New toxin detector has applications in fighting bioterrorism.

Department of Energy researchers at the Los Alamos National Laboratory have developed a technique for detecting the toxin that causes cholera. The technique should work equally well in detecting other protein-based toxins, including toxins that might be used in bioterrorism or biowarfare. Because the technique is highly sensitive, produces rapid results, and can pluck specific proteins from a sea of background molecules, it may be especially vital for first responders, who must act quickly to identify and minimize the impact of a terrorist attack.

The technique mimics natural cell-signaling processes, such as those used in the sense of smell; a single recognition event triggers a cascade of signals. To generate a detection signal, the researchers link receptor molecules designed to latch onto the cholera toxin with reporting molecules that fluoresce when struck with a pulse of laser light. The assemblage is lodged in a double-layer membrane formed from lipids, a class of insoluble organic compounds that are important constituents of living cells. Burying the fluorescent probes within the double layer isolates them from proteins and other molecules that are not of interest but that might otherwise affect the fluorescent response. The researchers use two different fluorescent probes, and the double signal minimizes extraneous effects from temperature and other changes. The cholera toxin receptors protrude from the surface of the double-layer membrane, and the fluorescent group resides within the double layer. When a cholera protein comes into contact with the surface, the protruding receptor grabs one of several docking sites on the toxin. (The toxin normally uses these docking sites to establish a toehold when trying to penetrate a cell.) The double layer of lipids allows molecules to migrate around the surface, just as natural cell membranes do. When one receptor nabs a cholera toxin, other receptors aggregate around it and grab onto the remaining toxin docking sites. This aggregation has two effects: First, by binding onto more than one site, the receptors hold the toxin tightly; second, as the receptors crowd together, the fluorescent signal from the reporting molecules changes. The fluorescent response is easily measured in a flow cytometer.

The next step will be to convert this technique from a successful laboratory demonstration to a device suitable for use in the field. The researchers are trying to adapt the technique to devices called "wave guides," which are like fiber-optic sheets. Wave guides are easy to handle and offer multiple channels, so even a small device could contain components that target a number of different toxins.

Such a device would offer advantages over existing detection methods. For example, immunoassays require a variety of reagents and aren't easily used in the field. DNA sequencing can be used only to detect the organism that produces a toxin, not the toxin itself.

In addition, the technique could have great value in the early diagnosis of infections - even before most symptoms appear. Other areas in which the method could be applied include the food industry and pharmaceutical development.

For further information, see the report on this research that appeared in the November 11, 1998, issue of the Journal of the American Chemical Society.