In vitro and in vivo affinity microdialysis sampling of cytokines using heparin-immobilized microspheres

Heparin-immobilized microspheres were included in microdialysis sampling perfusion fluids under both in vitro and in vivo conditions to improve the recovery of different cytokines, acidic fibroblast growth factor, vascular endothelial growth factor, monocyte chemoattractant protein-1 (or CCL2), and regulation upon activation normal T cell express sequence (or CCL5). Different strategies to dissociate captured CCL2 and CCL5 from the immobilized heparin were attempted, and both cytokines could be quantitatively eluted from the beads using a phosphate buffer (pH 7.4) containing 25% (v/v) acetonitrile which did not interfere with the subsequent detection of cytokine using an ELISA assay. Using these heparin-immobilized microspheres, a two to fivefold increase of microdialysis relative recovery (RR) was achieved for the four cytokines from a quiescent solution. Enhanced microdialysis RR of CCL2 using the heparin-immobilized microspheres from microdialysis probes implanted into the peritoneal cavity of a rat was performed to test the in vivo application. This work suggests that the heparin-immobilized microspheres provide an alternative affinity agent to the previously used antibody-immobilized microspheres for enhanced microdialysis sampling of cytokines.