Fluorescent Biosensor Is Sensitive, Fast

Dr. James P. Smith

By using polymers as fluorescent sensors, a new device has demonstrated detection sensitivity potentially a million times greater than conventional single-molecule-based fluorescence detection. The portable sensor has the potential to detect a variety of pathogens or biological markers almost instantly, making it possible to detect specific drugs or diagnose diseases quickly and easily in homes or clinics.

Researchers at Los Alamos National Laboratory and the University of California at Los Angeles described their instrument in the Oct. 26, 1999, issue of the Proceedings of the National Academy of Sciences. The relatively simple device holds the sample in either a small vial or on the tip of an optical fiber. The fluorimeter consists of a 470-nm light-emitting diode as the pump source, a red path filter and a photodetector. A laptop computer processes the data.

The polymer's fluorescence originates from long, conjugated chains comprising thousands of chromophores. Intermediary molecules turn the fluorescence off and on: A quenching molecule attaches to the polymer to quench the fluorescence; the analyte attaches to the quencher and pulls it away from the polymer, thereby restoring fluorescence. The system is extremely sensitive to small concentrations of quencher and analyte molecules -- a single quencher can turn off the entire chain.

Researchers are developing highly sensitive biological and chemical sensors based on fluorescence quenching in a conjugated polymer. Here, a biosensing polymer solution fluoresces when an analyte protein is added. Courtesy of Los Alamos National Laboratory.

"We have shown we can detect nanomole or subnanomole quantities of analytes," said Duncan W. McBranch, a researcher at Los Alamos. "And I don't think we're pushing the limit on that."

The most critical part of analysis is the bond between the quencher and the analyte, which must be selective and stronger than the quencher-polymer bond. Researchers do this by attaching a ligand to the quencher that is specific to the analyte. The technique has been demonstrated with proteins and with the cholera virus.

Instantaneous analysis

One of the advantages of the new technique is that it requires no washing or separation to prepare the sample, noted David G. Whitten, also at Los Alamos. "The analysis is made in a water system and is very fast; it is essentially an instantaneous analysis."

McBranch and Whitten have worked on this project for more than a year. The next hurdle is to use the technique on more biologically relevant systems, such as blood and other bodily fluids.