I've sacrificed a small number of mice via isofluorane + CO2 and quickly took all major organs and flash froze them with liquid nitrogen. The goal is to preserve the redox state of proteins in these major tissues for 2D gel proteomic analysis. I've been scanning some literature but it seems to me as though euthanasia methods aren't often published.
I'm wondering if cervical dislocation or decapitation (without isofluorane and CO2 beforehand) might be a better option. If you have any experience with this sort of thing, or could share the titles of some journal articles I could look into, I would really appreciate it. Thanks for your time!
I think you're on the right track. CO2 takes a while. When CO2 levels are carefully controlled, the method is considered humane, because the mice lose consciousness in a few seconds. However, over the next minute or so asphyxiation hypoxia generates gross physiological changes that affect metabolism at the cellular level in well-oxygenated tissues. Most ACUC will approve decapitation or cervical dislocation as more rapid acceptable alternatives, if the user exhibits expertise in these techniques.
Remember that tissues continue metabolism after the animal is sacrificed. So you need to harvest them in a rapid and reproducible fashion, especially tissues that normally exhibit very high or low oxygenation, and/or intrinsically relatively high or low metabolic rate (e.g., lung and colon, respectively). To slow redox changes after harvest do a quick rinse in ice-cold degassed saline containing a little EDTA, wrap in impermeable plastic food wrap to exclude air, then quick-freeze. Still, perform your analyses ASAP, because even frozen tissue metabolizes, albeit slowly.
if you already considered ethical aspects, then you still should consider refrasing your question. The term euthanasia is already taken and discussion can get ugly when used erroneously.
John Carter, thank you for getting back to me. I'm interested in your plastic wrap + saline/EDTA solution. What would you recommend as a final EDTA concentration in saline? 1-5mM?