Isolation of Mitochondria

[David George]

Guys do you have any idea how i can isolate mitochondria.If i centrifuge i think the mitochondria is going to get damaged.Please correct me if i am wrong...
Thanks

[canalon]

Centrifugation in a density gradient is the way to go, as far as I know. something like Ficoll is probably used, I cannot imagine sucrose being very good...

[hplc]

If you have homogenated cells you can use differential centrifugation. It's easy, and gives good results if you measure very exactly. If you sentrifugate the cells for 10 minutes with 1,000 g you should get pellet rich in nuclei and cellular debris. Another 20 minutes with 20,000 g and you should have pellet rich in mithocondria (and chloroplasts if you are talking about plant cells.) Then, extract the mithochondria carefully with a syringe or a vacuum-pipe, and you should have the mithocondria pretty isolated.