One of the ways to solve flickering or changes in intensity in a movie is to float the stack.
Left: brightfield.
Middle: reflection raw data
Right: reflection after float function applied.
(For bigger AVI version, click here.)
This is how we floated the stack:
//--------------------------------------------------------- //Written by Kenton Arkill [underwaterginge@GMAIL.COM] //Floating means that each image has the same mean pixel value and the same standard deviation. //I find it really useful as it means the same threshold can often be used on all the images. Here is my macro, //it floats the stack to the values of the first slice (so make sure it is a good image): macro "Float the stack"{ run("32-bit"); setSlice(1); // reference slice; could be set to something other than 1 getRawStatistics(nPixels, meanref, min, max, stdref, histogram); for (i = 1; i <= nSlices; i++) { setSlice(i); getRawStatistics(nPixels, meani, min, max, stdi, histogram); run("Subtract...", "value="+meani+" slice"); run("Divide...", "value="+stdi+" slice"); run("Multiply...", "value="+stdref+" slice"); run("Add...", "value="+stdref+" slice"); } run("Enhance Contrast", "saturated=0.35"); // could change to zero setSlice(1); }