Abstract
The development of solid-phase assays allows the measurement of anti-HLA antibodies with high sensitivity. Our goal was to address MFI inconsistencies between Luminex instruments as well as tech-to-tech variation when performing Single Antigen Class I & II assays (SAB).
The SAB assay from One Lambda was used for this study. Run-to-run variability was assessed by testing a pool of serums from sensitized patients (PC) on every tray. SAB was performed manually or using the LABXpress Pipettor (LXP). Trays were then run using the same Luminex or in 1 randomly selected instrument out of 4 available. MFI values from informative beads with different MFI intensities were evaluated. We performed Instrument correlation studies by testing the same patient sera on the LXP and reading the same tray on two separate instruments at a time. MFI values for each specificity were compared.
Comparison of PC individual beads MFI values between the manual and LXP methods read on the same Luminex showed an overall intra-run coefficient of variation (CV) of 17% and 10% respectively. An initial assessment of MFI values for PC using 4 different Luminex machines (CV13%) was higher than results obtained when the same Luminex was used (CV 10%). The run-to-run variability was greater for MFI ranges 9–15×103 (CV13%). Subsequent service (alignment & standardization) of all Luminex instruments did not show any improvement on MFI variance (CV 13%). MFI correlation studies revealed a 99% correlation coefficient for all instruments.
Our study revealed that implementation of the LABXpress significantly reduced the MFI variability and validates using this robotic approach for the SAB assay. Even though there was an excellent correlation between Luminex instruments and all QC parameters passed, Luminex controls do not assure instrument MFI comparability. Further efforts must be made to minimize MFI variations allowing the lab to compare MFI values from multiple instruments.
