Defining the non-sensitized patient: a comparison of HLA antibody screening and identification methods

Luminex Single Antigen beads (SAB) are widely used to identify HLA specificities and assign avoid antigens for kidney transplantation. However in our centers, “non-sensitized” waitlist patients are monitored using less sensitive screening kits. To confirm the lack of HLA sensitization, we evaluated sera by SAB. In this study, we compared several antibody screening and identification kits to uncover test discrepancies and investigate possible false reactions. During initial evaluation and listing, patients were defined as “non-sensitized” when the Class I/II FlowPRA screening beads (One Lambda) were negative. Presumed non-sensitized waitlist patients are screened monthly by the Lifecodes Lifescreen kit (Immunocor) and screened yearly by FlowPRA screening beads. To ensure that no HLA antibodies were missed by this protocol, 116 patient samples were tested by One Lambda Class I/II Labscreen Single Antigen beads (1000 MFI cutoff). When discrepancies between the three assays were identified, sera were tested by the Immunocor ID assay and/or One Lambda Labscreen PRA beads. Additional analyses by surrogate crossmatches are ongoing. Of the 116 samples tested from non-sensitized patients, 22% were positive by the Labscreen Class I SAB and 33% were positive by the Labscreen Class II SAB but negative by the screening kits. The positive specificities spanned all HLA loci, but there were some beads that were positive more than expected. Most notably, the DRB1∗04:04 bead was positive in 10% of cases (12/116) with a range of 1080–2524 MFI, and in three cases the patient’s HLA type was DR4. Other specificities observed three or more times included: A80, B45, B76, Cw15, DR18, DP1, DP11, bead DQA1∗01:04/DQB1∗06:04. In four cases, patients presented with new UNet unacceptable antigens (⩾3000 MFI), and in three cases this led to a positive cPRA (4–17%). Unexpected positive specificities were detected by the Labscreen SAB assay in otherwise non-sensitized kidney waitlist patients. Our data are consistent with other studies that compared multiple test assays to interrogate false reactions. Unexpected SAB reactions should be investigated with additional screening assays and/or surrogate crossmatches. In some cases, common false positive beads can be ignored during test interpretation.