Re: Colocalization question

Posted by Barlow, Andrew on
URL: http://confocal-microscopy-list.275.s1.nabble.com/Dyes-for-apoplastic-pH-in-leaf-tissue-tp5505884p5510339.html

Hello Leong,
You can measure correlation coefficients based around a region of interest in Volocity by selecting a region of interest in the Colocalization view using any ROI tool, or you can make a measurement protocol in the measurements view to select either single or multiple ROIs. You can then either add a "Calculate object colocalisation" task (for per object measurements) or select the colocalization view.

In the colocalization view coefficients are always based on the ROI if one is present.

If you wish to answer your bottom line question, "how much of the red protein co-localizes to green within the object and not the whole cell?" you should study Manders' coefficients which provide a good measurement of co-localization.  Use Pearson's correlation coefficient to get a good measurement of the quality of the relationship.

If you require any assistance with this please do call our technical support team or send me a message off list.
Regards,
Andrew


Andrew Barlow PhD | Applications Specialist
PerkinElmer | For the Better
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> -----Original Message-----
> From: Confocal Microscopy List
> [mailto:[hidden email]] On Behalf Of Teng-Leong Chew
> Sent: 07 September 2010 20:00
> To: [hidden email]
> Subject: Colocalization question
>
> Hi all,
>
> Pardon me for opening up a can of worms of colocalization issue.
>
> I wonder if anyone has experience in thresholding a biological
> object using a channel (say, green actin in a contractile ring)
> and use only that selected ROI to quantify Pearson coeefficient
> between green and a red-labeled protein? Bottom line question
> is: how much of the red protein co-localizes to green within
> the object and not the whole cell?
>
> We have Volocity, MetaMorph and ImageJ.
>
> Thanks in advance for your help.
>
> Regards,
> Leong
>
>
> --
> Teng-Leong Chew, Ph.D.
> Director, Cell Imaging Facility & Nikon Imaging Center
> Director of University Imaging Resources
> Feinberg School of Medicine & Office of Research
> Northwestern University
> 303 E. Chicago Avenue
> Chicago, IL 60611
> [phone]
> [phone] (W, F)