Specimen type validation and establishment of normal cytokine reference intervals in cerebrospinal fluid

Abstract

Cerebrospinal fluid (CSF) is a critical body fluid to examine in attempts to discover potential biomarkers for neuroinflammatory and other disorders of the central nervous system (CNS). Serum and/or plasma cytokine levels have been associated with a variety of inflammatory conditions, and some have been shown to be actionable therapeutic targets. Less is known, however, about cytokine levels in CSF. Serum and plasma cytokine testing is widely available in clinical and research laboratories, but cytokine testing in CSF is extremely limited and if performed, accompanied by a disclaimer that it is an unvalidated specimen type. In this study, we validate CSF as a suitable specimen type and determine normal reference intervals for multiple cytokines as well as a soluble cytokine receptor. CSF was validated as a specimen type for testing using a laboratory developed multiplexed cytokine assay previously validated to measure 13 cytokines/markers in serum and plasma. Performance parameters including specimen dilution, specimen interference, linearity and precision were examined. Reference intervals were established using 197 normal and control CSF specimens by non-parametric quantile-based methods. CSF cytokine analysis demonstrated within and between run precision of <10% and < 20% CV, respectively and linearity of ±15% for all analytes throughout the analytical measurement range of the assay. Reference intervals for the 13 cytokines/markers were established from 197 normal and control CSF specimens (78 Male; mean 44.8 y ± 21.7 SD, 119 Female; mean 42.8 y ± 20.3 SD). Cytokine concentrations in CSF from normal donors and controls were less than the lower limit of quantitation of our assay for 6 of the 13 measured cytokines/markers. The chemokine IL8 demonstrated the highest concentration of all analytes measured. CSF demonstrated acceptable performance as a specimen type in our multiplexed cytokine assay. By validating CSF as a specimen type and establishing normal reference intervals for cytokine concentrations in CSF, their potential as biomarkers for infectious, autoimmune and other inflammatory CNS disorders can be more appropriately investigated.