(A).The testis of wild type mice showed normal c-Kit protein expression (Aa,Ab), whereas the expression was absent in the testis of busulfan treated mice (Ad, Ae).a &d FITC; b& e; FITC & PI merge; c, f : respective negative controls wherein primary antibody was replaced with pre-immune serum (---Scale bars 50 μm).
(B). RT-PCR analysis of c-Kit mRNA expression in the testis of busulfan treated (Lane B1) and wild type mice (Lane B3) .c-Kit transcript was observed in the testis of wild type mice . Lane B2: No RT enzyme control. Cyclophiline-A was used as loading control (M = 100 bp DNA ladder).
(C).Western blot analysis of c-Kit in the testis of busulfan treated (Lane: C1) and wild type mice (Lane: C2). C-Kit protein (150 kDa) expression was seen in the testis of wild type mice. Negative controls (Lanes C3 and C4) where primary antibody was replaced with mouse serum. β-actin was used as loading control.
(D). RT-PCR analysis of Sertoli cell marker genes (GATA-1 & PEM-1) in the testis of busulfan treated (Lane: D1) and wild type mice (Lane: D2). Transcripts for GATA-1 & PEM-1 genes were seen in both these mice. Lane D3: No RT enzyme control. GAPDH was used as loading control.
