BACKGROUND: This study evaluated the prognostic impact of pretransplant donor-specific anti-human leukocyte antigen antibodies (DSA) detected by single-antigen beads and compared the three generations of crossmatch (XM) tests in kidney transplantation. METHODS: Thirty-seven T-cell complement-dependent cytotoxicity crossmatch (CXM) negative living donor kidney recipients with a retrospectively positive antihuman leukocyte antigen antibody screening assay were included. A single-antigen bead test, a flow cytometry XM, and a Luminex XM (LXM) were retrospectively performed, and the results were correlated with the occurrence of antibody-mediated rejections (AMRs) and graft function. RESULTS: We found that (1) pretransplant DSA against class I (DSA-I), but not against class II, are predictive for AMR, resulting in a sensitivity of 75% and a specificity of 90% at a level of 900 mean fluorescence intensity (MFI); (2) with increasing strength of DSA-I, the sensitivity for AMR is decreasing to 50% and the specificity is increasing to 100% at 5200 MFI; (3) the LXM for class I, but not for class II, provides a higher accuracy than the flow cytometry XM and the B-cell CXM. The specificity of all XMs is increased greatly in combination with DSA-I values more than or equal to 900 MFI. CONCLUSIONS: In sensitized recipients, the best prediction of AMR and consecutively reduced graft function is delivered by DSA-I alone at high strength or by DSA-I at low strength in combination with the LXM or CXM.