BrdU (B5002, Sigma-Aldrich) was dissolved in PBS to a stock concentration of 10 mg/ml. A total of 166.7 μg BrdU per gram of body weight was intraperitoneally injected for five consecutive days at different desired stages. At 6 dpi, injected fish were sacrificed, and the brains were dissected for further cryo-section. For each fish, two adjacent brain section slices were collected separately on two slides and were subjected to anti-BrdU and TUNEL staining, respectively.
BrdU staining was performed as previously reported (60 (link)). TUNEL assay was performed using Click-iT Plus TUNEL assay (Invitrogen, C10619). Subsequent immunostaining was performed as previously described (61 (link)). Primary antibodies used in this study are anti-GFP antibody (ab6658, Abcam), anti-DsRed antibody (632496, Clontech), anti-BrdU antibody (11170376001, Roche), anti–caspase 3 antibody (559565, BD Biosciences), and anti-Lcp1 antibody (30 (link)). The secondary antibodies used in this study are Alexa 488 anti-goat antibody (A11055, Invitrogen), Alexa 555 anti-rabbit antibody (A31572, Invitrogen), and Alexa 647 anti-mouse antibody (A31571, Invitrogen).
BrdU staining was performed as previously reported (60 (link)). TUNEL assay was performed using Click-iT Plus TUNEL assay (Invitrogen, C10619). Subsequent immunostaining was performed as previously described (61 (link)). Primary antibodies used in this study are anti-GFP antibody (ab6658, Abcam), anti-DsRed antibody (632496, Clontech), anti-BrdU antibody (11170376001, Roche), anti–caspase 3 antibody (559565, BD Biosciences), and anti-Lcp1 antibody (30 (link)). The secondary antibodies used in this study are Alexa 488 anti-goat antibody (A11055, Invitrogen), Alexa 555 anti-rabbit antibody (A31572, Invitrogen), and Alexa 647 anti-mouse antibody (A31571, Invitrogen).
