Selecting correct threshold algorithm

Posted by tmurp002 on Apr 18, 2014; 5:49pm
URL: http://imagej.273.s1.nabble.com/Selecting-correct-threshold-algorithm-tp5007356.html

Hi all,

Hopefully this question hasn't been answered before (I've searched back quite a bit into the archives and found nothing). I'm performing a set of experiments looking at cell swelling in brain slices under certain conditions over short intervals of time. To visualize this I'm using patch-clamp to load the cells with a membrane-impermeant fluorescent dye and imaging via CSLM. My standard protocol involves taking rapid z-stacks through the soma (to capture all edges of the cell body), then using max intensity projections through these stacks to compare the soma area between time points. Through a lot of trial and error I've come to the strategy of using Fiji to filter, align, crop, and threshold the images for each cell, and perform measurements on the thresholded area.

My problem is essentially figuring out how to pick the appropriate thresholding algorithm. I've basically settled on the Mean threshold method, because it is the only one which shows the same features I observe in the original image. It also would stand to reason (at least to me) that this method would appropriately account for intensity fluctuations in the soma (for example, lowering the threshold if average intensity drops, as is often the case when the cell takes on water).

Am I thinking about this correctly, or is there some large issue I'm overlooking which invalidates this method?