The Single Antigen Luminex Bead Assay for the Definition of HLA-Specific Antibodies Revisited: Improved Reactivity by Incubation at 37 Degrees Celsius

Background/Objectives: Sera from patients before and after organ transplantation were tested at two different temperatures, 21 °C and 37 °C. Currently, organs are transported under normothermic conditions (37 °C). This observational pilot study was conducted to define the effect of the incubation at 37 °C, comparing the results to the usual temperature of 21 °C for serum–bead incubation. Methods: We used the Luminex-based assay for the identification and characterization of HLA-specific antibodies. The assays were performed using single antigen beads for HLA class I and HLA class II. A total of 42 sera were assessed and tested, and 38 were analyzed on the Luminex 200 platform at both temperatures. Results: We noted varying outcomes: both an increase and a decrease in mean fluorescence intensity values. A shift from negative to positive values (n = 6) and vice versa (n = 1) was observed. Several sera (n = 4 for HLA class I and n = 5 for HLA class II) exhibited no alterations. In general, we observed an increase in the mean fluorescence intensity values by incubation at 37 °C. The analysis at the bead level revealed a significant deviation (37 °C vs. 21 °C) for the bead carrying HLA-A80 (p = 0.0006) and two HLA-DQ beads, DQA1*05:01-DQB1*02:01 (p = 0.0438) and DQA1*01:03-DQB1*06:03 (p = 0.0438). Conclusions: Mimicking physiological temperature conditions for the testing of HLA-specific antibodies will lead to the better and more accurate interpretation of the results. This method shows potential for use in the delisting strategy for highly sensitized patients as well, thus allowing a better and more reliable option for the patient awaiting a suitable crossmatch-negative organ.