrast), especially advantageous for specimens thicker than ~15 microns.

The ‘tilt’ part of the Tilt

You probably noticed that the whole Tilt Light Sheet module is slightly tilted and wondered why. By introducing a slight tilt (~3 degrees) the bottom-most light beam runs parallel to the coverslip and allows for a Light Sheet to be created in the focal depth of any objective. Meaning that the Tilt Light Sheet module can be used with any objective. This feature is unique to the Tilt.

Compatible with high magnification, high numerical objectives

Because of the ability to create a Light Sheet very close to the surface of a coverslip, the Tilt is compatible with any objective. Including high magnification, high numerical aperture objectives which allow scientists to combine Light Sheet with subcellular resolution. If you need to image with 60x or 100x high numerical aperture objectives for long periods of time, the Tilt might be perfect for you!

The technology behind the Tilt was developed by Paul Maddox’s lab at the University of North Carolina at Chapel Hill. Check out the publication for more details.

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