In Vitro Culturing Technique for Studying Cellular Dynamics in Zebrafish Scales

Our research explores the effects of environmental disturbances of bone tissue. We are specifically interested in understanding the effects of simulated microgravity and vibration on adult and developing bone. We use zebrafish as a model organism of choice because it offers many advantages for biological research.

Our protocol offers advantages over previous protocols because it limits the need for a carbon dioxide incubator, and it uses standard laboratory equipment. Our lab is trying to unravel the communication dynamics between different types of bone cells when they are subjected to an environmental disturbance such as gravity or vibration. In the future, we would like to be able to mitigate the negative effects of space travel on bone tissue.

To begin, dissolve 10 grams of DMEM powder and 5.95 grams of HEPES in one liter of distilled water and adjust the pH of the media to 6.91. Aliquot the solution into smaller containers and store them at four degrees Celsius. Then mix 88%DMEM medium with HEPES, 10%fetal bovine serum, and 2%penicillin-streptomycin.

To prepare the zebrafish, set up a container or a tank with approximately two liters of water for four to five adult zebrafish. Using a fish net, transfer four to five adult zebrafish into the prepared tank. Prepare separate containers with water for anesthetizing and recovering the zebrafish.

After anesthetizing an adult zebrafish, use a plastic spoon to transfer the fish to a petri dish lined with a wet paper towel and place the dish under a dissecting stereo microscope. Next, using fine forceps, gently tug in the anterior to the posterior direction to remove the scales. Place each individual scale into a separate 0.2ml tube containing the scale culture medium working solution, and put the tubes inside an incubator set to 28 degrees Celsius.

After scale removal, carefully return the zebrafish to the recovery tank and monitor it. Once the zebrafish has resumed movement, return it to the regular rearing tank.