The Cross Stain Index (CSI) value, quantifies the spread of a given dye (i.e. the source; reported in rows in the CSI matrix) into another dye (i.e. the target; reported in columns into the CSI matrix), and thus quantifies the impact of the source on the sensitivity of the target. The CSI matrix is a useful tool for the optimization of multicolor panels.

 

CSI values range between 0 and 1. The higher is the CSI value for a pair of dyes, the higher is the spread of the source dye into the target dye.

 

The CSI Matrix page is automatically created by FCS Express when either the Compensation or the Unmixing Matrix is calculated and depicts a spreadsheet with the CSI values for every pair of dyes included in the compensation/unmixing matrix respectively (Figure 9.62). The CSI automatically has conditional formatting automatically applied when created.

 

 

 

 

 

If errors are encountered when calculating the Cross Staining Index Matrix, the spreadsheet containing CSI values will report the error accordingly. Hovering the mouse over a spreadsheet cell reporting the error will display a hint with additional details on the source of the error. An example a hover over error hint is depicted in Figure 9.63 below. In the example below the error occurred because of a lack of stained events in one of the controls.

 

 

 

More details on the Cross Staining Index can be found in the white paper published at https://welcome.cytekbio.com/evaluate-multicolor-performance .