– Associate Director- Immunology and Flow, Atrium Health, United States
Aim: A previous study demonstrated that PAK LX (a Luminex bead-based assay that screen for platelet-specific glycoproteins and HLA class I antibodies) reactivity to HLA antibodies may not be an accurate indicator of HLA immunization. The data showed a 37% discrepancy with single antigen Luminex based assay (SAG) in a small cohort from a single center. Consequently, relying on PAK LX as a standalone assay for HLA class I antibody screening may be risky. The current study expands the cohort size to further investigate this phenomenon and identify the trends influencing this observed discrepancy.
Methods: A cohort of 137 individuals from two American National Red Cross Histocompatibility laboratories in both Charlotte NC and Portland OR, underwent screening using both PAK LX (Werfen, Inc) and SAG (Thermofisher, Inc) methods. Demographic data, antibody positivity rates, discordance rates, mean fluorescence intensity (MFI) values, and calculated percent reactive antibodies (cPRA) were obtained.
Results: The study population comprised 51% female and 49% male individuals, with a majority being Caucasian (52%) and 34% diagnosed with leukemia. Seventy-three percent tested positive for HLA class I antibodies by SAG, while 36% showed discordant results between PAK LX and SAG. Most discordant cases were false negatives (98%) from PAK LX, with two cases of false positives. The mean cPRA of discordant cases was 29% (range 0.02-96%) (figure 1a). Class I ABC antibodies showed a respectively a mean of MFI 4500 (range 1,000-17,000), 4079 (range 1,100-18,000), and 3758 (range 1,200 – 19,000) (figure 1b). The MFI of class I antibodies among discrepant cases was mainly for specificities with MFI below 6000 with a broad range from 2000 to 19,000 (figure 2 A & B).
Conclusion: Missing HLA class I antibody specificity is a significant risk factor for platelet transfusion refractoriness, potentially leading to ineffective treatment and an increased risk of bleeding. Our current study confirmed previous observation that PAK LX HLA class I bead should not be used as a stand-alone method for detecting HLA class I antibody. It also gave some insight of the nature of HLA class I specificities missed by PAK LX.
