Table 1 Flow cytometry controls

1. Mesonephroi × 4

No

• Click-iT 647

Negative sample for establishing gates in the 647 nm and 488 nm channels

• MVH-alexa fluor 488

• Propidium iodide

2. Gonads × 4

Yes

• Click-iT 647

Positive controls for setting the laser power in the 488 nm and 647 nm channels. Use females gonads for this if concentrating of male germ cell development and vice-versa if concentrating on female germ cell development (or a mix, if concentrating on both).

• MVH-alexa fluor 488

• Propidium iodide

3. Mesonephroi × 2 (optional)

Yes

• Click-iT 647 only

Single color control for staining compensation

4. Mesonephroi × 2 (optional)

No

• Propidium iodide only

Single color control for staining compensation

5. Gonads × 2 (optional)

No

• MVH-alexa fluor 488 only

Single color control for staining compensation