Amperometric Detection of Bacillus anthracis Spores: A Portable, Low-Cost Approach to the ELISA

Antibody-based detection assays are generally robust, a desirable characteristic for in-the-field use. However, to quantify the colorimetric or fluorescent signal, these assays require expensive and fragile instruments which are ill-suited to in-the-field use. Lateral flow devices (LFDs) circumvent these barriers to portability but suffer from poor sensitivity and subjective interpretation. Here, an antibody-based method for detecting Bacillus anthracis spores via amperometric signal generation is compared to ELISA and LFDs. This amperometric immunoassay uses antibody conjugated to magnetic beads and glucose oxidase (GOX) along with the electron mediator 2, 6-dichlorophenolindophenol (DCPIP) for production of a measurable current from a 0.4？V bias voltage. With similar sensitivity to ELISA, the assay can be completed in about 75 minutes while being completely powered and operated from a laptop computer. Immunoassay amperometry holds promise for bringing low-cost, quantitative detection of hazardous agents to the field. 1. Introduction Anthrax spores are a dormant viable offspring derived from the Gram-positive, rod-shaped bacterium Bacillus anthracis. They are responsible for 3 well-known disease states: cutaneous, gastrointestinal, and inhalation anthrax, the latter of which is characterized by a high mortality rate [1]. Due to spore stability and high mortality rate, anthrax spores are ranked high among the potential weapons of bioterrorists [2, 3]. Hence, a tremendous interest exists for developing quick, reliable, and field-deployable detection devices. DNA-based assays typically have the best sensitivity and specificity but are not readily suited for in-the-field use due to lack of portability and being highly sensitive to environmental contaminants [4]. Notably, during the investigation of the USA anthrax attack of 2001, a total of 4,639 real-time PCR reactions were performed and all were negative [5]. Positive results were only obtained after culture enrichment when environmental contaminants (dust, debris, and collection buffer surfactants) were eliminated. Alternatively, antibody-based assays are more appealing for in-the-field use due to their environmental robustness and ease of use. The typical ELISA generates quantifiable results and is fairly sensitive (104？cfu/mL) [6] but is too cumbersome and time-consuming to be performed entirely in the field. In addition, expensive and power-demanding instrumentation is needed. Lateral flow devices (LFDs, a.k.a. immunostrips) are a portable and low-cost immunoassay that can be completed in just 15