Figure 2. Comparison of whole lenses labeled with EdU and lens explants labeled with BrdU. A: EdU labels lens epithelial cells in S-phase in the germinative zone (GZ) of a 1-month-old lens. Lenses were placed on their equators and image fields were captured after moving one cells width anteriorly from the meridional rows (MR) which serve as a geographical marker for the posterior boundary of the lens epithelium. An example is provided in the inset, which is stained with Alexa Fluor488 phalloidin to label filamentous actin. The vertical dashed line is included to roughly depict the end of the transitional zone (TZ) and the beginning of the GZ. B: EdU-positive cells in the GZ of an 8-month old lens. C: An example of labeling of S-phase cells using an antibody to BrdU in an explant from a 1-month-old lens. Consistent identification of the germinative zone is complicated by distortion of the explant during dissection. During dissection, there is unavoidable damage to the lens epithelium (asterisks). Finally, the explant is difficult to keep flattened, leaving areas of the explant out of focus and less intense, making quantification more difficult. D: Lens explant from an 8-month-old mouse labeled with BrdU. E: Quantification of the percentage of lens cells in S-phase (EdU-positive) from an 8-month-old Balb/c mouse kept under normoxic conditions (room air; 21% O₂) compared to a mouse breathing 60% O₂. The asterisk indicates a p<0.05). Total cells (green) were stained with the vital dye, DRAQ-5. Scale bars: 10 μm (A, B) and 50 μm (C, D).