Glycerol / PBS at 4 before viewing. To examine the M-phase cells, the entire arrangement immunohistochemistry to detect phospho histone H3. The fish were placed in 400 M neomycin for 1 h, rinsed 4 times in fresh EM, and with the optimal concentration of the inhibitor Bleomycin Blenoxane drug GU 24 h at 28.5. The larvae were then incubated with MS 222, with 4% PFA overnight at 4 and rinsed several times for 20 in PBS T. hair cells labeling was before using rabbit anti-GFP and Alexa 488-conjugated goat anti-rabbit IgG immunostaining staining for phospho histone H3. The larvae were then washed in distilled water for 30 min with 5% normal goat serum in Blockierungsl Solution and incubated overnight with primary anti-rat Ren phosphohistone H3. Min after three washes with PBS-T for 20 samples for 5 h with Alexa 568 were called conjugated goat anti-rat. The larvae were closing Lich rinsed three times in PBS and present in the T 50% glycerol / PBS at 4, which. COLUMNS to the total number of cells in the neuromasts SECT Able to k, AB wild-type zebrafish larvae get Tet and in 4% PFA fixed overnight at 4 After several Rin Ages T Moxifloxacin Topoisomerase inhibitor in PBS, the fish in the pan nuclear dye SYTOX green, which stains the nucleic Acids with high affinity t and incubated to both hair cells and supporting cells. After 5 min, larvae were repeatedly washed in PBS-T and in 50% glycerol / PBS at 4 prior to imaging. The samples were dissolved in 50% glycerol / PBS on a Objekttr hunter deck mounted and observed on a Zeiss LSM5 Pascal confocal microscope under 40 × objective. Double-labeled cells were in seven neuromasts of fish gez just increments and compared with the values of controlled fish The vehicle alone. Images were analyzed using ImageJ, and Adobe Photoshop CS4 Pascal. The figures are presented as the mean total number of cells per fish. We calculated t tests and ANOVA to assess statistical significance.
The amputation of the tail fin. Wild-type AB zebrafish embryos were added to 2 3 dpf, anesthetized with MS 222 dechorionated and placed in a glass-Objekttr hunter depression. Using a scalpel, the tail fin primordia posterior notochord was amputated. The larvae were then photographed to document under the microscope Zeiss Axioplan 2 bright field of a filter differential interference contrast with a 10objective to the point of amputation. The fish was then placed in individual wells of a 48 well plate with the optimal concentration of a drug modulator in 1 ml of EM and recover for 72 h at 28.5. Each fish exerts bet Myricetin Was with MS 222 and the tail fin has been remapped. Caudal regeneration was determined by subtracting the distance between the tip of the tail fin of the notochord immediately after the amputation of the Ma Opinion on evaluated 72 h after the amputation. The tip of the tail as the point at the end of the rib defined directly along the center line of the notochord. Results of rapid tests for modulators of hair cell regeneration using the online system of zebrafish side, we have for new drugs and drug use as small compounds, the duration and Ausma affect the planned regeneration hair cells. Tion to induce the regeneration, we exposed zebrafish larvae at 5 dpf 400M to neomycin for 1 h, the tet 90% of the adult lateral line hair cells t. The regeneration occurs rapidly in wild-type animals, reaching a contr The survey.