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In many biological experiments the question that a researcher wants to ask is – ‘do some or all of my cells express a particular protein?’ There are many ways of doing this, which you will be familiar with e.g. Western blotting, immunoprecipitation, microscopic examination of stained cells and even mass spectrometry. Using Flow Cytometry to…
After panel design, and titration of the reagents, the next most important step in flow cytometry is setting the proper voltages on the photomultipler tubes (PMTs). These detectors take the photons of light emitted by the fluorochromes on the cells and convert them to electrons, which ultimately become the voltage current that is digitized and…
If you use a Becton Dickinson (BD) cytometer in your lab, the chances are you are acquiring your data using ‘Diva’ software. Diva software is used to acquire your cytometry data on LSRII, LSRFortessa, CantoII and Aria cell sorters. As well as acquiring your data using Diva software, you can also analyse your data after…
Part of my job in running a core flow cytometry facility is to make sure that the experiments that my users run have been optimised. But that optimisation can be split up into several areas. The first area is experimental planning: What do you want to know? Can you do this by flow cytometry? And…
No matter how we make measurements, there will be variation (a spread of data). Take 100 people and ask them to guess your age and you will get a range of results: some will be too low (excellent!), some too high (not so good!). It is the same with any of our laboratory experiments –…
Once you have been in the field of Flow Cytometry for a bit, the name Howard Shapiro will be familiar to you. However, for those who are new to Cytometry, you might not be aware of Dr Shapiro and his fabulous book ‘Practical Flow Cytometry’. Most Cytometrists have a paper copy of this bible of Flow…
After designing a multicolor flow cytometry panel and securing the necessary cells and reagents, the process of optimization of the panel can begin. The first step in that optimization is titration of your antibodies. In this process, following a standard protocol to be used in the final analysis, you stain a known amount of cells with…
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