Computational Imaging: Digital Microscopy

May 9, 2016 Christina Mayer

In the field of photography, the digital revolution has replaced analog recording technology using photographic plates with digital sensor chips. And it's not surpring that this same trend has changed microscopy. Before digital sensors were introduced, nearly all microscopes were equipped with an ocular, i.e. the eyepiece lens which enables the person using the microscope to directly observe the probe being studied.

In modern digital microscopes, however, oculars are a rarity. Digital sensors and monitor displays have become so advanced that digital technology has almost entirely replaced direct observation. The data observed is saved and further processed digitally. In other words: the many benefits of digital photography – just think of all the things the camera on your smartphone can do – are also useful for microscopic data:

  • HDR images: by combining images with different illumination levels and/or illumination times, images can be created that have much better contrast than a single image. This has proven advantageous when e.g. you're looking at specimens through a microscope which have very heterogeneous absorption or reflection. Higher contrast enables scientists to identify finer details in the shadow areas, even when very bright light sources, e.g. highlights, appear on the screen at the same time.
  • Stitching (panorama): most microscopes have a very small field of view because of the high magnification. That's why a motorized table is used for very large specimens. The individual images of an object are then combined to form a single image with extremely high resolution.
  • Image processing: in addition to the methods mentioned above, there are still more ways to process an image. Typical examples include enhancing contrast, noise reduction and deconvolution to sharpen the image. Many of these methods are now a part of every digital microscope.

Thus there's a significant overlap between the functionality of a basic digital microscope and a normal smartphone. You can even use your smartphone to perform a few basic tasks for which you'd normally use a microscope. All you need to do is to outfit your smartphone with some enlarging optics. These simple microscopes have already produced a few really lovely images (check out here).

Of course digital methods are also of interest in high-end microscopy because, by using some special tricks, it is possible to circumvent the classic microscopic limitations. That's why, for example, only one location is illuminated in a confocal microscope and thus the emitted fluorescence signal is detected. Rasterization is then used to capture the entire image. By selectively detecting the light emitted from the excited point, an optical cut can be made, making 3D imaging possible. In localization microscopy, which uses high-resolution, camera-based detection, the analysis of individual light points under certain conditions makes it possible to determine the location of the fluorescing points beyond the resolution limit. This can only be used for samples where fluorescing molecules are spread thinly or these samples emit thinly scattered fluorescent light via stochastic switching.

Lars Omlor and Ivo Ihrke

References:

Atcheson, B., Ihrke, I., Heidrich, W., Tevs, A., Bradley, D., Magnor, M., & Seidel, H. P. (2008, December). Time-resolved 3d capture of non-stationary gas flows. In ACM transactions on graphics (TOG) (Vol. 27, No. 5, p. 132). ACM.

M. L. Faulkner and J. M. Rodenburg. Movable aperture lensless transmission microscopy: A novel phase retrieval algorithm. Phys. Rev. Lett., 93:023903, 2004.

Levoy, M., & Hanrahan, P. (1996, August). Light field rendering. In Proceedings of the 23rd annual conference on Computer graphics and interactive techniques (pp. 31-42). ACM.

Levoy, M., Ng, R., Adams, A., Footer, M., & Horowitz, M. (2006). Light field microscopy. ACM Transactions on Graphics (TOG), 25(3), 924-934.

Mignard-Debise, L., & Ihrke, I. (2015, October). Light-field Microscopy with a Consumer Light-field Camera. In 3D Vision (3DV), 2015 International Conference on (pp. 335-343). IEEE.

Tian, J. Wang, and L. Waller, "3D differential phase-contrast microscopy with computational illumination using an LED array," Optics Letters, vol. 39, no. 5, pp. 1326--1329, March 2014.

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