BIOTECHNOLOGY SENSORS: Holograms measure enzyme concentration

Aug. 1, 1996
Two properties that render traditional silver-halide-based holograms unstable as an archival display medium are humidity-dependent color and breakdown of the gelatin.

Two properties that render traditional silver-halide-based holograms unstable as an archival display medium are humidity-dependent color and breakdown of the gelatin. These properties are now being exploited for chemical sensing by a group of researchers from the Institute of Biotechnology at the University of Cambridge (Cambridge, England). The group has been able to develop custom-designed holographic emulsions to produce holograms that collapse predictably in the presence of single specific enzymes. This technique is based on work using more-conventional holographic mirrors, immersed in a test solution, to sense proteaseschemicals present in fecesand water.

The Bragg effect means holograms change color when the fringe spacing is altered, such as might occur when the emulsion, if it is flexible, shrinks or swells. Display holographers have made use of this effect by preswelling emulsions before exposure, then allowing the fringe spacingand the wavelength of the reflected lightto shrink after processing. Thus, multicolor images can be made with a single wavelength of light. This same property, however, also can have a significant unwanted effect on holographic displaysthe color of untreated holograms in a gallery would change depending on the ambient humidity, sometimes rendering them invisible.

Turning this effect to advantage, the Cambridge researchers have demonstrated that the Bragg-reflected color from a holographic mirror can show precisely the concentration of water in a hydrophobic solution. The measurement can be made by eye or with a transmission spectrometer. When the mirror is inserted in xylene for example, the peak reflected wavelength of the hologram changes by as much as 55 nm for water content up to about 325 parts per million.

The relationship between wavelength and concentration is not linear. If the hologram is not heated to "reset" it between uses, the relationship reflects the order in which measurements are taken. Nevertheless, it can be characterized and used to accurately predict water content. Other solvents, including diethyl ether, tetrahydrofuran, butan-1-ol, propan-2-ol, and ethanol also have been tested, with similar results. The group has found that the change in wavelength measures not only the amount of water present, but also the water activity. According to researcher Jeff Blyth, this property may make holographic sensors particularly useful in the food, textile, electronics, and pharmaceutical industries.

Sensing enzymes

In the presence of certain chemicals and enzymes the gelatin used in conventional holographic emulsions degradesanother effect that the researchers have been able to turn to advantage. Trypsin, a protease enzyme that hydrolyses the peptide bonds of proteins, has proved a helpful indicator for certain diseases. Low levels in human feces, for instance, can indicate cystic fibrosis in children. Trypsin also breaks some of the peptide bonds in gelatin so when a hologram is immersed in a liquid containing the enzyme, the gelatin breaks down and the hologram becomes less efficient.

The reduced efficiency occurs partly because the holographic fringe spacing is compromisedinstead of constructively interfering, the reconstruction light tends to scatterand partly because gelatin may be removed altogether, thus eliminating the fringe. The hologram efficiency is, therefore, a function of the amount of trypsin present, and monitoring the intensity of reflected light provides a measurement of the trypsin concentration.

Unfortunately, other proteases can also break down gelatin. To produce a more-discriminating test, therefore, the researchers designed holographic matrices containing enzyme substrates that will collapse only in the presence of specific chemicals. In a report on their work at the Fourth World Congress on Biosensors in Thailand (May 1996), the Cambridge group described a trypsin-specific sensor containing poly(l-lysine) in a poly(vinyl alcohol) matrix.

According to researcher Roger Millington, the group plans to develop the technology in many different ways. Color changes viewed simply by eye could be used for some applications, resulting in a sort of "holographic litmus paper." Hand-held, color-sensitive electronic equipment could also be used to "read" the holographic response more accurately, as could a standard laboratory spectrophotometer. The group has demonstrated all of these techniques in the laboratory.

About the Author

Sunny Bains | Contributing Editor

Sunny Bains is a contributing editor for Laser Focus World and a technical journalist based in London, England.

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