Negative values in Watershed algorithm leading to black image - matlab

Question

I'm using the watershed algorithm to try and segment touching nuclei. A typical image may look like:
or this:
I'm trying to apply the watershed algorithm with this code:
show(RGB_img)
%Convert to grayscale image
I = rgb2gray(RGB_img);
%Take structuring element of a disk of size 10, for the morphological transformations
%Attempt to subtract the background from the image: top hat is the
%subtraction of the open image from the original
%Morphological transformation to subtract background noise from the image
%Tophat is the subtraction of an opened image from the original. Remove all
%images smaller than the structuring element of 10
I1 = imtophat(I, strel('disk', 10));
%Increases contrast
I2 = imadjust(I1);
%show(I2,'contrast')
%Assume we have background and foreground and assess thresh as such
level = graythresh(I2);
%Convert to binary image based on graythreshold
BW = im2bw(I2,level);
show(BW,'C');
BW = bwareaopen(BW,8);
show(BW,'C2');
BW = bwdist(BW) <= 1;
show(BW,'joined');
%Complement because we want image to be black and background white
C = ~BW;
%Use distance tranform to find nearest nonzero values from every pixel
D = -bwdist(C);
%Assign Minus infinity values to the values of C inside of the D image
% Modify the image so that the background pixels and the extended maxima
% pixels are forced to be the only local minima in the image (So you could
% hypothetically fill in water on the image
D(C) = -Inf;
%Gets 0 for all watershed lines and integers for each object (basins)
L = watershed(D);
show(L,'L');
%Takes the labels and converts to an RGB (Using hot colormap)
fin = label2rgb(L,'hot','w');
% show(fin,'fin');
im = I;
%Superimpose ridgelines,L has all of them as 0 -> so mark these as 0(black)
im(L==0)=0;
clean_img = L;
show(clean_img)
After C = ~BW; the whole image goes dark. I believe this is because the image pixels are all -inf or some smaller negative number. This is there a way around this and if so what could I change in my code to get this algorithm working? I've experimented a ton and I don't really know what's happening. Any help would be great!

Answer 1

The problem is with your show command. As you said in the comments this uses imshow under the hood. If you try imshow directly you'll see you also get a black image. However, if you call it with appropriate limits:
imshow(clean_img,[min(clean_img(:)), max(clean_img(:))])
you'll see everything you expect to see.
In general I usually prefer imagesc for that reason. imshow makes arbitrary judgements as to what range to represent, and I usually can't be bothered to keep up with it. I think in your case, your end image is uint16 so imshow chooses to represent the range [1, 65025]. Since all your pixel values are below 400, they look black to the naked eye for that range.