Lipids in Drosophila - (Jun/27/2008 )
Dear all,
My name is Joana, and I am currently starting to work with lipids.
I use Drosophila as a model system.
Since I am new to this field, I am having some difficulties in getting enough good quality lipid extracts in my samples.
Does anyone knows a protocol suited for experiments using Drosophila tissue?
Most protocols I see use the Folch or Bling Dyer Methods.
In these protocols, after concentration of the sample (by speed vac), in what solvent do you ressuspend your lipids? (I will be quantifying the lipids using colorimetric methods, so if I can avoid chloroform or methanol, that would be excelent, since the 96-well plates I have are not resistant to ant of these solvents).
Also, does anyone knows if this "concentration" step be done just by "air-drying" (I do not have a speed vac)?
Thanks,
Joana
My name is Joana, and I am currently starting to work with lipids.
I use Drosophila as a model system.
Since I am new to this field, I am having some difficulties in getting enough good quality lipid extracts in my samples.
Does anyone knows a protocol suited for experiments using Drosophila tissue?
Most protocols I see use the Folch or Bling Dyer Methods.
In these protocols, after concentration of the sample (by speed vac), in what solvent do you ressuspend your lipids? (I will be quantifying the lipids using colorimetric methods, so if I can avoid chloroform or methanol, that would be excelent, since the 96-well plates I have are not resistant to ant of these solvents).
Also, does anyone knows if this "concentration" step be done just by "air-drying" (I do not have a speed vac)?
Thanks,
Joana
I used glass vials, but with bigger insects. I directed a stream of nitrogen inside to evaporate the solvents and then I dried the samples until they had a constant weight in an oven (40°C) and then exsiccator for storage/transport. But I used a gravimetric method and all water had to be evaporated. If you're using a colorimetric method, then at least the solvent has to be evaporated completely as it is also has energy, if I think correct.
Instead of chloroform you can use also dichloromethane, not so harmful but also dissolving plastics. But perhaps you can pool the flies from one treatment for the analysis and then use bigger glass vials?
Thanks for the input.
My problem is in what solvent can I ressuped the lipids and still use it for colorimetric measurements...
Best,
Joana
My name is Joana, and I am currently starting to work with lipids.
I use Drosophila as a model system.
Since I am new to this field, I am having some difficulties in getting enough good quality lipid extracts in my samples.
Does anyone knows a protocol suited for experiments using Drosophila tissue?
Most protocols I see use the Folch or Bling Dyer Methods.
In these protocols, after concentration of the sample (by speed vac), in what solvent do you ressuspend your lipids? (I will be quantifying the lipids using colorimetric methods, so if I can avoid chloroform or methanol, that would be excelent, since the 96-well plates I have are not resistant to ant of these solvents).
Also, does anyone knows if this "concentration" step be done just by "air-drying" (I do not have a speed vac)?
Thanks,
Joana
I used glass vials, but with bigger insects. I directed a stream of nitrogen inside to evaporate the solvents and then I dried the samples until they had a constant weight in an oven (40°C) and then exsiccator for storage/transport. But I used a gravimetric method and all water had to be evaporated. If you're using a colorimetric method, then at least the solvent has to be evaporated completely as it is also has energy, if I think correct.
Instead of chloroform you can use also dichloromethane, not so harmful but also dissolving plastics. But perhaps you can pool the flies from one treatment for the analysis and then use bigger glass vials?