Methodological improvements for measuring eicosanoids and cytokines in exhaled breath condensate.
BACKGROUND
Exhaled breath condensate (EBC) is simple to collect and as such a non-invasive method that has attracted substantial interest in the last few years. However, several methodological concerns have been raised and it has been difficult to reproduce results between different centres. Because of low concentrations of inflammatory markers, potential loss in the sampling system may have great influence. The aim of the present study was to se if evaporation and plastic coating could facilitate detection.


METHODS
Through methodological improvements, we have now made it possible to measure EBC concentrations of eicosanoids and cytokines in our system. Due to absorbance of both fatty acid derivates and proteins to several plastics, the first step is coating of all surfaces with bovine serum albumin and Tween 20. Since several assays are sensitive to these factors, the methodology has to be standardised to avoid false results. Secondly, larger amounts of EBC have to be vacuum-dried, and thereafter resolved in the respective assay buffers. The EBCs have to be concentrated 5-10 times, depending on samples and assay sensitivity.


RESULTS
Due to these improvements we can measure, for example, cysteinyl-leukotrienes, leukotriene B4, prostaglandin E and 8-isoprostane. High sensitivity assays have also made it possible to measure cytokines, for example, interleukin (IL)-1beta, IL-8 and IL-13.


CONCLUSIONS
We are aware of different results from other labs. However, it seems essential to coat and to concentrate the samples in order to achieve reliable and measurable results.
Tufvesson
Ellen
E
Department of Respiratory Medicine and Allergology, University Hospital, 221 85 Lund, Sweden. ellen.tufvesson@med.lu.se
Bjermer
Leif
L
D000328:Adult; D001249:Asthma / Q000175:diagnosis*; D001944:Breath Tests / Q000379:methods; D016207:Cytokines / Q000032:analysis*; D015777:Eicosanoids / Q000032:analysis*; D006801:Humans; D008875:Middle Aged; D011136:Polysorbates; D012710:Serum Albumin, Bovine; D013501:Surface-Active Agents
D016207:Cytokines; D015777:Eicosanoids; D011136:Polysorbates; D013501:Surface-Active Agents; D012710:Serum Albumin, Bovine