replete with design, application, and sensitivity considerations. In addition to antibodies, a variety of other proteins, ranging from receptors in olfactory neurons to G proteins and bacteriorhodopsin have been explored as potential bioaffinity biosensor elements. In general, these biosensor systems have implementation requirements (size, power, ruggedness) that are not dissimilar to a spectrophotometer, laser, gas chromatography (GC), or mass spectroscopy (MS) based systems. In other words, biosensor systems have instrumentation requirements that may limit their use for standoff detection in the field.

In many cases, modern molecular biology has provided the tools to isolate and modify the genes for individual receptors or signaling proteins to make new biosensor capability. A recent example used computational biology to analyze a binding protein and predict changes to an amino acid sequence that would create new proteins capable of specifically binding to an analyte such as trinitrotoluene (TNT).3 By directed mutagenesis, the DNA encoding the binding protein was synthesized and cloned into bacteria. The fluorescent properties of the protein changed on binding to TNT, providing a direct real-time measure of TNT binding. Furthermore, the binding protein could be linked to a second signal via a signal transduction mechanism to activate a transcriptional fusion in living cells to produce a whole-cell TNT biosensor system. Engineered bacterial strains have previously been developed that couple bioluminescent (lux) or fluorescent (green fluorescent protein [GFP]) reporter proteins to transcriptional activation by chemicals such as TNT, to create biosensor organisms that have been explored for use in explosives detection4 or coupled directly to microluminometer chip technology for whole-cell biosensing.5 Efforts have been made to identify genes that are induced or activated by explosives such as TNT for both mammalian6 and plant systems.7 This raises the potential of engineering animals or plants with lu-

3  

Looger, L. L.; Dwyer, M. A.; Smith, J. J.; Hellinga, H. W. Nature 2003, 423, 185-190.

4  

Burlage, R. S.; Patek, D.R.; Everman, K. R. Method for Detection of Buried Explosives Using a Biosensor. U.S. Patent 5, 972,638, 1999.

5  

Simpson, M. L.; Sayler, G. S.; Applegate, B. M.; Ripp, S. A.; Nivens, D. E.; Paulus, M.J.; Jellison, G. E., Jr. Bioluminescent-bioreporter integrated circuits form novel whole-cell biosensors. Trends Biotech. 1998, 16, 332-338.

6  

Tchounwou, P.B.; Wilson, B. A.; Ishaque, A. B.; Schneider, J. Transcriptional activation of stress genes and cytotoxicity in human liver carcinoma cells (HepG2) exposed to 2,4,6-trinitrotoluene, 2,4-dinitrotoluene, and 2,6-dinitrotoluene. Environmental Toxicology 2001, 16, 209-216.

7  

Jackson, P. Development of Methods That Detect and Monitor Environment Munitions Contaminants Using Plant Sentinels and Molecular Probes. U.S. Department of Energy, 1996, NTIS, AD-A309 583.



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