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Published: Tuesday, February 27, 2007 - 23:00 Research engineer Phil Santangelo works in professor Gang Bao’s cell culture facility in the Wallace H. Coulter Department of Biomedical Engineering at Georgia Tech and Emory University in Atlanta. Photo courtesy of: Phil Santangelo Scientists have used a powerful molecular imaging technique to see inside living cells infected with the most pervasive and potentially fatal childhood respiratory virus known to medicine—respiratory syncytial virus (RSV). “That’s very different from how scientists have studied viruses in the past; they’ve looked at viruses in fixed (or preserved) cells,” he adds. “…Within the first week of studying human RSV in living cells, I learned something new because I was looking at it live.” Confocal microscopy images reveal the three-dimensional structure of bovine RSV viral RNA in living, infected bovine nasal cells using molecular-scale probes called molecular beacons. Photo courtesy of: Phil Santangelo Also, researchers were able to detect virion aggregates in bovine RSV within the first day in culture, Santangelo notes. Typically, veterinarians cannot detect RSV until after five or six days of incubation. This article was originally published in Georgia Tech Research News. Quality Digest does not charge readers for its content. We believe that industry news is important for you to do your job, and Quality Digest supports businesses of all types. However, someone has to pay for this content. And that’s where advertising comes in. Most people consider ads a nuisance, but they do serve a useful function besides allowing media companies to stay afloat. They keep you aware of new products and services relevant to your industry. All ads in Quality Digest apply directly to products and services that most of our readers need. You won’t see automobile or health supplement ads. So please consider turning off your ad blocker for our site. Thanks, For 40 years Quality Digest has been the go-to source for all things quality. Our newsletter, Quality Digest, shares expert commentary and relevant industry resources to assist our readers in their quest for continuous improvement. Our website includes every column and article from the newsletter since May 2009 as well as back issues of Quality Digest magazine to August 1995. We are committed to promoting a view wherein quality is not a niche, but an integral part of every phase of manufacturing and services.Molecular Imaging of a Virus
A door to earlier diagnosis
Scientists and engineers at the Georgia Institute of Technology and the University of Georgia are studying bovine and human RSV with molecular-scale probes, called molecular beacons, that are engineered oligonucleotides—short sequences of RNA or deoxyribonucleic acid (DNA) shaped like a hairpin with a fluorescent dye molecule on one end and a quencher molecule on the other end. They’re designed to fluoresce only when they bind to a complementary target, in this case, RSV genomic RNA.
“For the first time, we were able to visualize an important part of the RSV virus—its genome—in live, infected cells,” says Phil Santangelo, a research engineer in Georgia Tech’s Wallace H. Coulter Department of Biomedical Engineering and at Emory University. “Our molecular beacons attach to the virus and glow inside infected cells as the virus grows, replicates and infects other cells. We can now see that happen in real time in cultures in the lab.
Molecular beacons were originally developed at the Public Health Research Institute in New Jersey in the late 1990s. They were initially used for in vitro assays outside cells. But Santangelo and former Georgia Tech doctoral student Nitin Nitin, now a postdoctoral researcher at Rice University, devised methods for getting the beacons inside the cell without destroying the probe and without changing the cells.
Santangelo will give an invited presentation on his research on April 20 at the Materials Research Society meeting in San Francisco. The research is funded under a National Institutes of Health grant to Professors Shuming Nie and Gang Bao—both in the Department of Biomedical Engineering at Georgia Tech and Emory—to develop new, high-sensitivity live-cell probes. In this study, Santangelo, who works for Bao, collaborated with Amelia Woolums, an associate professor of large animal medicine at the University of Georgia.
They determined that their molecular beacon techniques deliver high-sensitivity and high-specificity results in bovine and human RSV strains. “The RSV genome is interesting in that it is 15,000 nucleotides long, and one of its RNA sequences repeats itself nine times,” Santangelo explains. “So we were able to bind up to nine probes to that sequence, and that helped us achieve very high sensitivity to the virus. In the human virus, in fact, we were able to see a single RSV virion.”
Bovine RSV can be a major problem in cows, which represent a good animal model for human RSV. Calves have RSV symptoms similar to those in human babies and the disease pathology is similar. So, studying bovine RSV yields information about the strain that infects humans, he adds.
Also in this study, researchers used confocal microscopy to view very thin sections of the RSV viral genome in live, infected cells. This technique allowed them to reconstruct the viral RNA aggregates in three dimensions.
“Most pathologists look at thick sections of RSV in formaldehyde, but our 3-D structures are more fluid and amorphous than the solid structures pathologists have observed,” Santangelo says. “The more we know about how RSV really looks, the more we’ll understand about its pathogenesis.”
RSV is the most important cause of respiratory infection in young children worldwide, infecting virtually every child in the first few years of life. Immunity is feeble and fleeting, and repeated infections are the rule. One in every 100 or 200 infected infants requires hospitalization, usually for bronchiolitis. There hasn’t been yet an effective vaccine for RSV, and current antiviral drugs are in their infancy in terms of efficacy, Santangelo notes.
Ultimately, researchers want to conduct in vivo testing, but they must first adapt their molecular beacons technology for that purpose, Santangelo says. “In the nearer term, we hope to use molecular beacons to detect RSV in clinical samples, like with those taken with a nasal swab. We might be able to detect RSV in its first day of incubation and make an early diagnosis,’’ he adds.
The researchers also hope their research will lead to the development of a suite of antiviral drugs for treating RSV and other viruses, including human influenza.
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