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Note on Virtual Bandpass Filter pipeline step for unmixing

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Unmixing Detector Extraction Automated Pipelines - Virtual band pass filters:

A Virtual band pass pipeline step is created automatically for each unmixing control when an umixing matrix is created, and can be viewed by clicking on the Transformations dialog (Tools tab→Transformations). The virtual band pass steps makes the brightest channel accessible on plots and histograms, even when the User Options are set to hide raw detectors in the parameter lists. This for example, makes possible displaying the brightest channel on an histogram to then identify the negative and the positive population.

Virtual band pass filters pipeline steps created by the unmixing wizard cannot be modified directly from the user. To modify virtual band pass pipeline steps the user needs to make adjustments in Step 5 - Adjust parameters (optional) as highlighted above. Adjustments to the Detector in the AutoUnmixing Controls table will automatically update the Virtual band pass filters in the pipeline steps. The change will be automatically displayed in the Spectral Detector Ranges of the Autogenerated Spectral Detector Extraction Step (Figure 9.56).

Although there is no need for the user to access and review the Virtual band pass filters transformations created by the unmixing wizard, it is useful to know why they are there and how they work.

Each Virtual band pass filters pipeline step is in charge of extracting the brightest channel to be used to define control populations on a control sample. The main options displayed in each Virtual band pass filters pipeline step dialog are:

•A list of spectral parameters. The spectral parameter selected in this section matches the spectral parameter selected by the user in Step 3 of the Unmixing Wizard.

•Spectral Detector Ranges. This is the number of the parameter that has been selected as brightest channel. This value matches the parameter defined in Step 5 of the Unmixing Wizard

•For more details on the additional options displayed, please refer to the Miscellaneous pipeline steps chapter of the User Manual.

(Figure 9.55). The brightest detector for APC has been manually modified in Step #5 of the Unmixing Wizard and the change is automatically displayed in the Virtual bandpass filters pipeline step for that dye.