Two Color RNA Intercalating Probe for Cell Imaging Applications

Abstract

Here we report on a phenanthridine derivative which has a covalently linked fluorescein molecule in order to increase the light absorption and hence fluorescence signal intensity when bound to duplex RNA. Steady-state fluorescence shows that the energy transfer efficiency from the fluorescein to the phenanthridine fluorophore is approximately 77%, which results in the probe being over 5x brighter than other phenanthridine derivatives when bound to RNA. Due to the relatively long lifetime (approximately 20 ns) of the probe, time-resolved fluorescence is used to increase the signal to background ratio in cell growth medium from 7 (steady-state value) to over 40. Moreover, fluorescence images of cells containing the probe show that the fluorescein signal is readily apparent along with that of the intercalated fluorophore, allowing this probe to be used as a dual color probe which simultaneously reports the probes' location and that of RNA.

Phenoxide-Bridged Zinc(II)-Bis(dipicolylamine) Probes for Molecular Imaging of Cell Death

“…Many of the dead and dying cell images with probes 1 and 2 showed punctate regions of high fluorescence intensity in the nucleus that were reminiscent of the nucleolus staining observed with RNA targeting probes. 40–41 Similar cell staining patterns with probe 1 were observed using dead and dying MDA-MB-231 breast cancer cells that had been treated with the drug etoposide (Figure S7). …”

The phenanthridine biguanides efficiently differentiate between dGdC, dAdT and rArU sequences by two independent, sensitive spectroscopic methods

“…4 In addition, it was shown that the chemical modulation of the phenanthridine exocyclic amines is a profitable option to tune the nucleic acid recognition properties of phenanthridinium dyes. 5 A lot of experimental data on ethidium derivatives have been collected over the last three decades 6 including a design of two-colour RNA intercalating probe for cell imaging applications 7 , photoaffinity labelling 8 and antibiotic as well as antitumor activity. [9][10][11][12] Also, recent study demonstrated that double amine-guanidine substitution at the 3 and 8 positions of the ethidium bromide changed the original intercalator into a minor groove binder of DNA.…”

“…25 Therefore, we decided to study an influence of biguanide substituents attached to the 3 and/or 8 position of the phenanthridine ring (compounds 5, 6; Scheme 1) on the binding mechanism of these ligands to DNA/RNA and on their cytotoxic activity. Moreover, to investigate the impact of biguanide-substituents, obtained results are compared with previously studied analogues (Scheme 1) equipped either with inert aliphatic substituents (7,8) or binding-active urea substituent (9). Furthermore, the antiproliferative activity of biguanide-phenanthridine conjugates against tumour cell lines and normal cells was also assayed.…”

Optical Characterization and Lasing in Three-Dimensional Opal-Structures

“…Studies of energy transfer in combinations of pyrene-perylene, two different coumarine dyes, and fluorescein-ethidium bromide, have been carried out with applications in biological imaging and bioanalytical science (Takakusa et al, 2003;Fujiwara et al, 2005;de Almeida et al, 2007;Stevens et al, 2009). Overlap of the fluorescence and absorption was estimated from our experiments as 6.9 × 10 −15 mol −1 cm 6 (Figure 8), and the critical Förster radius was calculated as 3.1 nm.…”