![]() ![]() The principle of this step is to ensure a good and even flow stream during the instrument’s run.Ĭlogging, back-pressure and other instrument-related issues can affect the flow, so eliminating cells that may have been affected by such problems is an important step to cleaning up the data. The details on this hierarchy, including how each fits together sequentially to produce the optimal flow cytometry figure for every experiment, are outlined below… 1. Backgating - to provide visualization of cells in final gate at higher level.This is where Fluorescence Minus One (FMO) controls become critical in defining the populations of interest. Using viability dyes and dump channels further narrow to the cells of interest. Subsetting gating - to rely on expression of markers and what they identify.Forward and side scatter gating - to remove debris and other events of non-interest while preserving cells based on size and or complexity.Pulse geometry gating - to remove doublets from the dataset.Flow stability gating - to capture events once the flow stream has stabilized, eliminating effects of clogging, back-pressure, and other instrument issues.To this end, the following hierarchy was created to help you gate your events correctly… Hopefully, you will objectively choose the right events to display. In other words, you can reuse and refine your gates and plots over and over again without actually losing cells, but you and you alone will determine which events you are displaying. While actual cells will not be lost in trying various gating strategies, data points can be eliminated from your population. 5 Gating Strategies For Publishing Flow Cytometry Data This is also where science becomes an art form. ![]() Thankfully, there are many ways to avoid shaping the results, and instead sifting for the real and actual data that is relevant to the flow cytometry experiment at hand.Ĭommunicating the results of a flow cytometry experiment is where the researcher has the power to make new or subtle findings instantly comprehensible to the audience. Science must be objective, or it is simply an exercise in creative sculpting, which does nothing to move science forward. ![]() The critical difference between sculptor and scientist is that while the sculptor is guided by a creative vision, the researcher is guided by very particular laws of nature and a specific method of working through a biological hypothesis to avoid shaping the results to his or her whims. There is a story inside the data, and it is the job of the researcher to unravel it. When sitting down to perform a new analysis of flow cytometry data, it is much like Michelangelo staring at a piece of marble. “Every block of stone has a statue inside it and it is the task of the sculptor to discover it.” - Michelangelo ![]()
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