Poster - #VH-02223 - Keywords: Flow Cytometry Instrument Standardization QC

What Your Instrument QC Isn’t Telling You; Implications for Instrument Standardization

Megan McCausland[1], Cameron Paterson[2], Nigel Cumming[2], Alistair Watt[2]
1 IQVIA Laboratories, Marietta, GA, USA 2 IQVIA Laboratories, Livingston, Scotland, UK

ABSTRACT:
Minimizing instrument-driven variability through standardization is crucial for generating reproducible flow cytometry data, essential in both research and clinical settings. Modern cytometry platforms often feature integrated software for instrument setup and QC monitoring to streamline daily startup and qualification. However, the use of hard-dyed QC beads, while efficient and cost-effective, does not accurately reflect the optical performance of fluorochrome-stained cells. Additionally, over-reliance on simple pass/fail outcomes from these automated systems can lead to a false sense of instrument stability and performance.
We present our ongoing efforts to minimize intra- and inter-instrument variability through advanced monitoring, including in-depth longitudinal QC tracking and fluorochrome-specific assessments to confirm both individual fluorochrome and multicolor panel performance. Our fluorochrome-specific assessments identified outliers for PE and BV421 MFI performance within our fleet of 5L (UV, V, B, Y/G, R) Cytek® Aurora spectral cytometers, despite standardized hard-dyed QC bead MFIs. Notably, standardized PE MFI performance for one outlier instrument had been previously verified, suggesting subsequent performance degradation was not detected by the daily QC workflows. Manufacturer intervention and corrective actions improved performance. Furthermore, in-depth longitudinal QC analysis revealed significant UV gain increases across most cytometers in the fleet, despite passing daily QC setup without warnings. The manufacturer’s investigation revealed burned UV optical components and decreased UV laser power after passing through these components, despite normal power measured at the laser during automated daily QC setup, correlating with extensive instrument powered-on time. Replacing the degraded optical components restored UV gains to expected ranges, and beta testing of a laser shutter mechanism to limit repeated burning is currently in progress.
Our findings underscore the need for an in-depth approach to instrument monitoring, including advanced longitudinal QC tracking and fluorochrome-specific assessments to monitor performance over time, applicable to various user scenarios. We urge instrument manufacturers to invest in reagent development and automate fluorochrome-specific particle-based standardization workflows to promote widespread adoption of this more accurate and reliable approach. Additionally, we recommend developing more robust QC pass/fail criteria, including new metrics and standards tailored to fluorochrome-specific assessments.