PLZF+ cells recover more slowly following busulfan treatment. (A) Eight-week-old wild-type and Tg(Gdnf) mice were injected with 10 mg/kg body weight of busulfan and analyzed 3, 6, 9, 12 and 15 days post-injection. (B) Immunostaining of whole-mount tubules stained for PLZF at 3, 6, 9 and 15 days post-injection. PLZF+ Aal chains were sensitive to busulfan, and by day 9, only As cells were present in wild-type testes. (C) Quantification of PLZF+ cells post-busulfan injection in wild-type and Tg(Gdnf) mice. Values are normalized to Sertoli cells and presented as a semi-log graph. The P values of the slopes were determined using log transformation of the total number of cells to stabilize the variance (performed using JMP 7.0 program). 3-9 days slope, P=0.06; 9 days-15 days slope P=0.0017 (**P<0.005). Scale bars: 100 µm.