Liposomes detect metal contaminants and viruses

Liposomes detect metal contaminants and viruses

By Susan English-Seaton

Albuquerque, NM — Scientists at Sandia National Laboratories are working to refine a biochemical technique that uses large molecular aggregates, or liposomes, platformed into in situ sensors to detect metal contaminants in semiconductor process streams. The liposomes can be tailored to react with certain metal ions in solution to detect contaminants in the parts-per-billion range in a matter of seconds. According to scientists, the technique holds promise for a variety of other practical possibilities, including water purity sensors in microchip fabs, metal detectors for environmental cleanup operations, and as rapid laboratory or in-home virus detectors. Sandia is even now working with researcher Deborah Charych at Lawrence Berkeley National Laboratory on recognition groups for viral particles. The team has already created a sol-gel-entrapped liposome that shows high sensitivity to a common flu virus and a working sensor targeted to the cholera toxin, which causes a potentially fatal bacterial disease.

The technique relies on a biochemical process discovered a few years ago at the California Institute of Technology, where researchers were studying ways to purify protein samples by patterning thin-film materials with liposomes. Depending on the chemical roles assigned to reactive molecular groups on the surface of the liposomes, areas on the film could selectively bond with certain proteins and reject others. Former Cal-Tech post-doctoral student Darryl Sasaki, who now leads the research at Sandia, found that when he added copper ions to a liquid solution containing liposomes he had created, the sample`s color emission under a fluorescence spectrophotometer very rapidly changed from green to blue. Researchers surmised that the introduction of charged metal ions prompted the liposomes to scramble their molecular arrangements to incorporate the new ions, thereby altering their fluorescence signal.

“Actually, it just came about serendipitously, our discovery of this sensor. We were trying to build a recognition system to bind proteins. When the protein binds to the surface, it rearranges the molecules in the liposome membrane, which will give us a fluorescent signal. But what happened when we made our first generation of these materials was that we made the liposome vehicle first and found that it gave off a certain fluorescent color. When we added metal into solution so that it could start binding proteins, the color changed very rapidly from green to blue. We didn`t know why, so we started studying,” says Sasaki.

The only problem, Sasaki continues, is that liposomes are not very stable in solution. They can be eaten up by bacteria inside solution and mold and fungus, and will spontaneously fall apart with time. “Liposomes in solution are not practical for a portable sensor. We had to find a way to entrap the liposomes in some solid medium that would also allow us to adhere it onto an optical sensor platform.”

Sol-gel encapsulation proved to be an excellent way to immobilize the liposomes, whereby the liposomes actually reside within cavities in the porous sol-gel matrix but are not chemically attached to the matrix. A class of solid, lightweight, silica-based materials, in contrast to polysaccharide and acrylate gels, sol-gels offer high matrix stability against microbial attack, temperature changes, and physical stress.

Not only did the sol-gel-entrapped liposomes react rapidly to metal ions, their sensitivities were 4 to 50 times greater than those observed for the liposomes in solution — the parts-per-billion range. Sasaki thinks the negatively charged silica surface (a product of sol-gel formation) acts like an ionic sponge increasing concentrations of positively charged metal ions near the sol-gel material`s surface and the odds that a metal ion will encounter and react with a liposome.

In addition, the very few in situ heavy metal sensors available today typically require minutes to hours to respond definitively in the parts-per-billion range. “I think a lot of companies have on-line instrumentation for monitoring wastewater. But typically, they`re somewhat large-scale apparatuses, and they go through a lot of multi-stage processing to finally get the identity or concentration of the metal,” Sasaki says. “Ours would be a good on-line system that should be cheap and quite small to put into a waste stream monitoring system. And it`s very rapid. When the liposomes are in the sol-gel, it takes a little bit longer, because there`s diffusion issues of the metal going through the material. I think we`re down to about 5 minutes now, but we`re still not perfected, so we`ll still have to work on that some more.”

For sensor applications, sol-gels have a number of advantages over other materials, according to Sasaki. They can be applied as a thin film to a variety of surfaces or cast in bulk form into nearly any shape. Optically clear, liposomal color changes are easily reflected. The sol-gel-entrapped liposomes seem impervious to fungal or bacterial attack even after months on a laboratory shelf. Durable, the liposomes remain intact even when the sol-gel structure is damaged. In pharmaceutical applications, says Sasaki, “You might put saliva or a blood sample on a strip of this sol-gel material, and the sensor`s color would change depending on whether it detected a virus the liposomes were looking for.”

A liposome is a spherical bi-layer of a lipid membrane, the inner portion of which is filled with water, the outer, bulk water solution. This gives the liposome its ability to trap substances within itself. “In this case,” says Sasaki, “we would use something like a fluorescent indicator dye. If you put the concentration value high enough, the fluorescence will be quenched, so if you shine light on it, there`ll be no fluorescence from the system. Then we could easily put antibodies on top of the liposome surface. And when the target molecule binds a virus or other pathogen, it will cause the membrane to disrupt, and the entrapped dye will be released, giving off a bright fluorescence. For assays, liposome-based biosensors might even perform a function similar to Rodac plates and bioluminescence in detecting bacteria.


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