There is limited data on the rate of late antibody-mediated rejection (ABMR) in unselected transplant cohorts. Here, we investigated the prevalence and characteristics of ABMR in a large cohort of long-term kidney allograft recipients. Patients were screened in the context of a randomized controlled trial (BORTEJECT study; ClinicalTrials.gov: NCT01873157) designed to investigate the impact of bortezomib on the course of late ABMR. The study (initiation in October 2013) includes a cross-sectional ABMR screening (key inclusion criterion: functioning graft at ≥ 180 days) to identify 44 recipients eligible for inclusion in the intervention trial. Patients were screened for donor-specific antibodies (DSA) applying solid phase technology and DSA+ recipients underwent protocol biopsies. Through November 2014, ABMR screening (after a median of 6.5 years post-transplantation) had been completed for 714 recipients. One hundred one patients (14%) had DSA above a threshold of 1,000 mean fluorescence intensity (MFI). Forty-four of 78 DSA+ recipients (6% of the overall cohort) subjected to biopsy were diagnosed with C4d-positive (n = 17) or -negative (n = 27) ABMR and 40 consented to participate in the intervention trial. DSA+ABMR+ and DSA+ABMR- patients differed significantly with respect to the MFI of the highest level DSA (P < 0.001), whereby ABMR or C4d positivity were moderately predicted by MFI values (area under the receiver operating characteristic curve: 0.75 and 0.84, respectively). In conclusion, the results of this cross-sectional analysis suggest a ≥ 6% prevalence of late ABMR. We demonstrate that the more frequent finding of circulating DSA may not necessarily associate with ABMR diagnosis, especially in patients with low antibody levels.