Holographic tomography vs Immunofluorescence: Zebrafish skeletal muscle

Zebrafish (Danio rerio) is a lively little schooling fish that only reaches about 6 cm in the aquarium. It’s easy to recognize with an attractively striped black and white “zebra” patterned body. The use of zebrafish (Danio rerio) as a model organism began in the 1960s: Zebrafish reproduce in large quantities, grow rapidly, and are transparent early in development; for these reasons, zebrafish have been used extensively to model vertebrate development and disease. From a genetic point of view, Zebrafish are very similar to humans (they share 70% of genes with us) and they have the same major organs and tissues as humans. Their muscles, blood, kidneys and eyes resemble that of the human systems. Interestingly, Zebrafish have an impressive ability to repair its muscles. For example, if part of their heart is removed they can grow it back in a matter of weeks. Scientists are working to find out the specific factors involved in this process to evaluate if this will help to develop new ways of repairing the heart in humans with heart failure or who have suffered heart attacks.

Here we want to show how Nanolive’s 3D Cell Explorer was able to explore zebrafish muscles in 3D and without any chemical staining in just a couple of seconds!

The micrograph on the left panel in the figure below shows an immunofluorescent staining on a cryosection (10 micrometer) of adult zebrafish skeletal muscle. The cytoplasm of the myofibers is rich in contractile proteins such as myosins. Slow-twitch myofibers are characterized by the presence of a particular isoform of myosin heavy chain, the myosin heavy chain 1 (green). Mitochondria (red) have been labeled with an antibody directed against Tom20, a protein specifically expressed in the mitochondrial outer membrane. Mitochondria are present in the sub-sarcolemmal compartment (red filaments) as well as between the myofibrils (little spots inside green regions). Nuclei are stained with Hoechst, showed here in blue. The images acquired with the 3D Cell Explorer are displayed on the central and right panels: the grey scale image is a map of the tissue based on the Refractive Index (RI) of its components. The main structures were identified and reconstructed in 3D through our fast, easy, inexpensive and non-invasive digital staining procedure. Judge the results by yourself 😉

comparison2

Figure. a. Confocal microscope immunofluorescent staining on a cryosection of adult zebrafish skeletal muscle. b. Tissue’s Refractive Index (RI) map in 2D. c. 3D Visualization as obtained with our software STEVE. The D-Stain panel shows how the different compartment are digitally stained based on their specific RI.

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