title: Comparison of multiplex platforms for cytokine assessments and their potential use for biomarker profiling in multiple sclerosis.
abstract: The levels of pro and anti-inflammatory cytokines can be altered in different autoimmune pathologies, such as multiple sclerosis (MS). It is likely that cytokines in bodily fluids can provide a good reflection of ongoing disease patho-physiology. In this study we aimed to validate multiplex cytokine platforms and evaluate whether these cytokines are differentially expressed in MS.
  
  
  
  Assay validation for simultaneous quantification of IL-1β, IL-6, IL-8 and TNF-α in serum and CSF were performed using both the Luminex-xMAP (Luminex) and Meso Scale Discovery (MSD) platforms. Next, the relation of the pro-inflammatory cytokine 4-plex with disease progression, symptoms and subtypes was studied in paired serum and CSF of MS patients (n=56), and compared with healthy controls (n=203), with the use of the MSD-platform.
  
  
  
  The MSD-platform showed overall better assay characteristics such as, sensitivity, recovery and linearity compared to the Luminex for the 4-plex cytokines in CSF and serum. IL-6, IL-8 and TNF-α (p<0.001) levels were significantly increased in MS serum compared to healthy controls. Moreover, serum IL-1β levels correlated with expanded disability status scale (EDSS) scores (r=-0.34, p<0.05). Additionally, IL-6 and IL-8 CSF levels were both significantly decreased in MS patients compared to non-inflammatory neurological disease controls. Noteworthy, higher IL-8 CSF levels than IL-8 serum levels were observed for MS patients, indicating intrathecal activation of macrophages in MS.
  
  
  
  We have demonstrated that the pro-inflammatory 4-plex kit of the MSD-platform shows better assay characteristics in comparison with Luminex kit for quantification of these cytokines in serum and CSF. Overall, the increased levels of IL-6, IL-8 and TNF-α in serum of MS patients compared to healthy controls, support the use of multiple cytokines for future MS biomarker and disease progression research.
authors.affiliation: Department of Clinical Chemistry, VU University Medical Center, Amsterdam, The Netherlands. Electronic address: a.malekzadeh@vumc.nl.
authors.forename: Arjan
authors.initials: A
authors.lastname: Malekzadeh
authors.affiliation: Department of Clinical Chemistry, VU University Medical Center, Amsterdam, The Netherlands. Electronic address: h.twaalfhoven@vumc.nl.
authors.forename: Harry
authors.initials: H
authors.lastname: Twaalfhoven
authors.affiliation: Department of Internal and Vascular Medicine, VU University Medical Center, Amsterdam, The Netherlands. Electronic address: nienke.wijnstok@gmail.com.
authors.forename: Nienke J
authors.initials: NJ
authors.lastname: Wijnstok
authors.affiliation: Department of Neurology, VU University Medical Center, Amsterdam, The Netherlands. Electronic address: j.killestein@vumc.nl.
authors.forename: Joep
authors.initials: J
authors.lastname: Killestein
authors.affiliation: Department of Clinical Chemistry, VU University Medical Center, Amsterdam, The Netherlands. Electronic address: ma.blankenstein@vumc.nl.
authors.forename: Marinus A
authors.initials: MA
authors.lastname: Blankenstein
authors.affiliation: Department of Clinical Chemistry, VU University Medical Center, Amsterdam, The Netherlands. Electronic address: c.teunissen@vumc.nl.
authors.forename: Charlotte E
authors.initials: CE
authors.lastname: Teunissen
mesh_terms: D015415:Biomarkers / Q000097:blood; D016207:Cytokines / Q000097:blood*; D005260:Female; D006801:Humans; D008297:Male; D009103:Multiple Sclerosis / Q000097:blood*; D011933:Reagent Kits, Diagnostic
chemical_list: D015415:Biomarkers; D016207:Cytokines; D011933:Reagent Kits, Diagnostic
keywords: CSF; Multiple sclerosis; Multiplex immunoassays; Pro-inflammatory cytokines; Serum