[David Pirman]

Ramin,

So for quantitation, it can be a bit tricky, but you are correct. You should definitely think about ng/area and then even volume (since you know your tissue thickness). I generate standard curves by spotting calibration standards beneath my tissue (in a fixed circular area) average the intensity within the spot and then calculate the volume of tissue. Then I would quantify with same exact area size somewhere else on the tissue, but I haven’t had much luck in this without an internal standard. So in my case, my calibration standards were in a nice confined area with a uniformly applied internal standard. If you are trying to generate quantitative data from a compounds with unknown area density information, I’d think it would be quite difficult since the calibration curve is heavily dependent on the calculation of area on the x-axis. I just had a paper accepted in Analytical Chemistry using this technique for quantitation.
For calculating the area accurately in DESI, I would suggest drawing an ROI and overlaying the image onto an optical image of your tissue and basically doing a manual measurement of the area you are interested in. Also, placing some distance registration marks on the MALDI target. I typically use glass slides, and will simply draw marks of interest on the backside so when I take an optical image I can make measurements and co-register the MS image.

Dave

[David Pirman]

Ramin,

I do remember something funny about the area when I was doing some quantitative work. I would typically measure my area by using my step size and laser diameter with a known amount of scans across the ROI, followed by a simple area calculation. This is what I would recommend.

There seems to be a few quirks with software when using it for quantitative applications. If I come across any more, I’ll post them on this forum.

Dave

[David Pirman]

When you click on statistics, you should see a window with some bar graphs and tabulated data. One of the columns is says volume (mm^2). I think this is the area since its in squared units. There is also a column of population which should be the # averaged spectra. I assume if the two ROI’s have the same population#, they should be same area.
If this statement is wrong, hopefully someone can comment further.

Dave

[David Pirman]

I believe the value N corresponds to the m/z. I’ve noticed there may be an error between the value N in the image pane and the statistics pane. It seems the stats pane should be N+1 of the image pane.

Dave

[David Pirman]

Here’s a screen shot.The top stats pane has the same N as the image and the bottom has N+1 [img size=679]http://www.maldi-msi.org/images/fbfiles/images/ROI_error.jpg[/img]

Attachments:

• ROI_error.jpg

[David Pirman]

You should be able to click on ROI–>statistics. You must ensure that the N(m/z) values match the particular m/z image you are interested in. Actually I think there’s an error, where the N value in the stats window should be +1 from the image.

Dave Pirman

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