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Sensing Trouble

Chemists approach homeland security in a variety of ways
Reservoir



OAK RIDGE NATIONAL LABORATORY
World events of the past two years have brought with them a number of new worries for the average American. The safety of the water supply, the risk of highjackers, and the threat of chemical and biological weapons being used on our shores have moved to the front of the country's collective consciousness. At the 225th national meeting of the American Chemical Society last month in New Orleans, presentations focusing on domestic security concerns were a noticeable addition to the program, with scientists outlining new ways to detect dangerous chemicals and describing novel applications of time-proven techniques.

RECRUITING NATURE

The nation's water reservoirs are a potential target for future terror attacks. In recognition of that, the Bioterrorism Act of 2002 authorized the use of $160 million in federal funds for vulnerability assessments of drinking water systems. In New Orleans, Elias Greenbaum of Oak Ridge National Laboratory outlined a unique approach to screening water reservoirs. Instead of spending a lot of time and energy designing and manufacturing a complicated sensor, his team recruited some of nature's own workers for the task: algae.

"If you look at any river, lake, reservoir, stream, pond--any surface water--even if it looks pretty clean, there will be algae growing in the water, " Greenbaum notes. He should know: his primary focus in the laboratory is studying the organisms' photosynthetic mechanisms. Because algae are sensitive to even the slightest change in their environment, they are suitable for monitoring for potential pathogens. The idea to put algae to work as aquatic sentinels started nearly two years before the 9-11 attacks with a routine call for proposals from the Defense Advanced Research Projects Agency (DARPA), the research and development arm of the Department of Defense. "The connection between using certain physiological parameters of algae and other photosynthetic organisms to try and detect either deliberate or unintentional toxins seemed like a reasonable thing to do," Greenbaum recalls.

During photosynthesis, green plants convert light energy into chemical energy, but like most such processes, this one is not 100 percent efficient. The lost energy leaks back out of the plant as it fluoresces, or emits radiation of a different wavelength than the incident light almost immediately after absorbing it. Different plants show varying patterns of fluorescence over time. The team at ORNL uses these so-called fluorescence induction curves--which are a measure of the algae's health--to monitor for toxins. Real-time, continuous measurements of the fluorescence from algae in surface waters, the researchers have found, can effectively reveal the presence of dangerous chemicals through their effects on the organisms. "In the last couple of years modern fluorometric instrumentation has advanced to the point where one can detect the fluorescence from algae in as-is, untreated water," Greenbaum says. "That is, you don't have to concentrate the sample in order to improve the signal."

So far, laboratory tests have shown that algae are reliable indicators of hazardous substances such as cyanide and the herbicides Paraquat, which destroys mucous membranes, and DCMU, which was often used in conjunction with Agent Orange. Greenbaum and his colleagues also monitored algae from the Clinch River near ORNL, which supplies Oak Ridge with its drinking water, and proved that their technique will work, in principle. Recently the laboratory joined with United Defense to manufacture a prototype system, dubbed AquaSentinelSM, that can be deployed in the field.

SHRINKING BENCHTOP INSTRUMENTATION

The key piece of ORNL's novel detection method may not even be visible to the naked eye. Robert J. Cotter and Ben D. Gardner of Johns Hopkins University described the progress being made toward shrinking what has typically been a laboratory behemoth, the mass spectrometer, in order to use the instrument for homeland defense. Mass spectrometry, in which compounds are ionized and then identified based on how the charged particles behave, is a good candidate for rapidly detecting and identifying biological agents. "Everything has a mass," Cotter points out, "so everything is up for grabs in a mass spectrometer."

But in order to be able to identify potential threats using the machine, scientists need to come up with reliable markers for bacteria or viruses of interest, such as anthrax or smallpox. That's because if potentially deadly bacteria were mixed with something more benign, the resulting spectrum would look different and perhaps allow the harmful agent to go undetected. These biomarkers therefore need to be consistent regardless of the other compounds included in a mixture with the dangerous agents.

To that end, genomic sequencing should help. "Because proteins and peptides are intimately linked to the genome, you can use the genome to predict the masses of all the proteins that could be expressed, regardless of whether they are or not," Cotter explains. "So you can predict all the ones that should be candidates for being expressed, and have a set of known masses for these things. Then all you need to see is a few." For example, a match of five peaks for a particular bacterium's known biomarkers in a complicated spectrum of 40 peaks would constitute a successful identification.

While scientists search for the particular proteins that will expose a bioagent, Cotter and his team are working on another part of the mass-spectrometer-as-field-agent problem: miniaturization. The researchers' latest design houses a cylindrical tube a mere three inches long, a fraction of the length of the tubes in stationary models, which can reach more than a meter. A major hurdle for smaller instruments is resolution: with a shorter flight path for the molecular fragments to traverse, it becomes more difficult to separate very similar masses. By changing the instrument's geometry and altering the standard electric field surrounding the tube so that it is no longer linear, the scientists significantly increased their machine's resolving power. Cotter predicts that further improvements for mini machines that can be used in a field setting will most likely depend on manufacturing constraints that exist for other parts of the machinery, such as vacuum pumps. Although work on the biomarkers is proceeding apace, real-world examples of the technique are probably still at least three years away, he says.

MONITORING FROM THE INSIDE OUT

Of course, an ideal homeland security defense would prevent people from any contact with harmful agents. But if exposure does occur, early detection is often key to a victim's survival. A presentation given by Anthony W. Czarnik, representing Sensors for Medicine and Science, Inc., outlined a method for in vivo monitoring that could one day alert a person to the presence of a toxin within his own body.

The company is developing an implant that can continuously monitor a person's blood sugar levels. The focus is on glucose levels for now, Czarnik says, because of the large market of patients suffering from diabetes. But implants sensitive to other molecules in the bloodstream, such as anthrax, could be developed in the future. "Applications to homeland security follow pretty easily," he remarks.

Roughly the size of a Tic TacR breath mint, the implant is designed to be placed under the skin at the wrist. Inside the tiny device, which would be powered by a specially designed watch worn above the sensor, is a functioning fluorometer that detects glucose molecules and relays the information to a display on the watch face. So far, the company has carried out tests in rabbits and found that the implant can reliably detect changes in blood glucose levels in the animals. An initial meeting with the Food and Drug Administration has been completed, Czarnik reports, and human trials could begin by the end of the year.

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