Purpose: In recent times, but especially since 2001, bioterrorism has been of increasing concern. In addition to the use of biological agents, including Bacillus anthracis (anthrax), there have been numerous hoax white powder "scares." It is imperative to rapidly and accurately identify any suspicious powder as hazardous or hoax. Classical methods for identification typically rely on time-consuming cultivation or highly specific molecular tests which are limited if the agent is unknown. Faster and field portable methods for analysis of suspicious powders are urgently required. Methods: Potential hoax agents, including Bacillus species and household powders, were analyzed using automated microfluidic capillary electrophoresis to determine if protein profiling can distinguish between, and identify, samples. Results: Distinctive protein profiles were produced for Bacillus species, with the presence and/or absence of certain bands, aiding identification. In particular B. anthracis Sterne strain contained a distinctive doublet band above 100 kDa which was not present in any other Bacillus species or hoax agents examined. The majority of powders produced distinctive banding that could enable the identification of the sample while simultaneously ruling out B. anthracis with a high degree of confidence. Conclusions: Results show automated microfluidic capillary electrophoresis can rapidly and reproducibly characterize Bacillus species and hoax agents based on protein profiles without the need for culture. Results were reproducible and there was enhanced resolution and rapidity compared to traditional protein profiling methods. Results show this technique is amenable to field use at a bioterrorism incident, thereby providing essential information to investigators regarding containment and treatment strategies.