Biomarkers in cerebrospinal fluid (CSF) can offer improved diagnostic accuracy for Alzheimer’s disease (AD). The present study investigated whether the glycoprotein and putative tumor suppressor Dkk-3 is secreted into CSF and evaluated its applicability as a diagnostic marker for AD. Using our highly specific IEMA Dkk-3 levels were measured in plasma and/or CSF of patients suffering from depression, mild cognitive impairment (MCI) or AD and compared with healthy subjects. Dkk-3 identity was verified by Western Blot and MALDI-TOF MS/MS. High concentrations of Dkk-3 were detected in CSF compared with plasma (28.2±1.3 vs. 1.22±0.04 nmol/l, respectively). Consistently Dkk-3 expression was demonstrated in neurons of the cortex and epithelial cells of the choroid plexus, the major source of CSF. Significantly increased Dkk-3 levels in plasma and CSF were observed for AD patients compared with healthy subjects but not patients suffering from MCI or depression. In summary our data indicate that elevated Dkk-3 levels are specifically associated with AD and might serve as a potential non-invasive AD biomarker in plasma.