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Updated: Jun 13, 2026
Standardization of a Cytometric Bead Assay Based on Egg-Yolk Antibodies
Published on: May 19, 2023
A model for continuous quality control incorporating sample-to-sample assessment of optical alignment, fluorescence
Denise Lawrie1, Lindi M Coetzee, Deborah K Glencross
1National Health Laboratory Services and Department of Molecular Medicine and Haematology, Faculty of Health Sciences, University of the Witwatersrand, Parktown, Johannesburg, South Africa.
Background:
Bead count rate (BCR) monitoring successfully identifies pipetting error during single platform CD4 enumeration. Despite rigorous prescribed quality control performed, preliminary data suggested that BCR outliers could also be attributed to occasional failure of flow cytometric volumetric operation. The aim of this report was to use counting beads in a model of continuous quality control (CQC) to monitor overall flow cytometric performance (laser alignment, fluorescence stability and volumetric operation).
Methods:
The proposed CQC model used FlowCheck and IMMUNOTROL blood controls daily. Extended monitoring of fluidics (FPV; beads and sheath only) and sample preparation (SPV; blood, IMMUNOPREP and beads) was done daily on five flow cytometers over five consecutive days prior to testing patient samples. Sample-to-sample CQC included monitoring BCR, selected time/fluorescence histograms (Time vs. Count; Time vs. Fluorescence and Forward Scatter vs. Fluorescence) and full peak coefficient of variation (FPCV) for 2000 samples tested.
Results:
Prescribed quality controls showed Half Peak CV values of <2% (FlowCheck) with Immunotrol within 0.5SD of the target means. Laser stability was confirmed (FPCV values <2%). However, fluidics (volumetric operation) fluctuated as indicated by a 3.2% BCR outlier rate of 2,000 samples tested (minus pipetting error) despite optimal fluidics performance verified at start-up (FPV CV < 3%).
Conclusions:
Sustained laser stability was confirmed with Time vs. Fluorescence histograms, but Time vs. Count histograms were insufficient to detect intermittent volumetric failure. The proposed CQC model, incorporating BCR monitoring with time/fluorescence histograms and FPCV monitoring can identify all volumetric inconsistencies in real-time.
