Selective improvement of tumor necrosis factor capture in a cytokine hemoadsorption device using immobilized anti-tumor necrosis factor.

Abstract

Sepsis is a harmful hyper-inflammatory state characterized by overproduction of cytokines. Removal of these cytokines using an extracorporeal device is a potential therapy for sepsis. We are developing a cytokine adsorption device (CAD) filled with porous polymer beads which efficiently depletes middle-molecular weight cytokines from a circulating solution. However, removal of one of our targeted cytokines, tumor necrosis factor (TNF), has been significantly lower than other smaller cytokines. We addressed this issue by incorporating anti-TNF antibodies on the outer surface of the beads. We demonstrated that covalent immobilization of anti-TNF increases overall TNF capture from 55% (using unmodified beads) to 69%. Passive adsorption increases TNF capture to over 99%. Beads containing adsorbed anti-TNF showed no significant loss in their ability to remove smaller cytokines, as tested using interleukin-6 (IL-6) and interleukin-10 (IL-10). We also detail a novel method for quantifying surface-bound ligand on a solid substrate. This assay enabled us to rapidly test several methods of antibody immobilization and their appropriate controls using dramatically fewer resources. These new adsorbed anti-TNF beads provide an additional level of control over a device which previously was restricted to nonspecific cytokine adsorption. This combined approach will continue to be optimized as more information becomes available about which cytokines play the most important role in sepsis.