Aug 29, 2024
Treatment of D. melanogaster with small molecules in fly food
- Natalie Kaempf1,
- Patrik Verstreken1
- 1VIB-KU Leuven Center for Brain & Disease Research, 3000 Leuven, Belgium.
Protocol Citation: Natalie Kaempf, Patrik Verstreken 2024. Treatment of D. melanogaster with small molecules in fly food. protocols.io https://dx.doi.org/10.17504/protocols.io.bp2l62rozgqe/v1
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: July 29, 2024
Last Modified: August 29, 2024
Protocol Integer ID: 104248
Keywords: ASAPCRN, fly food treatment, Q10, L-DOPA, R55
Funders Acknowledgement:
Aligning Science Across Parkinson’s
Grant ID: ASAP-000430
EMBO long-term postdoctoral fellowship
Grant ID: ALTF_299-2019
Research project, FWO Vlaanderen
Grant ID: G0A5219N
Research project, FWO Vlaanderen
Grant ID: G0B8119N
Methusalem project
Grant ID: METH/21/05 (3M210778)
Research project, KU Leuven Parkinson Fonds
Grant ID: EQZ-PARFON-O2010
Opening the Future grant, Leuvens Universiteitsfonds (LUF)
Grant ID: EQZ-OPTFUP-O2010
Research project, FWO Vlaanderen
Grant ID: G031324N
Abstract
This protocol describes the preparation of fly food with small molecules to treat flies during their aging. It can be adapted for any treatments. Details for small molecules used can be found in the "Materials" section
Materials
Coenzyme Q10, Merck, Cat# C9538-1G
- 50 mM stock solution in ethanol (99.8%), aliquoted in single-use portions and stored at -20°C
- final concentration of 1 mM with 2% ethanol
Retromer Chaperone, R55, Merck, Cat# 5310840001
- R55 (Calbiochem/Millipore) was dissolved in sterile distilled water as a 12 mM stock solution, aliquoted in single-use portions and stored at -20°C
- final concentration of 12 µM
Levodopa (L-Dopa), Merck, Cat# PHR1271-500MG
- 15.2 mM stock in sterile distilled water on ice and stored at -80°C in single-use aliquots
- 1 mM final concentration
3,4-Dihydroxy-D-phenylalanine (D-Dopa), Merck, Cat# D9378-250MG
- 15.2 mM stock in sterile distilled water on ice and stored at -80°C in single-use aliquots
- 1 mM final concentration
Food preparation
Food preparation
1m
1m
warm up the amount of fly food (standard corn meal and molasses food) required (e.g. 50ml for 45 XS vials) in the microwave, stir it in between, start with 1.5 min, mix the food, heat up for 1min again and continue until food has no clumbs anymore
prepare XS vials (45 for each condition when 50ml/condition) and label them, place them in plastic boxes
measure the amount of food in measuring cylinder and let it cool down in the beaker to around40 °C (immediately clean all glass ware after usage)
add compound/solvent and stir very well (00:01:00 ), to keep the food liquid put in water bath to maintain the temperature
1m
pipette around 1 mL per XS vial with 10 or 25 ml pipette
Make sure the food is filling the whole bottom of the vial and shows even surface to avoid flies getting stuck in small wholes when aging
store the food in darkness at 4 °C in the cold room and clean up
use the food not longer than 2 weeks
fly treatment
fly treatment
depending on the experimental condition flies are exposed to the treatment and corresponding control after they are collected (Q10 or R55 treatment) or 10 days prior the experiment (D-/L-DOPA treatment)
flies on treatment/control food are kept in a temperature and light-controlled incubator at
25°C and 12-hour light-dark cycle
flies are flipped to new food every two to three days
On the day of the experiment flies are flipped on fresh treatment/control food at Zeitgeber zero to one