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New Developments in SPR Biosensing

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Recent studies have advanced the potential of surface plasmon resonance biosensing, improving the efficacy of the technique and broadening the field of possible applications.

Gary Boas, News Editor

Surface plasmon resonance (SPR) biosensors have been developed for a variety of applications, from early diagnosis of disease to drug discovery and from water purity analysis to identification of pathogens possibly used for bioterrorism. Offering label-free detection, real-time monitoring and minimal consumption of reagents, the devices are advantageous especially for such applications. Accordingly, researchers are devoting a considerable amount of time and energy to advancing the technology further.

The need for SPR biosensors that can be easily deployed in the field -- to identify dangerous biotoxins, for example -- remains undiminished. Many such portable devices can be cumbersome and unwieldy, however. In a Biosensors & Bioelectronics paper published online Nov. 17, 2007, researchers with CSIRO Manufacturing and Materials Technology (now Materials Science and Engineering) in Clayton, Victoria, Australia, reported an SPR biosensor that is truly handheld -- one that is internally powered and that is operable without connection to an external computer or to any other controlling device.

Compact and off-the-shelf

The device -- which is 8 × 15 cm, weighs 600 g and is powered by a 9-V battery -- is based on a right-angle prism made by Thorlabs Inc. of Newton, N.J. A 3-mW Hitachi laser diode serves as the source of excitation, and a Hamamatsu photodiode array detects the light reflected back from the surface. The array contains 256 photodiodes, each 25 μm wide and 0.5 mm high.

In addition to keeping the device small, the developers sought to keep costs down as much as possible. “There were several design areas where it would have been easier and faster to use specialized components, rather than finding off-the-shelf alternatives or building our own components,” said researcher Bryce N. Feltis. This was particularly true for the fluid handling device, he said. They could have achieved much greater sensitivities with more expensive, custom-built microfluidic devices, but they opted for a not-quite-microfluidic flow cell that they developed themselves.

Researchers have described a truly handheld SPR biosensor. Measuring 8 × 15 cm and requiring no external control, the device can be deployed in the field for applications such as identifying dangerous biotoxins and detecting pathogens in areas with no access to laboratory facilities. Images reprinted with permission of Biosensors & Bioelectronics.

The handheld SPR biosensor could benefit a range of applications. Security and bioterrorism are, of course, important: If a white powder is found in a public space, for example, responders must identify the powder as soon as possible; however, Feltis also noted the need for pathogen detection and typing in remote areas with no available laboratory facilities. “A field-deployable unit that can detect strains of malaria or toxins in water would be of great use in regions where lack of such information often costs many lives.”

Other biosensors are available for such applications, particularly those related to security, but these do not always offer the same advantages. Most are briefcase- to toaster-size -- “certainly portable,” Feltis said, “but not the sort of devices that someone could, for example, carry on their belt.” Furthermore, these biosensors tend to use specialized components and thus can be very expensive.

Array format

The device has some limitations. Currently, it can detect only one substance per chip, in contrast with some of the “luggable” SPR devices, which can detect four or six substances. Also, the user must apply an oil layer between the prism and the sensor disc, an inconvenience when in the field.

The investigators are working to address both of these issues. Their immediate goal is to develop the device as an array, as opposed to its current single-spot format. “We also have some ideas for getting around the oil coupling issue and will be working on improving the user interface and chip insertion system,” Feltis said.

Researchers have established that surface-based methods such as SPR are highly dependent on transport of the analyte to the sensing surface: The biosensing performance can be reduced considerably if the mass transport is too slow. To address this problem, a group with Université des Sciences et Technologies de Lille in Villeneuve d’Ascq, France, developed an SPR biosensing system coupled to a digital microfluidic microstreaming system, reporting it in the Dec. 15, 2007, issue of Biosensors & Bioelectronics.

To address the issue of slow mass transport in SPR biosensing, which can reduce the performance of a device considerably, researchers have developed a digital droplet-based system coupled to a surface acoustic wave microfluidic platform. With this system, the substrate on the top slide enables surface acoustic wave microstreaming, which helps to overcome transport problems during real-time monitoring of interactions. SAW = surface acoustic wave.

