Watershed Algorithm setting removing all connected components - 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)
For whatever reason after C = ~BW; the whole image goes dark. This very same code block has worked on a handful of other images, all of which were more "solid" or not as grainy as these. However, I thought I compensated for this with BW = bwdist(BW) <= 1;. I've experimented a ton and I don't really know what's happening. Any help would be great!
Ps. this is the image after BW = bwareaopen(BW,8);

Answer 1

Before the top-hat, you should perform a closing and an opening in order to reduce the noise.
If you perform an area opening on a noisy image, you may end up with the result on your black and white image.
So it would be:
Closing and opening
Top-Hat
Area opening if still necessary
Thresholding
Erosion and dilation to find the inner and outer markers respectively
Watershed (never use a watershed without markers).