Photophysical processes in single molecule organic fluorescent probes

Abstract

The use of organic fluorescent probes in biochemical and biophysical applications of single molecule spectroscopy and fluorescence microscopy techniques continues to increase. As single molecule measurements become more quantitative and new developments in super-resolution imaging allow researchers to image biological materials with unprecedented resolution, it is becoming increasingly important to understand how the properties of the probes influence the signals measured in these experiments. In this review, we focus on the photochemical and photophysical processes of organic fluorophores that affect the properties of fluorescence emission. This includes photobleaching, quenching, and the formation of non-emissive (dark) states that result in fluorescence blinking in a variety of timescales. These processes, if overlooked, can result in an erroneous interpretation of the data. Understanding their physical origins, on the other hand, allows researchers to design experiments and interpret results so that the maximum amount of information about the system of interest can be extracted from fluorescence signals.

A Three‐Component Assembly Promoted by Boronic Acids Delivers a Modular Fluorophore Platform (BASHY Dyes)

“…Highly fluorescent dyes based, for example, on rhodamine,6, 7 fluorescein,3, 8 oxazine,3 coumarin,3 cyanine9, 10 or boron dipyrromethene (BODIPY)11, 12, 13, 14, 15 chromophores, have been extensively explored along these lines. In particular, BODIPY derivatives (Scheme 1 A) became the focus of different fields due to a combination of remarkable photophysical properties, which included very high quantum yields, narrow absorption and emission bands, and high molar absorption coefficients.…”

Chemo-mechanical pushing of proteins along single-stranded DNA

“…ScPif1 is an SF1B 5′ to 3′ ssDNA translocase that translocates with a rate of 81 ± 8 nt/s on a poly(dT) track at saturating [ATP] [50 mM Tris·HCl, pH 8.3, 100 mM NaCl, 5 mM MgCl 2 , 0.5 mM DTT, and 20% (wt/vol) glycerol, 22°C] (28). The arrival of Pif1 at the 3′ end of the 3′-dT-Cy3-(dT) 140 results in an enhancement of Cy3 fluorescence intensity that can be used to monitor ssDNA translocation (25,28,49) and has been referred to as PIFE (protein-induced fluorescence enhancement) (50,51). Pif1 also binds preferentially to an ss/double-strand (ds)DNA junction and remains bound to the junction while undergoing ATP-dependent ssDNA translocation, resulting in the transient formation of an ssDNA loop (41).…”

Spectroscopic Investigation on the Binding of a Cyanine Dye with Transferrin

“…[2][3][4][5][6] Recently, based on Tf can specifically recognized Tf receptor, Tf has been exploited as a potential strategy of drug to deliver drugs, which are loaded by either covalent conjugation or noncovalent binding, targeting cancer cells with over-expressed Tf receptors on its surface. [7] Meanwhile, cyanine dyes as fluorescent probes for labeling biological systems have attracted considerable attention because of their special properties [8][9][10][11] -wide spectral range, high fluorescence quantum yield, and high molar extinction. And their regulatory structure and a variety of state of aggregation are critical to bind with biomolecules.…”