Slow mass transport generally is attributed to the dimensions of the microfluidic channels and to the injection flow in the available SPR systems. Therefore, the researchers designed and implemented a digital droplet-based system coupled to a surface acoustic wave microfluidic platform. “We expected that droplet microstreaming, by maintaining some fresh analyte at the surface, could help overcome these transport limitations,” said Vincent Thomy, one of the authors of the study, along with Elisabeth Galopin, Maxime Beaugeois and others.

The microstreaming technique used in the study is made possible by absorption of surface acoustic waves into a droplet at the liquid-substrate interface. The researchers can control the amplitude of the waves while taking into account parameters such as droplet size and surface hysteresis: According to the power generated, they can reach various fluidic velocities and streaming phenomena.

The scientists demonstrated the technique by monitoring streptavidin binding in both surface acoustic wave streaming and static modes. The experimental setup contains a dye laser, a microfluidic platform composed of a gold surface, and a specific surface for surface acoustic wave microstirring (piezoelectric material) and a CCD connected to LabView software to measure, in real time, the SPR minimum deviation along the experimentation. They found that the technique effectively addressed the mass transport problems otherwise inherent in SPR biosensing.

The technique can benefit a range of applications. According to Thomy, it can be seen as complementary to classical flow SPR for investigating interactions where probing of the affinity, association and disassociation kinetic parameters is otherwise limited, noting that the method can be uniquely advantageous for such applications.

“Some studies have used electrothermal streaming in microchannels to enhance biosensing performance,” he explained. “However, we don’t believe this could be adapted to SPR sensing on account of temperature dependency.”

The researchers now are studying a number of additional antibody-antigen pairs -- with various kinetic parameters -- to explore the full range of the technique and its advantages. Soon-to-be-published simulations have shown that surface acoustic wave microstirring can be optimized efficiently according to the types of analytes studied.

Simplifying immobilization

SPR biosensors often use antibody-based assays in which immobilization of the antibody on the sensing film requires a covalent bond between the two, which typically is achieved using a reagent. This step can complicate the immobilization process, however. For this reason, in the Nov. 30, 2007, issue of Biosensors & Bioelectronics, a group with Jilin University in Changchun, China, reported an SPR biosensor system in which the antibody instead is coupled with magnetic beads that can be trapped on the film surface simply by magnetic force.

The magnetic bead-based sensor greatly simplifies the immobilization process and accelerates the detection of analytes, said Hanqi Zhang, the principal investigator of the study. He noted additional advantages: It can be stripped and reused, and it can use the same chemistry as that used to derive dextran-coated sensors.

Bioscientists have used magnetic beads since the mid-1970s, drawing on the beads’ unique properties for immunoassays, MRI and drug delivery, among other applications. More recently, researchers have developed the beads to be used as labels for biosensing because they are generally more stable over time than other types of labels.

Researchers have reported an SPR biosensor system in which the antibody is coupled with magnetic beads, thus allowing immobilization on the surface via magnetic force. This simplifies the immobilization process and accelerates the detection of analytes.

The Jilin University investigators used magnetic microbeads with an SPR biosensor system to detect the heat shock protein Hsp-70, which helps to regulate folding of proteins and to protect cells against stress. The concentration of Hsp-70 changes when a cell is exposed to stress. Therefore, precise, sensitive monitoring of the protein can be crucial for a variety of applications.

The SPR biosensor system they used employed a halogen tungsten lamp and a constant voltage transformer as a light source. After passing through a polarizer and two lenses, the parallel polychromatic light beam from this source traveled through an optical prism with a thin gold film, exciting surface plasmon where the gold film and the analytes met. The output light was transmitted to a spectrophotometer made by Ocean Optics Inc. of Dunedin, Fla., and was detected by a 1024-element CCD linear array detector.

The researchers introduced magnetic microbeads coupled with the antibody into a flow cell with a magnetic pillar positioned above the prism and then introduced a sample solution containing Hsp-70. After a 20-min incubation period, they measured the shift of the resonance wavelength attributable to the antibody-antigen immunoassay and found that the beads were “ideal for use even in a flow cell,” Zhang said.

He noted that the sensitivity of the system increases with an increase in concentration of the conjugate but that too high a concentration can interfere with determination because disassociation of the magnetic particles from the film lasts longer with higher concentrations, leading to wider dips in the SPR spectrum. The investigators are working to address this problem.

Feb 2008
Basic ScienceBiophotonicsbioterrorismdefenseFeaturesindustrialphotodiode arraySensors & DetectorsSurface plasmon resonance (SPR) biosensors

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