I want to collect mice blood(post sacrifice) for the analysis of metabolites by GC-MS.Will there be any effect of Chloroform anesthesia on the metabolites?
Hi Bhuwan
In all our mouse euthanasia experiments for organ collection we use euthanasia by overdosing with Isoflurane. These are performed singly per mouse so that blood can be collected by cardiac puncture immediately that the mouse heart stops. The blood is collected into chilled heparin pre-rinsed syringes and transferred to heparin pediatric blood tubes which are centrifuged to collect the plasma or into clean syringes and tubes where clotting can take place at ambient temperature for 30 minutes and centrifuged to get serum. The serum method does run a risk of metabolite chnges due to plasma/serum enzyme activity.
Chloroform takes longer and CO2 that also takes longer than Isoflurane alters the pH of the blood and organs that can affect the metabolite profiles.
Generally your animal house would have recommended SOPs in place for euthanasia that would meet the relevant Ethics Committee guidelines or requirements and these should be followed.
If you are going to use chloroform then the best way to ascertain if there is a biological respond to chloroform itself is to remove the chloroform signals from your dataset - the following paper gives GC related parameters (https://www.osha.gov/dts/sltc/methods/organic/org005/org005.html)
and then on the 'clean' dataset perform a pre/post analysis on the control group to identify metabolites that change after chloroform administration. This is good practice anyhow to look at the effect of the sham treatment on the control group.
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