Flowcytometry for immunostained mitohcondria - (Aug/11/2006 )

Can anybody tell me the detailed method to immunostain mitochondria for flowcytometric analysis. I have some information but it is not detailed (especially centrifugation steps)... would like to hear from somebody experienced in this method... any precautions etc????

On passing, can we detect an increase in a particular protein in mitochondria by this method? I mean the protein is in mitochondria (very less amount) under normal conditions but it is increased significantly after treatment... and can we detect this increase by fluorescence microscopy (like increase in number of immunostained mitochondria) ???

Better to use western blot to look for increase in protein.

Using fluorescence microscopy, I am not sure if u could quantify the amount of protein. Especially mitochondrial protein as detecting a mitochondrial protein by immunostaining looks punctate. Therefoe abundance of some mitochondrial protein, I am not sure.

I would prefer Western to compare protein levels and Use fluoroscence to detect presence of protein.