New technique for high speed, three-dimensional imaging
A limited number of approaches currently exist for 3D imaging and microscopy in biological research, medicine and industrial applications. Two-photon microscopy is both expensive and slow, owing to the need to scan a laser beam to every 3D location within a volume. Light-sheet technologies achieve optical sectioning through selective illumination, but are typically configured with two tightly aligned orthogonal objectives and a requirement to physically move a mounted sample. This limits both volumetric imaging speed and sample diversity (by shape, mounting ability, motion and scattering properties). Capturing volumetric imaging data from unmounted, living, and/or freely moving samples at faster-than-video rates is a major goal in neuroscience and biomedical research, and could find applications in medicine and a range of industrial processes. If available in a low-cost, simple-to-use benchtop platform, such a system could find a home in almost any biomedical research laboratory.
Single-objective, scanned light-sheet imaging permits high speeds and sample diversity.
Swept Confocally Aligned Planar Excitation microscopy (SCAPE) allows for the rapid 3D imaging of fluorescent (or other) samples without the need for sample mounting and translation, and can image in both intact scattering and non-scattering samples to within scattering limits. SOLiS utilizes a unique optical design that permits a stationary camera to acquire de-scanned images of a moving light sheet plane as it is scanned through the sample at an oblique angle. Samples require no preparation and can be positioned as in a normal epifluorescence microscope. Limited only by camera speed and signal to noise, SOLiS can acquire high-density volumetric, polychromatic images at between 20 and 400 volumes per second with commercially available hardware. Image formation does not require a reconstruction algorithm (although spatial corrections for spatial scaling and deconvolution can be applied if desired). The system's region of interest is scalable, permitting imaging ranging from diffraction-limited microscopy to large-scale imaging of non-scattering structures and 3D profilometry. The technique does not require (but could incorporate) non-linear excitation, and is thus compact and inexpensive. Preliminary SOLiS 4D image sequences of crawling transgenic drosophila larvae and in-vivo rodent brain (blood flow and GCaMP6 calcium activity in superficial dendrites) have been acquired at over 20 volumes per second using an inexpensive prototype dual-color SOLiS system.
Lead Inventor:
Applications:
- 3D live-cell imaging: capturing both cell motion and signaling dynamics in single cells, ensembles, intact and engineered tissues.
- Imaging small living organisms: drosophila (fruit fly), zebra-fish, c-elegans etc. for neuroscience, developmental, cardiovascular, genetics, disease and drug discovery research.
- In-vivo mammalian brain imaging: cranial-window based imaging of hemodynamics, intrinsic contrast, voltage and calcium sensitive dyes and fluorescent proteins.
- Human clinical imaging: intrasurgical, retinal, dermal and endoscopic imaging of superficial tissues.
- Exploiting the power of GCaMP and new transgenic proteins with sensitivity to voltage and other molecular and signaling targets.
- Fluid dynamics: detecting subsurface flows and convection and turbulent flow patterns
- Industrial materials: analyzing transparent and semi-transparent plastics, metal alloys and ceramics for defects.
- High-speed profilometry of reflective, scattering samples.
- Extensions to image Raman scattering, second harmonic generation, two and three-photon effects.
- Extension to superresolution via STED or structured light implementations.
Advantages:
- 3D volumetric images are generated at higher speeds than existing approaches, which are much more complex.
- 3D information is obtained without the use of multiple objective lenses, cameras or the need for sample mounting, translation or rotation.
- Unlike existing orthogonal light-sheet methods, this technology can image both cleared and intact scattering samples such as the rodent brain.
- The system design has almost no moving parts, uses inexpensive components, and could be portable, compact and even minaturized.
- The technology does not rely on computationally expensive model-based image reconstruction.
- Opportunity for development of proprietary 4D dynamic image analysis and visualization packages.
Patent Information:
Publication: Nature article