Confirmation of protective antigen functionality in an electrospun, nanofibrous membrane

The use of Bacillus anthracis as a bioterrorist agent, in 2001, resulted in the death of 5 individuals. The pathogenicity of anthrax is due to the proteolytic effects of one of its components: the enzyme lethal factor (LF). LF´s substrate is located within the cytosol of cells and its cleavage results in a silencing of the interleukin-6 (IL-6) cytokine signaling pathway. The entry of LF into the cytosol is facilitated by a binding protein: protective antigen (PA). Proposed here is a nanofibrous membrane, containing encapsulated PA, that can function as a transdermal anthrax vaccine. This membrane is produced using the voltage-intensive process of electrospinning. To confirm retention of protein biological activity throughout the process, Mono Mac 6 cells, a human monocytic cell line, were treated with electrospun PA after being activated with lipopolysaccharide (LPS) and dosed with LF. A down regulation of IL-6 production, determined through an ELISA, was used as an indicator for PA function. Functionality of electrospun PA was confirmed and the amount of functioning PA deposited on a 4 mm times 4 mm silicon wafer substrate was quantified using the dose-dependent response between PA present and IL-6 down regulated.