Intraperitoneal Injection: A Method of Solution Delivery into the Abdominal Cavity of an Adult Zebrafish
筆記録
– Fast an adult fish for 24 hours before injection に empty the gastrointestinal tract and create additional space で the abdomen. On the day of the procedure, anesthetize the fish with tricaine and place it into a slot で a wet sponge with the ventral side facing up.
Next, pipette the desired amount of injection solution onto a piece of laboratory film and pull it into an insulin needle. 役職 the needle between the pelvic fins at a 45 degree angle from the anterior-posterior body axis and gently push it で at the midline. Advance the needle approximately 1 に 2 millimeters into the abdominal cavity, the space between the abdominal wall and internal organs, and slowly inject the solution of interest. Wait for five seconds before pulling out the needle に avoid any spillage. Now, carefully remove the needle.
After injection, quickly transfer the fish into a recovery tank containing fresh tank water. If the fish doesn't begin に swim immediately, swirl the water near the gills に help it recover from anesthesia. In the example protocol, we will inject a mycobacterium marinum solution into RAG1 mutant fish に study infection.
– First, pipe out of 5 microliter droplet of the diluted bacterial solution onto a piece of paraffin film. Then, pull the droplet into a 30 gauge insulin needle. Use a five に eight-month-old fish for this experiment, with one being a wild-type fish and the other being a RAG mutant fish.
役職 these fish ventral side up で the slits of a piece of moist foamed plastic. Inject the insulin needle between the pelvic fins at a 45 degree angle. Keep the needle opening upwards に ensure that the entire opening is inside the abdominal cavity. Then, slowly inject the bacterial solution. After this, carefully remove the needle and immediately transfer the fish に a recovery tank filled with fresh tank water.
Take samples from the bacterial aliquot で use every 15 minutes on 7H10 plates. Incubate these samples at 29 degrees Celsius for five days に verify the infection dose. Check the well-being of the fish regularly, making sure に euthanize any fish with infection symptoms by incubating them で water with more than 0.02% of 3-aminobenzoic acid ethyl ester.
