Tests to identify cell organelles?

[DarpaChief]

We finished a lab in class a few days ago involving cell centrifugation to isolate specific organelles and one of the questions is asking, what are other tests to identify cell organelles?

I've been searching online for a while and cant seem to find anything...other than that we can see other cells using a transmission electron microscope

any help would be greatly appreciated

[JackBean]

youcan use some proteins. Either enzymatic activity (than it must be unique to the compartment) or blott (than there must be at least one distinguishable isoform in that compartment).

For some, you could use evolution of gases.

[DarpaChief]

interesting, what do you mean by evolution of gases?

[JackBean]

of course, I don't mean Darwin's evolution I mean release of gases like CO2 from mitochondria, O2 from chloroplasts or hydrogen peroxide from peroxisomes (although that's not gas, right?:( )

[sunghoo]

well ultimately, hydrogen peroxide becomes water and oxygen, so i guess it evolves into gas.. unless enzymes are denatured...

[MrMistery]

well if you have organelles isolated and your procedure didn't **** them up, then you can do EM and just look at them, you should be able to recognize what they are if your fraction is pure enough. EM would also have the advantage that it will give you information on how pure and intact the organelles are. If you just do enzyme assays and western blotting, you don't know purity - ex you can blot for Kar2p/BiP, see a band and determine you have an ER fraction, but really you might have a ton of mitochondria and lysosomes there, you have no way of knowing.
I guess it depends on what you want to do with them further on

[JackBean]

you can always determine markers of other organelles