Tomocube HT microscope allows unlabeled quantification of cellular lipid content

July 12, 2019

In a major breakthrough for research fields as varied as diabetes, atherosclerosis and biofuels, the mbad, concentration, volume and surface area of ​​individual lipid droplets were measured for the first time in living cells. unstained using a Tomocube Holotomography (HT) microscope. Eliminating any preparation or staining step and the 110 nm lateral resolution of the Tomocube HT microscope now allow researchers to easily track changes in lipid distribution over time at high resolution, allowing studies to be done. metabolic content of cellular lipids.

Holotomography of a human hepatocyte, Huh-7. 3D tomogram images (left), fluorescence image (right). The high RI droplets correlate with lipid droplets stained with red Nile. 95.5% of the organelles with RI values ​​greater than n = 1.375 overlap in lipid droplets stained with Nile red. Reproduced by Kim et al. Scientific reports 6: 36815.

Immediate quantitative images in phase of the intact morphology of living cells are a key advantage of HT microscopy compared to other imaging technologies. The holotomography detects the absolute refractive index (IR) of each voxel by measuring the phase delay. Since the IR of the lipid droplets is significantly higher than the cytoplasm or other cellular components, the contrast is easily sufficient to obtain a high resolution image. If you are using the Tomocube HT-2H microscope, which supports both holotomography and 3D fluorescence imaging, co-localization experiments are possible by labeling only the target protein or specific type of DNA. 39 fatty acid rather than dyeing both the lipid droplet and the target molecule. .

Compare this with light-field or phase-contrast microscopy, which conveniently visualizes cell morphology but provides only qualitative information. Confocal fluorescence microscopy captures high resolution 3D structural images, but involves invasive labeling procedures that inevitably induce phototoxicity or photobleaching. "

Aubrey Lambert, Marketing Director, Tomocube

Lipids play a crucial role in the modulation of many cellular functions and a breakdown in its regulation is badociated with diabetes, atherosclerosis and other metabolic diseases. The genetic engineering of lipid-rich bacterial and photosynthetic algae to increase the yield of fatty acids is also an important area of ​​research in the development of biofuels. In both cases, the amount of lipids must be quantified to study the metabolic dynamics in vitro.

Until now, to visualize the location of the lipids, the cells had to be prepared using lipid-specific staining probes, such as Nile red or red-oil. O, interfering with native cellular processes. Moreover, chromatography has been widely used to quantify lipid content, although it only measures the average of hundreds of algae after tedious experimental procedures.

According to Lambert, "the holotomography is very well suited to the detection of high-RI molecules, characteristic of lipid droplets. However, it is also applicable to the study of other high-RI molecules, such as polymers, nanoparticles or liposomes, and researchers may apply approaches identical to those of lipid imaging using HT microscopy.

"The rapid quantification of cellular lipid content is a unique feature of HT imaging and presents tremendous potential for adoption in cell biology as well as in the areas of bioengineering or the development of cell biology. biofuel industry. "