When we backgate - our goal is to identify certain endpoint criteria and apply them to the data set. As example in a whole blood assay, we can use scatter gating to identify which cells might be monocytes. However, there may be some overlap with other cell types making accurate analysis impossible. By adding a fluorescent marker that will identify those monocytes we can place a bitmap on the positive regions of the fluorescence histogram and backgate that to identify the scatter of only those cells that are positive with the fluorescence marker. This is now a backgated population. Backgating can be sequential based on several criteria.