PhotoBiotics Ltd

Thrapston, United Kingdom

PhotoBiotics Ltd

Thrapston, United Kingdom
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Chung P.-H.,King's College London | Levitt J.A.,King's College London | Kuimova M.K.,Imperial College London | Yahioglu G.,Imperial College London | And 2 more authors.
Progress in Biomedical Optics and Imaging - Proceedings of SPIE | Year: 2011

Meso-substituted boron-dipyrromethene (BODIPY-C 12) was used to monitor the viscosity in cells via fluorescence lifetime imaging (FLIM), and time-resolved fluorescence anisotropy measurements. Our results show that mesosubstituted BODIPY-C 12 senses the viscosity in HeLa cells and is insensitive to the surrounding polarity. The relationship between the fluorescence lifetime and the rotational correlation time of the dye in homogeneous solutions agree with the combination of the Förster Hoffmann equation and the Debye-Stokes-Einstein equation. © 2011 SPIE.


Levitt J.A.,King's College London | Kuimova M.K.,Imperial College London | Yahioglu G.,Imperial College London | Yahioglu G.,PhotoBiotics Ltd. | And 3 more authors.
Chinese Optics Letters | Year: 2010

Fluorescence liftime imaging (FLIM) of modified hydrophobic bodipy dyes that act as fluorescent molecular rotors shows that the fluorescence lifetime of these probes is a function of the microviscosity of their environment. Incubating cells with these dyes, we find a punctate and continuous distribution of the dye in cells. The viscosity value obtained in what appears to be endocytotic vesicles in living cells is around 100 times higher than that of water and of cellular cytoplasm. Time-resolved fluorescence anisotropy measurements also yield rotational correlation times consistent with large microviscosity values. In this way, we successfully develop a practical and versatile approach to map the microviscosity in cells based on imaging fluorescent molecular rotors. © 2010 Chinese Optics Letters.


Suhling K.,King's College London | Levitt J.A.,King's College London | Chung P.-H.,King's College London | Kuimova M.K.,Imperial College London | Yahioglu G.,PhotoBiotics Ltd
Journal of Visualized Experiments | Year: 2012

Diffusion is often an important rate-determining step in chemical reactions or biological processes and plays a role in a wide range of intracellular events. Viscosity is one of the key parameters affecting the diffusion of molecules and proteins, and changes in viscosity have been linked to disease and malfunction at the cellular level. While methods to measure the bulk viscosity are well developed, imaging microviscosity remains a challenge. Viscosity maps of microscopic objects, such as single cells, have until recently been hard to obtain. Mapping viscosity with fluorescence techniques is advantageous because, similar to other optical techniques, it is minimally invasive, non-destructive and can be applied to living cells and tissues. Fluorescent molecular rotors exhibit fluorescence lifetimes and quantum yields which are a function of the viscosity of their microenvironment. Intramolecular twisting or rotation leads to non-radiative decay from the excited state back to the ground state. A viscous environment slows this rotation or twisting, restricting access to this non-radiative decay pathway. This leads to an increase in the fluorescence quantum yield and the fluorescence lifetime. Fluorescence Lifetime Imaging (FLIM) of modified hydrophobic BODIPY dyes that act as fluorescent molecular rotors show that the fluorescence lifetime of these probes is a function of the microviscosity of their environment. A logarithmic plot of the fluorescence lifetime versus the solvent viscosity yields a straight line that obeys the Förster Hoffman equation. This plot also serves as a calibration graph to convert fluorescence lifetime into viscosity. Following incubation of living cells with the modified BODIPY fluorescent molecular rotor, a punctate dye distribution is observed in the fluorescence images. The viscosity value obtained in the puncta in live cells is around 100 times higher than that of water and of cellular cytoplasm. Time-resolved fluorescence anisotropy measurements yield rotational correlation times in agreement with these large microviscosity values. Mapping the fluorescence lifetime is independent of the fluorescence intensity, and thus allows the separation of probe concentration and viscosity effects. In summary, we have developed a practical and versatile approach to map the microviscosity in cells based on FLIM of fluorescent molecular rotors. © 2012 Creative Commons Attribution License.


