Deposition of α-synuclein (αS) aggregates inside brain cells is a pathological hallmark of several neurodegenerative diseases, including Parkinson's disease (PD), and dementia with Lewy bodies (DLB). Recently, extracellular αS was detected in cerebrospinal fluid (CSF) and plasma of humans. We investigated whether CSF influences αS aggregation in vitro using fluorescence spectroscopy with thioflavin S and electron microscopy. We found that CSF obtained from Alzheimer's disease (AD) and DLB patients enhanced the αS fibril formation compared with tauopathy and non-central nervous system disease. Thus, CSF of AD and DLB characterized by aggregation of Aβ or αS might promote fαS formation.