Journal: Frontiers in Cell and Developmental Biology

Article Title: Spermatogonial fate in mice with increased activin A bioactivity and testicular somatic cell tumours

doi: 10.3389/fcell.2023.1237273

Figure Lengend Snippet: Antibody details.

Article Snippet: Fixed and permeabilized cells were washed in PBS with 2% FCS then stained with the following antibodies diluted in PBS with 2% FCS: eFluor 450-conjugated anti-KI67 clone SolA15 (Thermo Fisher Scientific, 1:500, 48–5698–82, RRID: AB_11149124 ) and phycoerythrin (PE)-conjugated anti-KIT clone 2B8 (Thermo Fisher Scientific, 1:250, 12–1171–81, RRID: AB_465812 ), rabbit anti-mouse EOMES clone 1219A (R&D Systems, 1:1,000) and Alexa 647-conjugated anti-PLZF clone 9E12 ( ).

Techniques:

Journal: Frontiers in Cell and Developmental Biology

Article Title: Spermatogonial fate in mice with increased activin A bioactivity and testicular somatic cell tumours

doi: 10.3389/fcell.2023.1237273

Figure Lengend Snippet: Flow cytometry analysis of testis tubule cells from adult Inha WT and KO ‘normal’ testes. (A) Gating strategy used to identify (a) undifferentiated SPG (PLZF+/KIT–), (b) early differentiating SPG (PLZF+/KIT+), and (c) late differentiating SPG (PLZF low /KIT+) cells. Percentages of cells within gates are indicated. (B) Graph showing subpopulations (a–c) of cells as a percentage of tubule cells (interstitial depleted). (C) Gating strategy for EOMES+ cells within the PLZF+/KIT- population. (D) Graph showing EOMES+ fraction as a percentage of (a) undifferentiated cells. (E) Representative flow cytometry and (F) graphs showing proportion of proliferating (KI67+) cells in populations from (A) and (C) . All graphs show mean percentage of cells in each population ± SD ( n = 4 animals/genotype). Significance determined using a two-tailed unpaired t-test, * p < 0.05, ** p < 0.01.

Article Snippet: Fixed and permeabilized cells were washed in PBS with 2% FCS then stained with the following antibodies diluted in PBS with 2% FCS: eFluor 450-conjugated anti-KI67 clone SolA15 (Thermo Fisher Scientific, 1:500, 48–5698–82, RRID: AB_11149124 ) and phycoerythrin (PE)-conjugated anti-KIT clone 2B8 (Thermo Fisher Scientific, 1:250, 12–1171–81, RRID: AB_465812 ), rabbit anti-mouse EOMES clone 1219A (R&D Systems, 1:1,000) and Alexa 647-conjugated anti-PLZF clone 9E12 ( ).

Techniques: Flow Cytometry, Two Tailed Test

Journal: Frontiers in Cell and Developmental Biology

Article Title: Spermatogonial fate in mice with increased activin A bioactivity and testicular somatic cell tumours

doi: 10.3389/fcell.2023.1237273

Figure Lengend Snippet: Tubules adjacent to stromal tumour regions are associated with clusters of cells with SSC characteristics. (A) Representative image of tumour region (white dashed line) in adult Inha KO testes. Numbered boxes show location of tumour associated tubules (designated TAT) (in B) relative to tumorigenic region. (B) Representative images showing clusters of GFRA1+/EOMES+ cells. White arrow heads indicate GFRA1+/EOMES+/GILZ+ spermatogonia. (C) High density GFRA1+ cells are LIN28– or low. White arrow heads indicate GFRA1+/LIN28-/GILZ+ spermatogonia. (D) Representative negative control images. Scale bars: main image = 200 μm, panels (B–C) = 20 μm, negative control panels = 50 µm. (E) Graphs show average (AVG) tubule perimeter (µm), AVG number of GFRA1+ cells and SOX3+ cells/mm tubule perimeter, and ratio of SOX3+/GFRA1+ cells in Inha WT (grey bars) and KO ‘normal’ (N, purple bars) and TAT tubules (orange bars). All graphs show mean ± SD (n = 3/genotype). Significance determined using a two-tailed unpaired t-test, * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: Fixed and permeabilized cells were washed in PBS with 2% FCS then stained with the following antibodies diluted in PBS with 2% FCS: eFluor 450-conjugated anti-KI67 clone SolA15 (Thermo Fisher Scientific, 1:500, 48–5698–82, RRID: AB_11149124 ) and phycoerythrin (PE)-conjugated anti-KIT clone 2B8 (Thermo Fisher Scientific, 1:250, 12–1171–81, RRID: AB_465812 ), rabbit anti-mouse EOMES clone 1219A (R&D Systems, 1:1,000) and Alexa 647-conjugated anti-PLZF clone 9E12 ( ).

Techniques: Negative Control, Two Tailed Test