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RESEARCH PROFILE Multireporter microarray Cy3, and Cy5) and 7 quantum dots Tufts University, where Shepard conA lthough DNA microarrays can assay ducted his doctoral research. It contains (QDs 525, 560, 585, 620, 655, 705, tens of thousands of genes at once, they a bundle of 50,000 optical fibers, each 3 and 800) that together spanned the visitypically compare just two samples, such µm in diameter. A bead coated with sin- ble spectrum. By obtaining images beas cancerous and noncancerous tissue. fore and after labeled targets were hygle-stranded DNA (a probe) is embedThis limitation interested Jason Shepard bridized, he determined the of the Wadsworth Center at reporters that could be excitthe New York State Departed and measured only at parment of Health. “Lung canticular wavelengths and those cer, for example, can be dividthat also emitted into adjaed into many subtypes, such cent reporter wavelengths. as small-cell carcinoma, largeTwo of the quantum dots cell carcinoma, squamous-cell (585 and 705) bled substancarcinoma, and adenocarcinotially into the wavebands of ma,” he says. “So microarrays neighboring reporters and would be better diagnostic therefore were rejected for tools if you could directly further use. compare a patient sample to The next steps were to those four subtypes of the label each PCR product with disease.” one of the 8 remaining fluoMost microarray users serescent reporters, mix the lect a red fluorophore to label samples together, and incua control sample and a green bate the mixture on the array fluorophore to label a test for ~30 min. By exposing the sample. They then excite the Fluorescent spots (3 µm in diameter) on a polychromatic microarray array to eight different wavefluorophores with two differwith (blue) and without (green) hybridized DNA sequences. bands of light and recording ent lasers. Exciting >2 dyes the emissions, Shepard detected all eight ded in the end of each fiber. When a lawould require additional lasers and make beled sample of DNA (a target) hybridizes samples. Thus, each sample contained a scanners bigger and more costly. Laser to the probe and its fluorescent reporter sequence complementary to part of the beams also tend to bleach organic dyes. is excited, the emitted light travels down 16S rRNA gene of B. anthracis, and Shepard has devised a way to make a multireporter microarray without involv- the optical fiber to a camera. “The spots all eight could be detected in a single experiment. on this array are round and perfectly ing lasers. In the April 15 issue of AnaIn the event of an anthrax scare, such uniform, so there is no question about lytical Chemistry (pp 2478–2486), he as the one in 2001, this type of experitheir boundaries,” Shepard says, adding describes a polychromatic optical fiber ment would enable samples from multithat conventional microarrays often microarray that can be reused dozens of ple sources to be tested at one time. For contain irregular spots or “dead” spots times. With fluorophores and quantum example, seven letters mailed from difthat fail to signal. dots in tandem, the instrumentation can To test the polychromatic microarray, ferent places could be analyzed simultareport from eight different samples Shepard coated microspheres with DNA neously along with a standard control. simultaneously. “The marriage between Shepard says his system might be imsequences directed at the 16S ribosomal microarrays and quantum dots seems proved by using a camera that is more RNA gene specific to Bacillus species. very natural,” Shepard says, explaining sensitive to the red area of the spectrum, To obtain the targets, he used DNA that quantum dots give a stronger signal devising algorithms to compare fluorofrom B. anthracis, the bacterium that than organic dyes, fluoresce over a narphore and quantum-dot emissions, and rower waveband, and resist photobleach- causes anthrax. The isolated DNA was optimizing the microscope’s filters. But subjected to asymmetric PCR with biing. Quantum dots are semiconductor having proved the principle, he predicts nanocrystals that fluoresce brightly in re- otinylated primers, which served to amthat polychromatic microarrays will have plify it as well as to incorporate biotin sponse to a beam of light, which in this many applications in basic research, for later attachment of a streptavidin recase comes from an epifluorescence mirapid diagnoses, tailoring of drug theraporter complex. croscope with many filters. pies, and control of bioterrorism. a Shepard tested 10 fluorescent reportThe optical-fiber microarray was —Linda Sage ers one by one: 3 fluorophores (Cy2, devised in the 1990s by David Walt at M A Y 1 , 2 0 0 6 / A N A LY T I C A L C H E M I S T R Y
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