Infinicyt™: Flow Cytometry Software

Merge of FCS files

What does File Merge consist on?

With this function, the user can select several FCS files and merge its information in a single file.
The file resulting from the merger will contain all the events of all the files selected for the merger process. All parameters measured will be merged into a single data file but each parameter is saved separately.

Please see detailed information about file merge process in chapter 4 of the User’s Manual and several illustrative examples in the Merge Tutorial available in the “Support” section of our web site: www.infinicyt.com.

What do I need to know before designing a panel of tubes to merge?

Please be aware of the following, in order to correctly use the File Merge option:

1. The acquisition settings and the sample preparation protocol (surface staining / cytoplasmic staining) have to be the same for each tube to be merged if we want to consider FSC and SSC as common parameters.
2. The appropriate marker panel design is fundamental in order to obtain satisfactory results with the File Merge mode. The design of the necessary antibodies panel must be done taking into account that in each tube of the panel must appear:
   • A series of common parameters that allow to clearly identify the population of interest for our particular study. The more common parameters we use, the more precise the identification of the population at interest will be, e.g. to identify the population of B lymphocytes from a peripheral blood sample FSC / SSC / CD19, at least, could be used as common parameters.
     • It is necessary to include in each tube all the markers that will allow us to identify the population of interest (common parameters), e.g. if our population of interest was the helper/inducer T cells, all the tubes in the panel must have at least CD3 and CD4 markers for their correct identification.
     • The common markers used in the panel must be marked with the same fluorochrome.
   • In fluorescences not occupied by common parameters, the rest of antibodies to be used in our study will be included.

Infinicyt™ includes an internal control system that verifies if the previously mentioned conditions apply. Please see detailed information about merger process controls in chapter 4 of the User’s Manual.

Can I merge FCS files, which include common markers (parameters) stained with different fluorochromes?

Although Merge process has no requirements in the data files to be merged, the software will consider as common parameters the ones containing the same label. Therefore common markers have to be stained with the same fluorochrome (and same label typing) to be recognized by the merger algorithm as common parameters.

How many common parameters do you need to include in each FCS file in order to do a file merge?

The original files must have as many as necessary common parameters that allow the identification of the cell population of interest.

In case we are analyzing a chronic lymphocytic leukaemia we need identify the pathological B-cells so we will have as common parameters the Forward and the Side Scatter characteristics and the CD19 parameters in all the files.

Example: Panel of Chronic Lymphocytic Leukaemia

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