Background: Neurofilament phosphoforms (Nf) are principal components of the axoskeleton released during axonal injury. Cerebrospinal fluid (CSF) levels of Nf phosphoforms might be useful surrogate markers for disability in multiple sclerosis (MS), aid in distinguishing clinical subtypes, and provide valuable prognostic information. Method: Thirty four patients with MS were included in a three year follow up study along with 318 controls with other non-inflammatory neurological diseases. CSF levels of two Nf heavy chain (NfH) phosphoforms (NfHSM135, NfHSM134) were quantified at baseline and three year follow up using new ELISA techniques. Levels of NfH phosphoforms, the degree of phosphorylation (NfHSM134:NfHSM135 ratio), and changes in NfH levels between baseline and follow up (ΔNfH) were related to the clinical phenotype (RR or SP/PP), to three clinical scales (Kurtzke's EDSS, ambulation index (AI), and nine hole peg test (9HPT)), and to progression of disability. Results: A significantly higher proportion (59%) of patients with SP/PPMS experienced an increase in NfHSM135 levels between baseline and follow up compared with those with RRMS (14%, pSM134 levels at baseline were higher in patients with SP/PP (11 pg/ml) compared with RR (7 pg/ml, pSM135 levels were higher at follow up in SP/PP (129 pg/ml) compared with levels below assay sensitivity in RR (pSM135 correlated with the EDSS (rs = 0.54, ps = 0.42, ps = 0.59, pSM135 with all clinical scales at follow up suggests that cumulative axonal loss is responsible for sustained disability and that high NfHSM135 levels are a poor prognostic sign.