Mitochondrial Distribution

Hi all,

I’m a new user of Cell profiler, and therefore have no expertise in building a suitable pipeline for my needs. I was wondering if anyone might have a pipeline already made for the analysis I need to do, or whether anyone could advise me on which modules to use.

The overexpression of our protein of interest leads to a shift in the distribution of the mitochondria from a diffuse cytoplasmic distribution to a perinuclear one. We would like to enumerate this change in distribution, so that we can then try and reverse the phenotype experimentally using siRNAs, compounds, or to compare mutants of our protein of interest to the wild-type protein.

I attach two images to represent the sort of phenotype I’d like to analyse.

http://i3.photobucket.com/albums/y81/LeInspector/MitoDistribution_zps0fe0b1cb.png

Many thanks in advance!

Luke (UCL)

Hi Luke,

Yes, in fact we have published a paper with just this type of analysis:
Abnormalities in mitochondrial structure in cells from patients with bipolar disorder.
Cataldo AM, McPhie DL, Lange NT, Punzell S, Elmiligy S, Ye NZ, Froimowitz MP, Hassinger LC, Menesale EB, Sargent LW, Logan DJ, Carpenter AE, Cohen BM.

I see that we didn’t publish the pipeline on our normal page for this on, for some reason. I will look for it and post a link here soon.

But in general, you would want to:
LoadImages
IdentifyPrimaryObjects (using nuclear channel)
IdentifySecondaryObjects (call them “Cells”, using whatever your red channel is)
IdentifyTertiaryObjects (“Cytoplasm” = Secondary minus Primary)
then add various measure modules…
But most importantly, MeasureRadialDistribution, which defines concentric annuli from the nucleus outward and measures whatever you want it to in each annulus.

(Note: there may be some updates to this module that are not in the 11710 release version, though I can’t recall offhand. Post back here if it doesn’t allow you to choose the nuclei as a starting point to measure your annuli).

There are finer questions – do you want to measure the coverage of mito, or weight them by their intensity? You can do both, but the analysis can be more complicated.

In any case, let us know how that sounds.
David

Hi David,

Fantastic, thanks. If at all possible, I’ll wait for the pipeline to be located before giving it a go, but it’s very encouraging to know it’s been done.

Best wishes, and looking forward to giving the analysis a go.

Luke

I am also interested in that pipeline -was it ever posted?

No, but it will be soon! I will make a note to reply again here when done.

The Cataldo et al. 2010 pipeline is now posted here: http://www.cellprofiler.org/published_pipelines.shtml.

Thank you so much!! That’s exactly what I was looking for.

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