Syntheses, Photophysical Properties, and Application of Through‐Bond Energy‐Transfer Cassettes for Biotechnology

Abstract

We have designed fluorescent "through-bond energy-transfer cassettes" that can harvest energy of a relatively short wavelength (e.g., 490 nm), and emit it at appreciably longer wavelengths without significant loss of intensity. Probes of this type could be particularly useful in biotechnology for multiplexing experiments in which several different outputs are to be observed from a single excitation source. Cassettes 1-4 were designed, prepared, and studied as model systems to achieve this end. They were synthesized through convergent routes that feature coupling of specially prepared fluorescein- and rhodamine-derived fragments. The four cassettes were shown to emit strongly, with highly efficient energy transfer. Their emission maxima cover a broad range of wavelengths (broader than the four dye cassettes currently used for most high-throughput DNA sequencing), and they exhibit faster energy-transfer rates than a similar through-space energy-transfer cassette. Specifically, energy-transfer rates in these cassettes is around 6-7 ps, in contrast to a similar through-space energy-transfer system shown to have a decay time of around 35 ps. Moreover, the cassettes are considerably more stable to photobleaching than fluorescein, even though they each contain fluorescein-derived donors. This was confirmed by bulk fluorescent measurements, and in single-molecule-detection studies. Modification of a commercial automated DNA-sequencing apparatus to detect the emissions of these four energy-transfer cassettes enabled single-color dye-primer sequencing.

Construction of Energy Transfer Systems within Nanosized Polymer Micelles and their Fluorescence Modulation Properties

“…It can also be observed that with the repeated cycles of UV/visible light irradiation, a slight decrease in the fluorescence intensity in the "On" state (upon UV irradiation) occurs, indicating that some spiropyran moieties might undergo irreversible photodamage. [30][31][32] Moreover, other triblock copolymer micelle samples also exhibit similar spectral behavior as that of the sample VII (PEO 110 -b-PS 85 -b-PSPMA 3 ) in aqueous dispersion, except that at the same micelle concentration, the triblock copolymer micelle samples with longer PSPMA block (spiropyran-containing block) display higher fluorescence intensity at 645 nm upon UV irradiation.…”

From Byproducts to Efficient Fluorophores: A Route to the Synthesis of Fluorubines

“…For instance, the thermal cyclization reaction of 4-bromo-benzaldehyde with substituted 2-amino-hydroxyphenoles into the well-known rhodamines occurs in yields between 5 and 35 % respectively. [17] After anion exchange (Tf À /Cl À into BF 4 À ) of derivative 3 f, we were able to achieve single crystals. The result of the single-crystal X-ray analysis and some bond parameters are depicted in Figure 6.…”

Fluorescence Lifetime Measurements and Biological Imaging

“…21). Alkynyl groups are known to be an effective bridge for energy transfer processes175-179 and have been found to be highly effective in facilitating through-bond electron transfer channels with a low attenuation factor (0.04 per Å) 180,181. Although such constructs have not yet been applied for lifetime imaging, they have some promising characteristics as unique labels.…”