Fluorescence and Optical Fiber
In the previous chapter, we learned about spectrophotometric detection for biosensor applications, namely, detection of light absorbance at a certain wavelength. This method works well for a variety of applications, including, obviously, glucose sensing.
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Fluorescence and Optical Fiber
In the previous chapter, we learned about spectrophotometric detection for biosensor applications, namely, detection of light absorbance at a certain wavelength. This method works well for a variety of applications, including, obviously, glucose sensing. Despite its popularity, it has some limitations. The glucose assay utilizes the use of the enzyme glucose oxidase (GOx) that can only detect glucose. To detect other target biomolecules, we need to find an appropriate enzyme that specifically binds to and oxidize them. This is not an easy task, especially when the target is a complicated biomolecule, such as a protein, virus, or even bacterium, where its oxidation is sometimes not at all possible. A more generalized approach is to “label” the target molecule with a specific “dye,” and quantify the concentration of the dye. This dye can be conjugated directly to the target, but a more generalized approach is the use of a secondary bioreceptor that is conjugated with a dye. Figure 9.1 graphically illustrates this concept: Bioreceptors (primary antibodies in Fig. 9.1) are pre-immobilized on a solid surface prior to the assay. A solution that may contain target molecules (antigen to the primary antibody in Fig. 9.1) is added. Upon washing, all unbound molecules are washed away. If dye (fluorescent dye in Fig. 9.1) is conjugated to the target prior to the assay, we simply need to monitor fluorescent emission coming from the solid surface. This monitoring confirms the existence of target molecules and possibly quantifies its concentration through light intensity measurement. However, this direct dye conjugation is impractical, as the other molecules in a test solution may also be conjugated with fluorescent dyes. Therefore, secondary antibodies, identical to the primary antibodies but pre-conjugated with fluorescent dyes, are added to the surface followed by washing. These secondary antibodies will bind to captured target on the surface if it is present. Again, measurement of fluorescent emission from the surface can confirm the existence of target and/or quantifying its concentration.
This chapter is jointly written with Lonnie J. Lucas (Applied Energetics) J.-Y. Yoon, Introduction to Biosensors: From Electric Circuits to Immunosensors, 141 DOI 10.1007/978-1-4419-6022-1_9, # Springer Science+Business Media New York 2013
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9 Fluorescence and Optical Fiber
Fig. 9.1 Use of fluorescent dye in biosensing
Obviously other types of bioreceptors can also be used, including DNA/RNA and enzymes. Two different types of dyes have been used—radioisotope and fluorescent dyes. Radioisotopes are extremely powerful, as a tiny amount of it can exhibit significant magnitude of radioactive decay, leading to extreme sensitivity. Due to the strict regulation on the use of radioisotopes and the difficulty of its use, however, radioisotopes are losing their popularity. Fluorescent dyes have mostly replaced the applications of radioisotope labeling. Due to the recent advancements in optoelectronic componen
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