Optical and Fluorescent Biosensors

The principle of fluorescent detection systems is that an organic molecule, or in some cases a metal complex, can absorb photons in a particular range and then reemit them at a longer wavelength, illustrated by a Jablonski diagram in Fig. 6.4. For this to be used as a detection method the fluorescent species must have an affinity for a specified target or be attached to a molecule that does. 

The receptor-spacer-reporter motif may appear quite simple yet, as the group of de Silva has shown, can result in some extremely sophisticated responses [3], Through a careful choice of fluorophore reporter, short spacer and nitrogen-containing probe it is possible to construct a series of molecules that can perform Boolean logic in response to external stimuli, illustrated in Fig. 6.5. These mechanisms are currently used in electronic logic components and form the basis of many computational functions. 

The Anslyn group has pioneered supramolecular displacement assays as a distinctive method of analysis. A typical example used a tripodal molecule, or podand, to determine the amount of citrate in drinks and the quality of wine made from Pinot Noir grapes [7], The initial work on citrate utilized a complex between a tris(guanidinium) derivative of 1,3,5-triethylbenzene with a fluorescent dye, car-boxyfluorescein. In the presence of citrate the dye is ejected and replaced with the 

Squaraines are derivatives of 3,4-dihydroxycyclobut-3-ene-l,2-dione, or squaric acid, so called because the central structural motif consists of four carbon atoms linked to form a square [8], Numerous squarylium dyes, such as those in Fig. 6.8, 

The purpose of the rotaxane is to protect the squaraine from nucleophilic attack, likely to occur under physiological conditions, and so enhance the lifetime of the probe in vivo. Indeed, the half-life of the probe is 100 times longer than a non-rotaxane analogue that incorporates a commercial sulfonated carbocyanine dye in place of the squaraine moiety. Furthermore the near infrared spectral nature of the dye means that it gives a strong signal even through centimetres of soft tissue. 

---