Investigating Cytokine Binding Using a Previously Reported TNF-Specific Aptamer

Cytokines are of chief importance in the pathophysiology of sepsis and other systemic inflammatory response syndromes. We are designing and testing an extracorporeal cytokine adsorption device (CAD) that can remove cytokines via adsorption on biocompatible, microporous beads. The goal of this study was to determine whether a previously reported TNF binding DNA aptamer, 5’-GCGGCCGATA AGGTCTTTCC AAGCGAACGA ATTGAACCGC-3’, could be immobilized our hemoadsorption polymer surface to increase the removal rate of TNF. A reservoir consisting of horse serum spiked with a known concentration of TNF was perfused through our CAD packed with aptamer modified or unmodified (control) polymer beads. The binding affinity of the TNF aptamer was characterized using an enzyme-linked oligonucleotide assay (ELONA). As a positive control a well-established DNA aptamer that binds PDGF BB was also subjected to the same ELONA to validate the assay. TNF capture using the CAD showed no TNF removal over four hours for both the aptamer modified and unmodified control beads. Additionally, the results of the ELONA showed no binding of TNF to the reported aptamer; however the PDGF BB aptamer did bind PDGF BB. Based on these results we are able to conclude that the reported TNF specific aptamer does not bind TNF. These results will be of importance to other studies exploring aptamers for specific binding of TNF.