Cell Counting (DAPI stain)

Dear Sir/Madam,

I would like to count my cells (white dots in black background) using the following code, I would highly appreciate your help if you kindly adjust this code to read only one frame TifF file.

Best wishes Peyman

%___________________ Cell Counting

Open This Example This example shows how to use a combination of basic morphological operators and blob analysis to extract information from a video stream. In this case, the example counts the number of E. Coli bacteria in each video frame. Note that the cells are of varying brightness, which makes the task of segmentation more challenging.

Introduction Initialization Stream Processing Loop Summary Data Set Credits Introduction This example illustrates how to use the morphological and BlobAnalysis System objects to segment individual cells and count them.

Initialization Use these next sections of code to initialize the required variables and objects.

VideoSize = [432 528]; Create a System object™ to read video from avi file.

filename = 'ecolicells.avi'; hvfr = vision.VideoFileReader(filename, ... 'ImageColorSpace', 'Intensity',... 'VideoOutputDataType', 'single'); Create an Autothreshold System object to convert an intensity image to a binary image.

hautoth = vision.Autothresholder( ... 'Operator', '<=', ... 'ThresholdScaleFactor', 0.8); Create a BlobAnalysis System object to find the centroid of the segmented cells in the video.

hblob = vision.BlobAnalysis( ... 'AreaOutputPort', false, ... 'BoundingBoxOutputPort', false, ... 'OutputDataType', 'single', ... 'MinimumBlobArea', 7, ... 'MaximumBlobArea', 300, ... 'MaximumCount', 1500);

% Acknowledgement ackText = ['Data set courtesy of Jonathan Young and Michael Elowitz, ' ... 'California Institute of Technology']; Create a System object to display the video.

hVideo = vision.VideoPlayer; hVideo.Name = 'Results'; hVideo.Position(1) = round(hVideo.Position(1)); hVideo.Position(2) = round(hVideo.Position(2)); hVideo.Position([4 3]) = 30+VideoSize; Stream Processing Loop Create a processing loop to count the number of cells in the input video. This loop uses the System objects you instantiated above.

frameCount = int16(1); while ~isDone(hvfr) % Read input video frame image = step(hvfr);

    % Apply a combination of morphological dilation and image arithmetic
    % operations to remove uneven illumination and to emphasize the
    % boundaries between the cells.
    y1 = 2*image - imdilate(image, strel('square',7));
    y1(y1<0) = 0;
    y1(y1>1) = 1;
    y2 = imdilate(y1, strel('square',7)) - y1;

    y3 = step(hautoth, y2);       % Binarize the image.
    Centroid = step(hblob, y3);   % Calculate the centroid
    numBlobs = size(Centroid,1);  % and number of cells.
    % Display the number of frames and cells.
    frameBlobTxt = sprintf('Frame %d, Count %d', frameCount, numBlobs);
    image = insertText(image, [1 1], frameBlobTxt, ...
            'FontSize', 16, 'BoxOpacity', 0, 'TextColor', 'white');
    image = insertText(image, [1 size(image,1)], ackText, ...
            'FontSize', 10, 'AnchorPoint', 'LeftBottom', ...
            'BoxOpacity', 0, 'TextColor', 'white');

    % Display video
    image_out = insertMarker(image, Centroid, '*', 'Color', 'green');
    step(hVideo, image_out);

    frameCount = frameCount + 1;
end

release(hvfr); % close the video file