Background: HLA-specific memory B cells may contribute to the serum HLA antibody pool upon antigen re-exposure. The aim of this pilot study was to investigate the presence of concurrent donor-specific memory B cell-derived HLA antibodies (DSA-M) in renal allograft recipients with pre-transplant donor-specific HLA antibodies (DSA) and its association with occurrence of antibody-mediated rejection (ABMR) using a recently developed method. Methods: Twenty patients with Luminex single antigen bead (SAB) assay-defined DSA but negative complement-dependent cytotoxicity crossmatches were enrolled. Plasma samples and peripheral blood mononuclear cells (PBMC) were collected at 3 timepoints (pre-transplant, month 6, month 12). We analyzed IgG-purified and concentrated culture supernatants from polyclonally activated PBMC using SAB assays and compared HLA antibody profiles with same day plasma results. Results: Plasma SAB analysis revealed 35 DSA in 20 patients pre-transplant. DSA-M were detected in 9/20 (45%) patients and for 10/35 specificities (29%). While median mean fluorescence intensity (MFI) values of DSA with concurrent DSA-M (5877) were higher than those of DSA without DSA-M (1476), 3/6 patients with ABMR and low MFI DSA (<3000) had DSA-M. Overall, pre-transplant DSA/DSA-M pos allograft recipients showed a higher incidence of biopsy-proven (sub)clinical ABMR (p=0.032) and a higher extent (g≥1+ptc≥1) of microvascular inflammation (67% versus 9%, p=0.02). In 17 patients (28 DSA) with post-transplant analyses, persisting DSA post-transplant had more often DSA-M (6/12; 50%) than non-persisting DSA (2/16; 13%). Conclusion: Assessment of DSA-M might be a novel tool to supplement serum HLA antibody analysis for pre-transplant risk stratification in patients with DSA