Intensity over time measurements

[Shane Harding-2]

Hello,
I am attempting to measure the intensity of a ROI from a series of  
time lapse images with a GFP-labelled protein. There are two cases. In  
the first, there is a stable background nuclear expression of the  
protein which bleaches briefly and then recovers over ~5mins. The  
second case starts with a bright spot (the ROI) within the nucleus  
that resolves over ~1h.

My question is what is the best way to measure the intensity of a  
selected ROI over time while also accounting for photobleaching. I  
have tried the FRAP profiler and it seems to do a reasonable job. Is  
this appropriate for this type of analysis? What exactly does this  
algorithm do for normalization?

Thank you very much,
Shane

[Aryeh Weiss]

I find that when I set the max memory using
Options-->Memory&threads...

to a value over 1600 (eg, 2300MB), then I receive a warning that setting to a
value over 1600MB may cause IJ to not start. Fair enough -- but I am using
JRockit and in fact can  use a larger max memory.
So I click OK, but the max memory will be unchanged. If I put in a "legal" value
(ie under 1600MB), then it will be changed after the next startup.
I can work around this by manually entering the max memory in the ImageJ.cfg
file as -Xmx####m .

I think that this behavior is new, and might have been introduced as an
unintended side effect of putting in the warning.

--aryeh
--
Aryeh Weiss
School of Engineering
Bar Ilan University
Ramat Gan 52900 Israel

Ph:  972-3-5317638
FAX: [phone]

[Wayne Rasband]

On Jan 17, 2009, at 4:02 PM, Aryeh Weiss wrote:

> I find that when I set the max memory using
> Options-->Memory&threads...
>
> to a value over 1600 (eg, 2300MB), then I receive a warning that  
> setting to a value over 1600MB may cause IJ to not start. Fair  
> enough -- but I am using JRockit and in fact can  use a larger max  
> memory.
> So I click OK, but the max memory will be unchanged. If I put in a  
> "legal" value (ie under 1600MB), then it will be changed after the  
> next startup.
> I can work around this by manually entering the max memory in the  
> ImageJ.cfg file as -Xmx####m .
>
> I think that this behavior is new, and might have been introduced  
> as an unintended side effect of putting in the warning.

This bug is fixed in the v1.42h daily build.

-wayne

[Joris FA Meys]

Hi,

maybe a bit late to answer, but my plugin Frap_Norm aids with both measurement and normalization, and can be found at :
http://imagejdocu.tudor.lu/doku.php?id=plugin:analysis:frap_normalization:start

Hope this helps

Kind regards
Joris

Shane Harding-2 wrote

Hello,
I am attempting to measure the intensity of a ROI from a series of  
time lapse images with a GFP-labelled protein. There are two cases. In  
the first, there is a stable background nuclear expression of the  
protein which bleaches briefly and then recovers over ~5mins. The  
second case starts with a bright spot (the ROI) within the nucleus  
that resolves over ~1h.

My question is what is the best way to measure the intensity of a  
selected ROI over time while also accounting for photobleaching. I  
have tried the FRAP profiler and it seems to do a reasonable job. Is  
this appropriate for this type of analysis? What exactly does this  
algorithm do for normalization?

Thank you very much,
Shane