Pitfalls in cytokine measurements – Plasma TGF-βI in chronic fatigue syndrome

Background: Serum TGF-βI concentrations are reported to be elevated in chronic fatigue syndrome (CFS). However, measurement of circulating cytokines is a complex procedure and control of pre-analytical procedures is essential. The objective of the current study was to measure circulating TGF-βI concentrations in CFS patients compared to healthy controls, taking into account differences in pre-analytical procedures. Methods: Two cohorts of female CFS patients were included. In both studies patients were asked to bring a healthy, age-matched control. At baseline, TGF-βI levels were measured in plasma and additionally P-selectin, a marker of platelet activity, was determined in a subgroup of participants. Results: 50 patients and 48 controls were included in cohort I, and 90 patients and 29 controls in cohort II. Within the cohorts there were no differences in TGF-βI concentrations. However, between the cohorts there was a large discrepancy, which appeared to be caused by differences in g-force of the centrifuges used. The lower g-force used in cohort II (1361 g) caused more platelet activation, reflected by higher p-selectin concentrations, compared to cohort I (p < 0.0001), which was confirmed in a second independent experiment. There was a correlation between TGF-βI and p-selectin concentrations (r 0.79, p < 0.0001). Conclusion: These results demonstrate that control of pre-analytical procedures is an essential aspect when measuring circulating cytokines. No evidence for enhanced TGF-βI in patients with CFS was found.