The effect of pre-analytical handling on the stability of fractalkine, monocyte chemoattractant protein 1 (MCP1), interleukin 6 and interleukin 8 in samples of human cerebrospinal fluid

J Immunol Methods. 2021 Jul;494:113057. doi: 10.1016/j.jim.2021.113057. Epub 2021 Apr 18.


Cytokine networks in cerebrospinal fluid (CSF) are important to our understanding of several neuroinflammatory diseases. Knowledge about optimal handling of samples is limited but important to minimize bias and reduce costs in CSF biomarker studies. The aim of this study was to examine the effect of storage temperature and time delay from CSF sample collection until freezing on the concentration of 11 different cytokines thought to be associated with chronic pain. CSF samples from 21 individuals undergoing hip or knee arthroplasty under spinal anesthesia were divided between two tubes. One tube was stored and centrifuged (within 30 min) at room temperature, and one tube was stored in ice water and centrifuged (within 30 min) at 4 °C. Each tube was split into six vials that were frozen at -80 °C, 0.5, 1, 2, 3, 4, or 5 h after collection. Cytokines were analyzed using a multiplex panel. A random effect panel data regression was conducted for each biomarker including the variables of storage temperature until freezing and time delay. Four cytokines had detectable levels: Fractalkine, monocyte chemoattractant protein 1(MCP-1), interleukine 6 (IL-6), and interleukine 8 (IL-8). There was no significant effect of storage temperature and time delay on MCP-1, IL-6, or IL-8 concentrations. Fractalkine concentration showed no clear trend. No concentration differences were observed between samples kept in ice water and those at room temperature except at the 3-h time point, and there was no overall significant effect of time delay on fractalkine concentration. We found no clear effect of storage temperature and time delay up to five hours from sample collection until freezing on the CSF concentrations of fractalkine, MCP-1, IL-6, or IL-8.

Keywords: CCL-2 (C–C motif chemokine ligand 2); CX3CL1 (C-X3-C motif chemokine ligand 1); Pre-analytical handling; Storage temperature; Time delay.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Arthroplasty, Replacement, Hip
  • Chemokine CCL2 / cerebrospinal fluid*
  • Chemokine CX3CL1 / cerebrospinal fluid*
  • Freezing
  • Humans
  • Interleukin-6 / cerebrospinal fluid*
  • Interleukin-8 / cerebrospinal fluid*
  • Osteoarthritis / diagnosis*
  • Specimen Handling / methods*
  • Temperature


  • CCL2 protein, human
  • Chemokine CCL2
  • Chemokine CX3CL1
  • Interleukin-6
  • Interleukin-8