Note that you can manually type in the boundaries of a gate if you wish; you can specify the gate boundaries in terms of channel number, absolute fluorescence, or percentile within the subset.

Or you can create gates by using a gate tool in the graph window. Gate tools are shown in the floating graph tools window, and include a range tool (for histogram plots), as well as 2D tools such as rectangles, polygons, ellipses, and quadrants. The selection tool (arrow) is used to select gates before moving them, deleting them, or modifying them. Selected gates are indicated with square "handles".

Depending on what you select in the preferences, FlowJo may automatically select a gating tool when you click in a graph window.  If your preference is to have FlowJo automatically select a gating tool, you can also choose to have the polygons or rectangles tool as you default for bivariate displays. Note that you can hold down the option key while clicking, to toggle between the rectangle and polygon gating tool.

The first gate is a one-dimensional gate, consisting of an upper and lower bounds. It can only be drawn on a histogram or CDF plot. Simply point the cursor in the graph window where you wish to have the upper or lower bounds, click on the mouse, and while holding the mouse down, drag to the other extent of the gate. FlowJo draws a horizontal line in the graph to show you the bounds of the gate that you have selected. Later, you can click on the line to move it right or left (changing the bounds of the gate), or up or down (which does not change the gate itself). Alternatively, you can move the upper or lower bound to extend the gate.

The other three types of gates are drawn on any bivariate plot (contour, density, or dot plot). A polygonal gate with any number of vertices is started by clicking within the graph window. Move the cursor to the next vertex; click to generate another point in the polygon. Holding down the shift key will force the edge of the gate to be horizontal or vertical. You may close the polygon either by clicking on the originating point, or by double-clicking on the next-to-last point. (You can cancel the generation of a polygon by hitting the "esc" key on the keyboard at any time). You can press the "delete" key to erase the last vertex you created for any polygon.

To draw a rectangular gate, select the appropriate tool (or option-click in the graph window). Hold the mouse key down as you drag to the opposite corner of the rectangle. Rectangular gates can be computed considerably faster than polygonal gates; you may wish to use them when you analyze large data files.

An ellipse gate can be created by selecting the ellipse tool. Ellipses can only be horizontally or vertically oriented. Click and drag the outline of the enclosing rectangle for the ellipse.

Use the quadrant tool to create four non-overlapping rectangular gates. For more information on creating and using quadrant gates, click here.

Once you have created a gate (except when you use the quadrant tool), FlowJo asks you to name it. FlowJo supplies a default name based on the parameters used to draw the gate; you may type in any name you wish. Note that gate names must be different than any other "sibling" of that gate; i.e., a population cannot have two gates with the same name (see information on naming subpopulations).

You can change a gate by moving it or moving one of its vertices; the gate is automatically recomputed as you change it. You can delete a gate by pressing the delete key when a gate is selected.