LAUSR

BACKGROUND 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. METHODS Assay validation for simultaneous quantification of IL‐1&bgr;, IL‐6, IL‐8 and TNF‐&agr; 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. RESULTS 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‐&agr; (p < 0.001) levels were significantly increased in MS serum compared to healthy controls. Moreover, serum IL‐1&bgr; 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. CONCLUSION 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‐&agr; in serum of MS patients compared to healthy controls, support the use of multiple cytokines for future MS biomarker and disease progression research. HIGHLIGHTSAssessment of multiplex immunoassays for IL‐1&bgr;, IL‐6, IL‐8 and TNF‐&agr; quantification.MSD platform overall showed better assay characteristics for IL‐6, IL‐8 and TNF‐&agr;.IL‐1&bgr;, IL‐6, IL‐8 serum levels were higher in MS patients compared to controls.IL‐6 and TNF‐&agr; CSF levels are lower in RRMS compared to NINDC.