Figure 7. An example of gating strategy used to eliminate aberrant binding events due to dead cells that may change tetramer analysis.

This figure shows an example of the utility of using amine reactive dyes to eliminate erroneous frequencies due to non-specific binding of dead cells. In this example, CMV antigen specific CD8+ T cells are measured in a staining panel containing a specific CMV tetramer reagent. The first row shows a traditional gating strategy. The final gate indicates that 1.57% of the CD8+ T cells are tetramer positive. However, when including the viability gate (second row) the CD8+ tetramer + T cells were determined to be at a lower frequency (0.62%) than reported using traditional gating. Overlaying the total CD8+ tetramer + T cells from the first row onto the viability gate (bottom row) shows that the majority of the cells are in fact ViViD + and hence should not be included in the final analysis.