Friday, April 17, 2009

What is MFI?

If you've read any papers with flow cytometry data in it, undoubtedly you've come across the abbreviation, MFI. Generically, people expand this to Mean Fluorescence Intensity, but ironically, you'd rarely use the actual Mean of the population. Basically what the MFI is suppose to measure is the shift in fluorescence intensity of a population of cells. In cases where the entire population stains with different levels of an antibody (like measuring expression level of antigen x), it would be appropriate to report relative MFI values based on some sort of control (unstained, isotype, FMO, etc...) to demonstrate an increase or decrease in expression of this marker (assuming that each sample was stained with saturating amounts of antibody, and all samples were run under the same conditions and instrument settings blah, blah, blah). So, if you wanted to make measurements like this, what statistics would you use? When you analyze your data in software (e.g. FlowJo) you are given options to calculate the Mean, Median, Mode, and Geometric Mean. I've included a link which explains these measures in terms of flow cytometry data pretty well, so i won't bother going through that here. But, I will give you the punchline. When in doubt, use Median Fluorescence Intensity. Mean is pretty much useless, it doesn't work too well on a log scale, and for non-normal distributions, it is easily affected by outliers. I don't mean to be so mean when talking about the mean, but hey, for flow data on a log scale, why bother (sorry, i couldn't resist with the 'mean' pun). If you feel you must use an arithmetic average on a log scale, use Geometric Mean.

12 comments:

  1. Hello
    I am having very basic problem, since i have started using the FACS recentely. i can't find out the way to calculate MFI. i am using CellQuest pro. please help me my email is mazhar@gizi.dote.hu
    thanking you.

    ReplyDelete
    Replies
    1. Ucflow - Flow Cytometry News, Reviews, And Tips.: What Is Mfi? >>>>> Download Now

      >>>>> Download Full

      Ucflow - Flow Cytometry News, Reviews, And Tips.: What Is Mfi? >>>>> Download LINK

      >>>>> Download Now

      Ucflow - Flow Cytometry News, Reviews, And Tips.: What Is Mfi? >>>>> Download Full

      >>>>> Download LINK 86

      Delete
  2. CellQuest is for noobs...
    Use Flowjo.

    ReplyDelete
  3. hi,
    i'm using FlowJo, but my problem is that i do not know how to calculate the MFI, if there's any option with FlowJo to do it and how!
    please if can anybody help me.

    thank you.

    ReplyDelete
  4. Regarding MFI using FlowJo, you can find some documentation here: http://www.flowjo.com/v9/html/statistics.html

    ReplyDelete
  5. This really helps~
    http://flowlab.dfci.harvard.edu/pdf/FlowJo_Tutorial.pdf

    ReplyDelete
  6. In the text above under 'What is MFI?" you say "I've included a link which explains these measures in terms of flow cytometry data pretty well" . I'm probably missing something but I can't see this link - can you please post it in reply to this so I can look at it?

    Thanks ever so much - great blog!

    ReplyDelete
    Replies
    1. hmm. The word 'link' in that sentence is suppose to be a hyperlink to this page on the FlowJo blog: http://flowjo.typepad.com/the_daily_dongle/2007/10/mean-median-mod.html

      Delete
  7. Hi Ryan,
    What would be the best statistic in order to reproduce dose response curve as obtained from Western blots?

    ReplyDelete
    Replies
    1. I think you're looking for a flow cytometry stat that would mimic the results you currently have showing a dose response curve using a Western Blot method. And, if that's your question, then the answer is the Median Fluorescence Intensity. This is a similar relative measurement as one would get from measuring bands in a western.

      Delete
  8. Based on the above information, I gather that if a specific type of cells was exposed to different concentrations of an enzyme to assess the expression of a surface receptor and the MFI obtained was by comparing the experimental samples with untreated control cells, the data should have been expressed as Median Fluorescence Intensity and not as Mean± one SEM?

    Also, is it acceptable to set the MFI of the untreated control cells at 100% and then calculate the MFI of the experimental cells (treated as described above)as a percentage of the 100% in which the MFI of the untreated control was arbitrarily set? Is it further acceptable to get a statistical significance of this MFI percentages?

    ReplyDelete
  9. Ucflow - Flow Cytometry News, Reviews, And Tips.: What Is Mfi? >>>>> Download Now

    >>>>> Download Full

    Ucflow - Flow Cytometry News, Reviews, And Tips.: What Is Mfi? >>>>> Download LINK

    >>>>> Download Now

    Ucflow - Flow Cytometry News, Reviews, And Tips.: What Is Mfi? >>>>> Download Full

    >>>>> Download LINK

    ReplyDelete