Microdialysis recovery of the lipophilic analytes prostaglandin B2, leukotriene B4 and C4 was studied in vitro. Relative recovery (RR) through different commercially-available microdialysis probes for prostaglandin B2 and leukotrienes was examined using different flow rates. The enhancing effect at different concentrations of binding agents such as alpha, beta, gamma-cyclodextrins (alpha, beta, gamma-CD) on the microdialysis RR for different eicosanoids was evaluated. Small organic molecules such as ethanol, propylene glycol and dimethyl sulfoxide (DMSO) were studied in terms of their effect on enhancing RR. Inclusion of arachidonic acid in either the perfusion fluid or the sample medium caused the microdialysis RR for these hydrophobic analytes to be increased.