Beckman Coulter listmode files acquired on the FC 500, Gallios™, and Navios™ instruments actually contains two datasets inside. The first data set is 10-bit FCS 2.0 data. This is the dataset that you are used to seeing. The second dataset is 20-bit FCS 3.0 data. Users can decide to analyze one or both datasets in FCS Express.
You can choose which dataset to use in the FCS File Options category of FCS Express User Options. To have access to both datasets, you will need to check the box next to Allow Multiple Datasets. Then, when you select the listmode file, you will be given the option to choose which dataset to load.
Leave the box unchecked to always select a single dataset. Enter the data set you wish to use (one (1) for the FCS 2.0 dataset or two (2) for FCS 3.0 dataset) in the Default Dataset to use edit box.
If you have a Beckman Coulter flow cytometer, you probably already know that they have a feature called "baselines offset" that allows you to visually enhance your data display. FCS Express also has this functionality and here is a short video to show you some tips for handling this type of display.
Using Baseline Offsets
The Gallios allows you to define Acquisition and Elapsed Time settings. For more details on the Elapsed Time setting please see our elapsed time knowledgebase article. FCS Express can be used to convert the Time parameter in the Gallios file to real time through the use of the parameter math feature. To convert the Time parameter stored by the Gallios to real time please use the following formulas with parameter math:
|For the FCS 2.0 dataset||For the FCS 3.0 dataset|
If P14 is the time parameter:
New Parameter=P14*(Elapsed time/1024)
If P14 is the time parameter:
New Parameter=P14*(Elapsed time/1048576)
The values for the Elapsed Time can be found in the Gallios Cytometer Control dialog or the Set Times dialog as seen below.
If you want to see what Elapsed Time was used for your data file, you can look at the @PnX keyword in the data file, where n is the time parameter in that data file. The @PnX keyword is only located in the FCS 2.0 dataset of the Gallios file.
Displaying Summit data in FCS Express as it appears in the Summit Software
FCS data in the Summit software is displayed on a 0-256 linear or 0-100,000 logarithmic scale. However, the data is stored in the data file as 32-bit data with a range of 0-4,294,967,295. When the data is viewed in FCS Express, the 32-bit native values are displayed on the axis and used for subsequent calculations such as means and medians.
There are two options for rescaling or displaying the data in FCS Express so that it is in the same range as the Summit Software
- Use the Divide by Axis Labels feature: Using the Divide by Axis Label feature will display the axis as they are seen in Summit but will not change the native raw data or statistical values.
- Use the Parameter Math Feature: Using Parameter Math will create a new parameter based on a scaling factor. The native raw data values will also be scaled accordingly.
|Summit data in FCS Express before applying any axis or parameter math changes.||Summit Data after applying the Divide By Axis labels change. Notice the Axis values have been updated according to the scaling factor but the X and Y Median values have not changed.||Summit Data after performing parameter math. Notice the Axis values have been updated according to the scaling factor and the X and Y Median values have changed to reflect the newly created parameters.|
Gallios, Navios, Summit are registered trademarks of Beckman Coulter Inc.