![]() ![]() You should see windows appear for all your channels in the “Live View” window. When the machine is ready to start, the next button, “Play”, will change from a grey arrow to a black arrow. First click the “Ready” button (bottom left). You will use the start controls in the “Registered Point List” window instead of the “Image Acquisition Control” window. The coordinates for each square should then appear in the Registered Point List window. When you have entered all the squares, click “Apply” in the “Mosaic Outline” window. Click on the next square on interest in the MATL Controller window and repeat. Click the 1st button (red on black) to enter the coordinates. A window (“Mosaic Outline”) will open that will show the coordinates of each entered square. Click a square you want included, and then click the “Mosaic Outline” button. If you define an irregular matrix, the program will fill in squares to create a rectangle. The “Mosaic Outline” button (2nd button from the left on the MATL Controller window) will allow you more direct control over the location of the matrix, but all the points must be entered manually. The coordinates for each square will be loaded into the Registered Point List. The “Define Matrix” button (3rd from the left in the MATL Controller window) will allow you to specify a set number of squares across and down.Ĭlick “Set” and the yellow crosshairs will be centered in the defined matrix. You can also double click any square on the MATL controller grid to move the field of view to that position. When you find a region of interest, it is a good idea to jot down the coordinates in case you stray too far from it. The windows at the top are particularly useful as they will allow you to go back to any set of coordinates on your side. The stage controller allows you to fine control x-y movement from the computer. You can zoom in or out with the slider bar to the right. The x y coordinates of the yellow crosshair (relative to the center) are shown to the right of the grid (and in the windows of the stage controller). The red cross marks the center of the grid. The yellow crosshair shows the position of the sample in the view field on the grid (“you are here”). Each square on the grid represents the x-y plane that can be imaged through the objective. ![]() The MATL Controller window displays a grid. The “Normal” scan pattern is recommended. ![]() You can still do stitching, but seams are possible in the final image. If you must do any quantitation on your images, you cannot use overlap. X zoom#)/ 8 ) + 3 = ideal overlap%įor example, if you are using a 60X objective with a zoom of 1.5, the ideal overlap is around 14%, and you would set the overlap # to 86%. The optimal amount of overlap is determined by the total magnification: Some overlap is required for optimal image stitching to produce the most seamless combined image. The overlap setting is in the window next to the % sign. The 3rd button from the right on the MATL Control window opens the “Preferences” window. You also have the option to name your image files (this will show up later on the Registered Point List spreadsheet). If you want to do Time Lapse, see below (“Setting up time-lapse mosaics”).Ĭlick the 4th button from the right (disc & wrench) on the Registered Point List window, which will allow you to set the folder destination (Folder Name window). If you experience a problem with getting the MATL program to accept Z-coordinates, follow the instructions HERE. It is essential to click “Depth” BEFORE setting up your matrix, otherwise the program will function only in 2 dimensions. If you wish to merge Z-stacks, set your start and end points, then click the “Depth” button in the “Image Acquisition Control “ window. Load your dyes (virtual phases cannot be done in combination with MATL), find a region of interest on your slide, set exposure levels, etc. You should see 3 new windows: Stage Control, MATL Control, and Registered Point List (more detail on those later). When it is finished moving, you can place your slide/dish on the stage. This will cause the stage to move around. The “XY stage warning” window will pop up with the statement: “This stage will move in a wide range to detect its mechanical origin”. Mosaic stitching allows you to take a series of overlapping images and merge them together into a seamless picture of the entire subject.īEFORE you place your slide/dish on the microscope, start the FV1000 software, and select “Multi Area Time Lapse” (MATL) from the “Device” tab on the top of the screen. Many samples are larger than the microscope’s field of view, but sometimes you wish to produce an image that shows the whole sample.
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