Neurofilament (NF) subunits NF-H, NF-M and NF-L are hyperphosphorylated and elevated in Alzheimer disease (AD) brain. We investigated the level and phosphorylation states of NF subunits in lumbar cerebrospinal fluid (CSF) from living patients by bienzyme substrate-recycle enzyme-linked immunosorbent assay. We found: (i), that the levels of phosphorylated NF-H/M (pNF-H/M), non-phosphorylated NF-H/M (npNF-H/M) and NF-L were significantly higher (pNF-H/M, approximately 12-24-fold; npNF-H/M, approximately 3-4-fold) in neurologically healthy aged people than young control individuals; (ii), that in AD, the levels of npNF-H/M, and NF-L were similar to vascular dementia (VaD), and higher than in age-matched controls; and (iii), that the levels of pNF-H/M were significantly higher than in aged controls, non-AD neurological disorders and VaD. Based on these findings, it is suggested that the increased level of total NF proteins in CSF could be used as a marker for brain aging and neurodegenerative disorders in general, and the levels of pNF-H/M as a marker to discriminate AD from normal brain aging and as well as neurological conditions including VaD.