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    • CommentAuthorelpitts
    • CommentTimeDec 18th 2010
    1) Open Photoshop on the computer
    2) Click File > Open
    3) Select the images that you want to overlay and open them in photoshop
    4) Make sure there is a saved copy of all the original images
    5) Save the images under new names before you begin manipulating them, so that you will always have an unchanged original
    6) Once you have opened all of your images, pick one to be your base image (In this example, the base image will be green, and the other two images will be red and blue).
    7) Bring the blue fluoresent image to the front
    8) Go to Image> Select> All
    9) Then, go to File> Copy (or Ctrl C)
    10) Once you have a copy of the blue image, go back to your base green image and click on the channels tab (next to the layers tab at the bottom right)
    11) Highlight the blue channel
    12) Paste (File> Paste or Ctrl V) your copied blue image into the blue channel
    13) Repeat these steps copying the red image into the red channel
    14) Now save the overlay image as a photoshop and a JPEG file (make sure that you are saving it under a new name and not replacing your original green image).
    • CommentAuthorroclemente
    • CommentTimeSep 29th 2011
    Be sure that when you highlight the channel you desire, you aren't simply clicking on the eye. The eye only makes the channel visible or invisible but doesn't actually select the channel you want to work on.
    • CommentAuthorvibeamer
    • CommentTimeSep 30th 2011
    Make sure the levels of the fluorescent image have been increased to a point that it shows. The separate images may show a great representation of the fluorescence, but the composite image can be more difficult to see distinctions. Adjusting the levels can make the visibility of each distinct fluorescing piece easier.
    • CommentAuthoralfleming
    • CommentTimeOct 1st 2011
    It is not completely necessary to bring the blue image to the front. Sometimes your image could be better if you place the green fluorescent or red fluorescent image at the front of the channel. Also save the image as a photoshop image in case you need to make adjustments later. Once the file is saved as a JPEG you cannot undo or change edits you made in photoshop.