Figure 1.

The Helios mass cytometer shows divergent performance when analyzing EQ beads and cells in the same sample. Replicate aliquots of CD45-barcoded PBMCs were spiked with EQ beads and run in parallel on a CyTOF2 and a Helios mass cytometer. (A) When gating specifically on EQ beads in the sample, the Helios shows higher median intensity and lower CVs for bead-associated channels including 165Ho and 175Lu. (B) In contrast to the beads, expression of CD45–165Ho and CD45–175Lu antibodies on cells in the same sample shows lower median intensity and higher CVs. (C) This higher CV does not reflect a progressive change in marker expression over time; higher variability persists consistently throughout the duration of the acquisition. (D) CD45 expression across defined gated cell subsets, highlighting that the reduced median and increased CV are not population-specific phenomenon. (E) Due to divergent behavior between cells and beads, the reduced signal intensity cannot be corrected by bead-based normalization, which also has no effect on marker CVs. Bars for all graphs represent the mean and S.D. of three aliquots of the same stained PBMC sample acquired independently on three separate days on one CyTOF2 and one Helios mass cytometer.