Holotomography
Label-free quantitative imaging : A completely new way of investigating cells and tissues
Tomocube’s Holotomography (HT) technology provides label-free 4D quantitative imaging solutions for imaging and analyzing cells and tissues. Without using any preparation including fixation, transfection, or staining, details of dynamics and mechanisms of live cells, subcellular organelles, and tissue structures can be seen. HT not only enables observation of nanoscale, real-time results based on quantitative phase imaging (QPI) but also provides quantitative information of cells and organelles.
The Principle
Conventional methods used to image unstained biological samples, such as Phase Contrast or Differential Interference Contrast microscopy, only image in two dimensions when biological samples are in reality, three dimensional. Tomocube’s Holotomography (HT) solves this problem.
Just as a CT scan uses X-ray absorptivity as the imaging contrast to see inside a patient’s organs without invasive procedures, HT uses the refractive index (RI), an intrinsic optical parameter describing the speed of light passing a specific material, to visualize living cells and tissues. As light traverses through a sample, the various constituents scatter light differently based on their refractive index. By rotating an imaging beam 360° around the sample, we can capture a sequence of holograms from different angles. The resulting multiple 2D hologram images of the sample obtained from various illumination angles can be reconstructed into a 3D RI tomogram.
The Principle
How is this achieved?
The sample is located on a stage between an objective lens and a condenser lens. A light source is split to follow two paths, the sample beam and the reference beam (the principle behind a Mach-Zehnder interferometer). The sample and the reference arms when combined generate a 2D hologram, which is recorded by a digital image sensor (CMOS). This reflects a QPI measurement principle of the phase shift relative to the reference beam.
The imaging beam illuminates the sample with an incident angle of 63°. The beam is rotated through 360° with respect to the optical axis. From the 48 overlapping captured holograms RI tomogram of the sample is then reconstructed.
Patented Beam Rotation Technology
The DMD consists of several hundred thousand micromirrors arranged in a rectangular array. Each individual mirror can be rapidly tilted electronically to create a mirror pattern which can rotate the beam through 360° around the optical axis at a desired angle.
More than just a 3D image: Quantitative Bioimaging Meets Kinetics.
Using our TomoStudio™ software, we can effectively pseudo-color bands of RI to highlight structures within the captured volume and visualize 3D, color-coded structures that were previously undetectable without staining.