Biosensor FRET-based

Small molecules such as dopamine,ascorbic acid, amino acids, nucleotides, NADH, and glucose have been successfully detected with QD-based biosensors. FRET-based fluorescence microscopy and electrochemistry are the most commonly used methods for small biomolecule detection using QD-based biosensors. ... 

The first FRET-based biosensors employing fluorescent proteins were developed over 10 years ago. These protease sensors consisted of a BFP donor fused to a GFP acceptor by a protease-sensitive linker [44, 119]. BFP and GFP have well separated emission spectra, resulting in little fluorescence bleed-through (Figs. 5.5A and 5.6A). This facilitates data analysis for FRET ratio imaging... 

The requirement of near-UV excitation for BFPs can be a limiting factor. A possible solution would be to use two-photon excitation alternatively a 405 nm diode laser could be used for excitation of BFP. A unique advantage of the BFP/GFP FRET-pair in principle is that it can be used in combination with a red-shifted VFP FRET pair. This enables the simultaneous expression of two different FRET-based biosensors (see Chapter 6). 

One way to reduce the number of independent variables in the FRET-adjusted spectral equation is to use samples with a fixed donor-to-acceptor ratio. Under these conditions, the values of d and a are no longer independent, but rather the concentration of d is now a function of a and vice-versa. This approach is typical for the situation of FRET-based biosensor constructs. These sensors normally are designed to have a donor fluorophore attached to an acceptor by a domain whose structure is altered either as a result of a biological activity (such as proteolysis or phosphorylation), or by its interaction with a specific ligand with which it has high affinity. In general, FRET based biosensors have a stoichiometry of one... 

FRET-based protein biosensors have been developed using CPEs as the lightharvesting donors in conjugation of lock-key recognition. Streptavidin is a... 

Ko, S., and Grant, S. A. (2006). A novel FRET-based optical fiber biosensor for rapid detection of Salmonella typhimurium. Biosens. Bioelectron. 21,1283-1290. 

In the last years, the number of publications related to QD-FRET-based systems has increased continuously. In 2008, a FRET-based nanosensor was developed for the rapid detection of human cardiac troponin I, which is a key factor for the early detection of myocardial infarction.96 In this system, a donor(QD)-labeled protein A is bound to an acceptor-labeled capture antibody and the presence of troponin I antigen generates a conformational change within the structure of the antibody. This results in a change of the distance between the donor and acceptor and, therefore, a shift in energy transfer is observed. A limit of detection of 55 nM of troponin in human plasma and a very short time of analysis (1 minute) were reported using this biosensor. 

NEW METHODS FOR DEVELOPMENT OF FRET-BASED BIOSENSORS WITH EXPANDED DYNAMIC RANGE... 

Development of FRET-Based Biosensors with Expanded Dynamic Range... 

Type 1 Forster (or fluorescent) resonance energy transfer (FRET)-based biosensors Type 2 Bimolecular fluorescence complementation (BiFC)-based biosensors Type 3 Single FP-based biosensors... 

Figure 1.14 Two common examples of (a) FRET-based biosensors. Sensory...

In this FRET-based biosensor (a) one of the FPs is linked to the MRE and the other is linked to the analyte protein. When the sensory protein domain binds with the substrate, the donor and acceptor FPs are brought together, thus increasing the acceptor fluorescence intensity while reducing the donor fluorescence intensity. This strategy is commonly used to tag protein—protein interactions in live cells. 

In this FRET-based biosensor (b) The donor FP and the acceptor FP are fixed to the opposite ends of the MRE. When the analyte binds to the MIEE, the conformation of the sensor protein changes thus placing the donor and acceptor FPs side by side. This increases the FRET efficiency. This is usually used for the detection of glucose, maltose, glutamate, and cyclic nucleotides. 

Burroughs-Tencza, S. FRET-based peptide biosensors for detecting anthrax lethal factor protease and Bacillus anthracis. PCT Int. Appl. WO 2001059149, 2001 Chem. Abstr. 2001, 135, 177260. 

Medintz, I.L., Clapp, A.R., Mattoussi, H., Goldman, E.R., Fisher, B., and Mauro, J.M. (2003) Self-assembled nanoscale biosensors based on quantum dot FRET donors. Nat. Mater. 2, 630-638. 

Pu KY, Liu B (2010) Fluorescence reporting based on FRET between conjugated polyelectrolyte and organic dye for biosensor applications. In Demchenko AP (ed) Advanced fluorescence reporters in chemistry and biology. II. Springer Ser Fluoresc 8 417 -53... 

Fluorescence Reporting Based on FRET Between Conjugated Polyelectrolyte and Organic Dye for Biosensor Applications... 

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