Stability of pro- and anti-inflammatory immune biomarkers for human cohort studies

Abstract

BackgroundAlthough discovery research has identified the importance of dozens of pro- and anti-inflammatory immune mediators in the pathogenesis, maintenance, exacerbation and resolution of inflammatory diseases, most human cohort studies have incorporated few or no immunological intermediate phenotypes in their analyses. Significant hindrances have been (1) the limited panel of biomarkers known to be readily detected in healthy human populations and (2) the stability, hence utility, of such biomarkers to repeated analysis.MethodsThe frequency and stability of 14 plasma biomarkers linked to in vivo immune regulation of allergic and autoimmune inflammatory disorders was determined in 140 healthy pediatric and adult participants. The impact of initial and multiple subsequent freeze/thaw cycles on pro-inflammatory (CCL2, CXCL10, IL-18, TNFα, IL-6), anti-inflammatory (IL-10, sTNF-RII, IL-1Ra), acute phase proteins (CRP, PTX3) and other biomarkers (sST2, IL-1RAcP) was subsequently quantified.ResultsMultiple biomarkers capable of providing an innate immune signature of inflammation were readily detected directly ex vivo in healthy individuals. These biomarker levels were unaffected when comparing paired data sets from freshly obtained, never frozen plasma or serum and matched aliquots despite extensive freeze/thaw cycles. Neither age nor sex affected stability. Similarly, no quantitative differences were found following repetitive analysis of inflammatory biomarkers in culture samples obtained following in vitro stimulation with TLR and RLR ligands.ConclusionsA broad panel of in vivo and ex vivo cytokine, chemokine and acute phase protein biomarkers that have been linked to human chronic inflammatory disorders are readily detected in vivo and remain stable for analysis despite multiple freeze thaw cycles. These data provide the foundation and confidence for large scale analyses of panels of inflammatory biomarkers to provide better understanding of immunological mechanisms underlying health versus disease.Electronic supplementary materialThe online version of this article (doi:10.1186/s12967-017-1154-3) contains supplementary material, which is available to authorized users.

Blast traumatic brain injury and serum inflammatory cytokines: a repeated measures case-control study among U.S. military service members

“…Changes in peripheral blood composition due to a post-blast chronic progressive or healing/neuroprotective condition may also affect the detected cytokine levels, as previously reported [58][59][60]. The cytokine stability in serum may be affected by storage conditions [61,62], but according to one study [63], freeze thaw cycles should not significantly affect levels of some of the cytokines measured in our study (TNF-α and IL-6). The results reported here suggest that long-term cytokine profiles may not reflect changes seen immediately following blast results; however, we cannot completely exclude comorbidity with other clinical conditions and we cannot rule out the influence of drug therapy influencing these levels.…”

Pentraxin-3 is not related to disease severity in cirrhosis and hepatocellular carcinoma patients

“…Unfortunately, plasma was not available from this cohort. Measurement of plasma and serum from the same donor yielded higher values in serum [37]. Therefore, PTX3 levels of the HCC and cirrhosis patients could not be compared.…”

Association Between Visfatin and Hepatic Steatosis in the General Population During Long-Term Follow-Up

“…rable covariate in epidemiological studies as, for example, age or gender [26]. On the other hand, Graham et al could not find any differences between the results of several blood samples that were directly evaluated or that underwent up to five freeze-thaw cycles [27]. Moreover protein in blood samples stored at -80 °C has been shown to be stable for years [28].…”