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One of the best things about computers is how they handle repetitive tasks. Show the computer how to do something once, and it will do it a million times without the slightest complaint. In FlowJo's Layout Editor, you can create a layout based on graphs and statistics from one of your samples. With a single click, you can ask it to produce the same layout from all of the rest of your samples. Or any subset that makes sense for your experiment. There are a couple of terms that we use in this explanation that are important to get clear from the start. They are attribute and value. The keywords in an FCS file are organized in attribute-value pairs. The attribute is a the keyword that is found in most files. The second is the value of that keyword in any given file. For example, most files will contain, in their header, a $Date attribute, and a value containing the date on which that sample was collected. When we talk about iteration attribute we mean the attribute that is changed for each frame of the stack that is generated, for instance the date of the collection. Generally, you would select a keyword that groups sets of tubes into distinct units--for example, choosing a keyword that connotes patient ID, tissue type, or experimental condition. Then you can place multiple graphs from different tubes (from different FCS files!) into the same layout and still generate a batch output. For example, if you set the iteration attribute to be the keyword "$Cells", where you have a Patient ID entered into the $Cells keyword, then FlowJo will examine all of the tubes in the current workspace, and generate a list of the unique values of $Cells. These are then displayed in the "Iteration" popup menu. By selecting any of these values from this menu, FlowJo will change the current layout view to show graphs & statistics drawn only from tubes with that value of the Iterator. (By setting this popup menu to "Off", you are telling FlowJo to use all of the original graphs that dragged and dropped into the layout). This is the way in which you can build template reports which select graphs and statistics from different tubes and put them all on the same page, but retain the ability to batch the outputs. The most common case remains that where the iteration attribute is the sample (by which we generally mean the tube run through the cytometer, as opposed to the sample taken from the subject). Iterating in the Layout Editor The first thing to be aware of is what is the currently active group in the Workspace window. This will be the selected item in the top half of the window. This is important to know because the current group will determine the set of samples, from which the layout will be constructed. Next you should look at the popup menu in the layout. In most cases, you'll probably be iterating by sample, and the popup will appear like this: Changing the Iteration Value There are several ways to change the iteration value. You can use the popup menu in the layout editor window to show the layout for any value in the current group. Alternately you can use the yellow up and down arrows to move to the previous or next values. From the keyboard, you can use the home, page up, page down and end keys to move to the first, previous, next and last iteration values respectively. Setting the Iteration Attribute If you want to edit the iteration attribute, then select the command from the Layout menu called Set Iteration OptionsÖ This will bring up the following dialog box:
The popup menu in this dialog lists all the keywords found in all of the samples of the current group. Select the keyword that you want common to all images in a frame of your layout, but to vary through each of the frames in the stack. |
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