Recently we used single staining to make a compensation matrix. But, for each single staining we saw that 13 to 25 percent of our negative population had been shown below Zero. In this situation MFI would be negative after calculation I think. Am I right? what can I do? Can I use flowjo software to correct it? Or sth else?
Have a look for such rules: https://www.med.unc.edu/flowcytometry/files/2018/06/compensation.pdf and https://depts.washington.edu/cfar/sites/default/files/uploads/core-program/user213/Slides-%20Flow%20Cytometry%20Advanced%20Topics-%2001.30.2020-%20Part%201.pdf
I'm not sure what you are asking. Are you calculating the MFI for compensation? Why? Compensation is a setting on the machine itself (or if you back compensate during analysis, it's just a way to adjust data issues in your samples), but this is only to give you a clear distinction between positive and negative populations so the machine can tell what is consider positive within any given channel, and then to adjust for spillover.
Also MFI is the mean of the population, and if only 20% is
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