CyTOF Software v8.0 generates flow cytometry standard (FCS 3.1) files. Events FCS files contain events that meet the event detection criteria specified in the template used to acquire the samples. This is the file used for downstream analysis.
The LMD (linear mode data) file contains all the raw data generated during acquisition. We recommend saving the LMD file in case processing with different event detection parameters is required later.
Additional FCS files containing specific data can also be generated, as well as files intended for service and support purposes.
Select the files that you want to acquire in the Template used for manual tuning or data acquisition.
File |
File Extension |
Description |
Events |
.fcs |
IMPORTANT Select the Events file type for downstream sample analysis. When selected, an FCS file is created for each sample in a batch. Events, Randomize, and Normalize are selected by default, creating a single FCS files with all three processing functions applied. This file name is appended with _Processed. • When Events is selected, the FCS file includes information about the beginning of each cell event (leading push), event duration (in pushes), summed ion signal per event for each channel specified in the settings template, and four additional channels (Center, Offset, Width, and Residual) derived from a Gaussian fit to each event transient. These channels can be used to discriminate between events and nonevents, such as debris and doublets. • When Randomize is selected, data presentation is improved when events are viewed on a bivariate plot. Randomization affects the way events with low signal (1–10 ions) are shown on the plot. • When Normalize is selected, beads added to the sample are used as an internal standard for quantitation of signal coming from cells and to compensate for variation in instrument performance. Variations of signal coming from cell events are corrected by scaling recorded bead signals to the values in a Beads Passport. The default bead type for CyTOF XT™ analysis is EQ6 beads. EQ4 beads are added to samples prepared with a Maxpar Direct Immune Profiling Assay kit. IMPORTANT If you did not add EQ6 beads to your samples, be sure that Add EQ Beads is selected in the Acquisition Protocol. The sample introduction system will add 10% (v/v) EQ6 beads. |
LMD (linear mode data) |
.lmd |
Contains the ion signal for each push from all mass channels. LMD files can be processed to FCS files with different event detection parameters or converted to text using the data processing tools on the Data Processing tab. NOTE CyTOF Software Users must acquire an LMD file for all samples. Templates that do not have the LMD file type selected cannot be applied. |
SPR (signal per reading) |
.fcs |
Contains the summed dual counts of all channels per reading for the duration of sample acquisition. Acquire this file if you want to review potential contaminants and background after acquisition is complete. |
TOF (time-of-flight) |
.txt |
The distribution (spectrum) of ions by time-of-flight accumulated per reading. This file type is not intended for daily acquisition, but for troubleshooting purposes. It may be requested by Fluidigm Service and Support when providing remote assistance.
|
BgPR (background per reading) |
.fcs |
The average dual count per reading of all 135 mass channels during pushes between cell and bead events. Acquire this file if you want to review background signal after acquisition is complete.
|
MBPR (mean beads per reading) |
.fcs |
Contains the number of bead singlets accumulated, the mean dual counts, and CVs calculated per reading for every mass channel specified in Beads Passport. Acquire this file if you want to monitor all bead-positive events during sample acquisition.
|
IPPR (intensity and pulses per reading) |
.txt |
Records the cumulative intensity and number of pulses (ions) per reading detected for all xenon isotopes and 127Ir. This file type is not intended for daily acquisition, but for troubleshooting purposes. It may be requested by Fluidigm Service and Support when providing remote assistance.
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