Fig 1: Age dependent loss of undifferentiated spermatogonia in absence of GCNA. (A) Immunofluorescence staining of GCNA (Tra98) and PLZF on adult mice testis sections. PLZF was displayed in red, GCNA in green and DNA (DAPI stained) in blue. Cells positive for both PLZF and GCNA are highlighted in yellow and PLZF only cells are highlighted in white. GCNA is expressed in mice USGs. (B) Immunofluorescence staining of GCNA and PLZF on human testis sections. GCNA was displayed in red, PLZF in green and DNA (DAPI stained) in blue. GCNA is expressed in human USGs. (C) Frequency of PLZF positive cells also positive for GCNA in human and mouse. A minimum of 50 PLZF positive cells are scored per experiment. Data represent the mean and S.D. Data represent three independent experiments. (D) Immunofluorescence staining of PLZF on testis sections of 3, 6 and 12 months old wild-type and Gcna−/Y mice. PLZF is displayed in green and the DNA staining DAPI in blue. Yellow arrows highlight USGs, which are PLZF positive. Bottom panels are magnifications of regions highlighted by the yellow boxes. (E) Number of PLZF positive cells per seminiferous tube in wild-type and Gcna−/Y testes, between 1.5 months and 12 months. A minimum of 50 seminiferous tubes are scored per mouse. Data represent the mean and S.D. (n = 4 mice for each genotype and age). P values were calculated by using an unpaired t-test. (F) Frequency of seminiferous tubes displaying no PLZF positive cells in wild-type and Gcna−/Y testes, between 1.5 months and 12 months. A minimum of 50 seminiferous tubes are scored per mouse. Data represent the mean and S.D. (n = 4 mice for each genotype and age). P values were calculated by using an unpaired t-test.
Fig 2: Accumulation of undifferentiated spermatogonia in S-phase in absence of GCNA. (A) Immunofluorescence staining of wild-type and Gcna−/Y testis sections from 6 months old mice. KI67 was displayed in red, PLZF in green and DNA (DAPI stained) in blue. Cells positive for both PLZF and KI67 are highlighted in yellow and PLZF only cells are highlighted in white. (B) Frequency of PLZF positive cells negative for KI67 (quiescent USGs) at 1.5, 3, 6 and 12 months old. A minimum of 50 PLZF positive cells are scored per mouse. Data represent the mean and S.D. (n = 3 mice for each genotype and age). P values were calculated by using an unpaired t-test. (C) Immunofluorescence staining of wild-type and Gcna−/Y testis sections from >6 months old mice. Phosphorylation of H3 (pH3, Ser10) was displayed in red, PLZF in green and DNA (DAPI stained) in blue. Cells positive for both PLZF and pH3 are highlighted in yellow and PLZF only cells are highlighted in white. (D) Frequency of PLZF positive cells also positive for pH3 at 1.5 and >6 months old. A minimum of 50 PLZF positive cells are scored per mouse. Data represent the mean and S.D. (n = 3 mice for each genotype and age). P values were calculated by using an unpaired t-test. (E) Immunofluorescence staining of wild-type and Gcna−/Y testis sections from > 6 months old mice. Cyclin A2 (CCNA2) was displayed in red, PLZF in green and DNA (DAPI stained) in blue. Cells positive for both PLZF and CCNA2 are highlighted in yellow and PLZF only cells are highlighted in white. (F) Frequency of PLZF positive cells also positive for CCNA2 at 1.5 and >6 months old. A minimum of 50 PLZF positive cells are scored per mouse. Data represent the mean and S.D. (n = 3 mice for each genotype and age). P values were calculated by using an unpaired t-test. (G) Immunofluorescence staining of wild-type and Gcna−/Y testis sections from >6 months old mice. EdU was given to mice through intraperitoneal injection and mice were culled for histological analyses 4h after injection. EdU was displayed in red, PLZF in green and DNA (DAPI stained) in blue. Cells positive for both PLZF and EdU are highlighted in yellow and PLZF only cells are highlighted in white. (H) Frequency of PLZF positive cells also positive for EdU at 1.5 and >6 months old. A minimum of 50 PLZF positive cells are scored per mouse. Data represent the mean and S.D. (n = 3 mice for each genotype and age). P values were calculated by using an unpaired t-test.
Supplier Page from MilliporeSigma for Anti-GCNA antibody produced in rabbit