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Enhanced Microdialysis Sampling in vitro Recovery of Heparin‐Binding Cytokines
Author(s) -
Newhart William Henry,
Stenken Julie,
Herbaugh Anthony,
Duo Jia
Publication year - 2009
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.23.1_supplement.700.4
Subject(s) - heparin , microdialysis , cytokine , chemistry , monocyte , pharmacology , in vitro , proinflammatory cytokine , chemokine , chromatography , biophysics , immunology , biochemistry , medicine , receptor , inflammation , extracellular , biology
Microdialysis sampling, a well known diffusion‐based separation method, has been used to collect solutes from various tissues in selected organisms. Cytokines have low aqueous diffusion coefficients and can frequently be in low (fM to pM) concentrations. The purpose of this study was to analyze heparin for its ability to increase the RR of selected cytokines. Heparin is a naturally‐occurring glycosaminoglycan and has high binding affinity to many types of cytokines. Two cytokines, monocyte chemoattractant protein‐1 (MCP‐1) and RANTES were chosen since we have confirmed their binding to heparin using surface plasmon resonance (SPR) to be K D = 5.08 ± 1.63 nM and 474 ± 110 nM respectively. For each cytokine, it was important to ensure that heparin did not affect the chemical analysis. Heparin concentrations in the range of 10 µM to 0.001 µM did not interfere with ELISA analysis. In this study, the relative recovery of the cytokine MCP‐1 was shown to increase with heparin concentrations 1 µM (18.34%) and 10 µM (21.20%), but possible interference by heparin of the ELISA analysis was observed. Cytokine RANTES was suspected to cause high dose hook effect with a modified ELISA protocol.