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Materials list is empty in the source — the heading is present with a single bullet containing nothing. Someone should fill it in.
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Advance preparation
Bleach synchronize worm strains and hatch 1,000 embryos on unseeded 10 cm NGM plates for L1 assays, or seeded 10 cm NGM plates for adult assays.
Adult assay
- Four days after bleaching, wash worms off the 10 cm plates with M9 into 1.5 mL Eppendorf tubes.
- Wash and prepare worms:
- Wash worms from 10 cm NGM plates into 1.5 mL Eppendorf tubes using 1 mL of M9.
- Allow worms to settle for 2 minutes.
- Remove the supernatant without disturbing the worm pellet.
- Add 1 mL of M9 and invert to wash.
- Repeat the settle, remove and wash steps twice more, for a total of three M9 washes.
- After the final wash, add the desired drugs plus 10 mM serotonin and wait 20 minutes.
- Pipette worms and solution onto an unseeded 10 cm plate and wait 1–2 minutes.
- Pick 5–8 worms without using OP50 to unseeded 6 cm assay plates, GFP+ where needed. Wait 10 minutes before counting pharyngeal pumps.
- Use the Axio injection scope to count pumps for 10 seconds. One pump is the movement of the terminal bulb down and back to its starting position.
L1 assay
- One day after bleaching, wash worms off the 10 cm plates with M9 into 1.5 mL Eppendorf tubes.
- Prepare worms by washing them from 10 cm NGM plates into 1.5 mL Eppendorf tubes using 1 mL of M9.
- Add the desired drugs and wait 20 minutes.
- Pipette worms and solution onto an unseeded 10 cm plate and wait 5 minutes.
- Pick 5–8 worms to seeded 6 cm assay plates, GFP+ where needed. Wait 10 minutes before counting pharyngeal pumps.
- Use the Axio injection scope to count pumps for 10 seconds. One pump is the movement of the terminal bulb down and back to its starting position.
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Check the L1 assay. Step 2 omits the three M9 washes described in the adult protocol, and step 5 uses seeded plates where the adult assay uses unseeded. Both may be deliberate, but the difference isn't explained.
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