Why should you consider using the free PC program MFI as an alternative to your existing flow cytometry listmode file analysis software? Speed, convenience for certain tasks, being warned about unexpected problems with your data, and inexpensive decentralization of certain analysis tasks are the main reasons. Another is the ability to view time kinetic results with cytometers that don't record time.
These benefits are further explained below. MFI has extensive built-in on- line help, and comes with a detailed tutorial that walks new users through its capabilities, using sample data files included. MFI is in routine use in flow cytometry facilities around the world. MFI supports many common Coulter and BD file formats (including XL, Vantage, FACStation) as well as Winlist, Pro2FCS, and CyCLOPS.
MFI was designed with these goals: to get fluorescence intensity results quickly from large numbers of files, including quick, compact viewing/printing of histograms and dotplots; to provide all necessary corrections to the intensity data; and to alert the user to situations that might otherwise lead to inadvertant misinterpretation of the intensity data. MFI achieves the first goal by allowing unattended batch runs (without viewing graphics) to tabulate intensity results. After MFI was well along, another goal was added: the ability to display time kinetics, even for cytometers that do not record time as an event parameter.
For large numbers of files, MFI gives you intensity results in closer to final form, in a more compact table, with less fuss, and faster. It allows you to scan through histograms or dot plots quickly for large numbers of files, or skip this entirely if you only need intensity results. Because of the many ways in which MFI alerts you to possible problems, you may see things in your data with MFI that you miss with other software.
MFI fosters decentralization of your data analysis. You can make as many copies of MFI as you wish, so there is no reason to confine your analysis to the flow facility computers. With MFI, you can analyze or review your data from your lab or office PC or at home.
MFI's speed and convenience were achieved by sacrificing some of the flexibility offered by alternative software. MFI complements other software, but does not replace it. MFI does not offer quadrant analysis, DNA analysis, 3-D plots, contour plots, density plots, different colors for each gate, complex gating logic, or font control. With the exception of DNA analysis, these features are offered by Joseph Trotter's free program WinMDI.
Getting fluorescence intensity results quickly means printing a compact table without examining histograms or dot plots for all files. Doing this safely means that MFI automatically detects multiple peaks in histograms when they occur, reports drifting of an event cloud relative to its scatter gate, and warns when >10% of events are off-scale. When MFI notices more than one peak in a histogram, it automatically reports the percentages of events and intensities for each peak.
Getting graphics results quickly means that MFI shows you histograms for all parameters on one screen. You can arrive at this screen for the first data file as quickly as pressing the Enter key 5 times after starting the program with new data. Similarly, all possible dot plots (including kinetic dot plots if desired) are displayed with one more press of the Enter key.
When graphics are displayed, MFI takes care to offset the axis lines from the data, so that off-scale data are clearly evident (unlike much other software). Although off-scale end-pileups can be clipped for scaling the Y axis of histograms, a warning is always displayed when this option is enabled.
MFI deals with large numbers of files by allowing the user to tag a subset of the files in the current directory for a given run. The run organization screen also allows assignment of different gates to different files, marking of control files for subtraction, and designation of histogram overlay sets. Up to 16 runs can be configured for a given directory of files.
MFI provides many convenience features. A good example is the ability to add a 1-line sample description or "label" to each list-mode file. These labels make organizing an analysis run and interpreting the resulting printed table much easier. Did you acquire your control sample last? No problem, MFI allows you to rearrange the automated sequence of files from alphanumeric order. The control can be marked for subtraction from the intensities of subsequent files in the run, and its histogram can be overlayed on those for subsequent experimental files. Log-acquired intensities are converted to a linear scale, and intensities can be reported in MESF. Of course MFI remembers all of your configuration choices between uses.