Structural mode-matching techniques have enabled researchers at the Naval Research Laboratory (NRL; Washington, DC) to develop an evanescent wave immunoassay sensor capable of detecting bacteria, toxins, and environmental pollutants in parts-per-billion concentrations. In field tests at military Superfund sites, sponsored by the Environmental Security Technology Certification Program, the instrument has measured explosives in ground water for site characterization and remediation monitoring. Led by Frances Ligler, head of the Biosensors and Biomaterials Laboratory at NRL, the team has also automated the sensor so that it can operate from an unmanned plane, sending data by radio signal to observers on the ground.
Fiber sensing tip
The instrument essentially consists of an optical fiber attached to a detector/ transmitter unit. For both biological and nonbiological sensing applications, an antibody is chemically immobilized on the 5-cm-long unclad end of the multimode fused silica fiber. When an antigen contacts the sensing tip of the fiber, it is bound by the antibody; at the same time, a fluorescent tag binds to the antigen. Light from a 10-mW diode laser operating at 635 nm passes down the fiber to excite fluorescence from the tag. The fluorophore propagates back up the fiber to a dichroic mirror, which diverts it to a photodiode detector.
In prototypes, mode-mismatch in the interface region between the clad and unclad fiber resulted in significant signal loss, limiting sensitivity. To address this issue, the group tapered the diameter of the unclad sensing tip from 600 to 100 µm in two stages. Beginning at the interface between the clad and unclad regions of the fiber, a steep taper is applied to match the number of propagation modes excited in the fiber to the number of modes sustained by the unclad region at the sensing end. A second, gradual taper converts new propagation modes excited in the fiber into an evanescent wave, thus maintaining uniform illumination.
Adding the taper, says Ligler, increases the sensitivity of the instrument by three to four orders of magnitude; it can detect concentrations of 1 to 10 ng/ml over assay periods of 5 to 10 min. Although the researchers have experimented with imparting the taper during the drawing process, they have achieved best success by etching the fiber with hydrofluoric acid, using a computer-controlled dipping process.
Automated operation
Ligler's group successfully demonstrated a fully automated system for remote air-sampling applications, flying the battery-operated instrument on a small, unmanned plane to detect benign test bacteria released into the atmosphere. To minimize power requirements, the unit incorporates a ram air sampler—air flowing through a collection tube creates a vortex of fluid through which air passes. Bacteria from the air remain in the fluid, which is drawn off and passed to the sensor tip. An EPROM chip governs the fluidics, while the sensor sends data to a ground-based observer by radio signal.
The research, funded by the Defense Advanced Research Projects Agency (DARPA), has applications in bacterial warfare countermeasures, says Ligler. "It gives you a way to find out what's coming before it gets to you. We can detect bacteria, viruses, environmental pollutants, food contaminants, and explosives."
Research International (RI; Woodenville, WA) has licensed the technology and commercialized nonautomated versions of the sensor, producing an instrument that interrogates four fibers simultaneously, to detect four different substances at once. The design also uses a fiber bundle to provide input/output to the unclad tapered fiber sensing tip. To eliminate the fabrication difficulties of tapering by acid etch, the company has successfully incorporated molded plastic fibers into these devices. In collaboration with NRL, RI has also produced a portable version of the sensor featuring quantitative or qualitative evaluation capabilities with LED readout.