Levitt J.A.,King's College London | Chung P.-H.,King's College London | Kuimova M.K.,Imperial College London | Yahioglu G.,PhotoBiotics Ltd. | And 3 more authors.
ChemPhysChem | Year: 2011

We present polarization-resolved fluorescence measurements of fluorescent molecular rotors 9-(2-carboxy-2-cyanovinyl)julolidine (CCVJ), 9-(2,2-dicyanovinyl)julolidine (DCVJ), and a mesosubstituted boron dipyrromethene (BODIPY-C12). The photophysical properties of these molecules are highly dependent on the viscosity of the surrounding solvent. The relationship between their quantum yields and the viscosity of the surrounding medium is given by an equation first described and presented by Förster and Hoffmann and can be used to determine the microviscosity of the environment around a fluorophore. Herein we evaluate the applicability of molecular rotors as probes of apparent viscosity on a microscopic scale based on their viscosity dependent fluorescence depolarization. We develop a theoretical framework, combining the Förster-Hoffmann equation with the Perrin equation and compare the dynamic ranges and usable working regimes for these dyes in terms of utilising fluorescence anisotropy as a measure of viscosity. We present polarization-resolved fluorescence spectra and steady-state fluorescence anisotropy imaging data for measurements of intracellular viscosity. We find that the dynamic range for fluorescence anisotropy for CCVJ and DCVJ is significantly lower than that of BODIPY-C12 in the viscosity range 0.6<η<600 cP. Moreover, using steady-state anisotropy measurements to probe microviscosity in the low (<3 cP) viscosity regime, the molecular rotors can offer a better dynamic range in anisotropy compared with a rigid dye as a probe of microviscosity, and a higher total working dynamic range in terms of viscosity. © 2011 Wiley-VCH Verlag GmbH & Co. KGaA.


Wu Y.,Imperial College London | Stefl M.,J. Heyrovsky Institute of Physical Chemistry | Stefl M.,Charles University | Olzynska A.,J. Heyrovsky Institute of Physical Chemistry | And 8 more authors.
Physical Chemistry Chemical Physics | Year: 2013

Understanding of cellular regulatory pathways that involve lipid membranes requires the detailed knowledge of their physical state and structure. However, mapping the viscosity and diffusion in the membranes of complex composition is currently a non-trivial technical challenge. We report fluorescence lifetime spectroscopy and imaging (FLIM) of a meso-substituted BODIPY molecular rotor localised in the leaflet of model membranes of various lipid compositions. We prepare large and giant unilamellar vesicles (LUVs and GUVs) containing phosphatidylcholine (PC) lipids and demonstrate that recording the fluorescence lifetime of the rotor allows us to directly detect the viscosity of the membrane leaflet and to monitor the influence of cholesterol on membrane viscosity in binary and ternary lipid mixtures. In phase-separated 1,2-dioleoyl-sn-glycero-3- phosphocholine-cholesterol-sphingomyelin GUVs we visualise individual liquid ordered (Lo) and liquid disordered (Ld) domains using FLIM and assign specific microscopic viscosities to each domain. Our study showcases the power of FLIM with molecular rotors to image microviscosity of heterogeneous microenvironments in complex biological systems, including membrane-localised lipid rafts. © the Owner Societies 2013.


Stamati I.,Imperial College London | Kuimova M.K.,Imperial College London | Lion M.,Imperial College London | Yahioglu G.,Imperial College London | And 3 more authors.
Photochemical and Photobiological Sciences | Year: 2010

Photodynamic Therapy (PDT) is a minimally invasive procedure used for treating a range of neoplastic diseases, which utilises combined action of light and a PDT drug called a photosensitiser. The efficiency of this treatment depends crucially on the properties of the photosensitiser used, namely on its efficient uptake by cells or by the surrounding vasculature, intracellular localisation, minimal dark toxicity and substantial phototoxicity. In this report we compare the spectroscopic properties, cell uptake and in vitro phototoxicity of two novel hydrophilic photosensitisers derived from pyropheophorbide-a (PPa). Both new photosensitisers have the potential to form bioconjugates with antibody fragments for targeted PDT. We find that the photophysical properties of both new photosensitisers are favourable compared to the parent PPa, including enhanced absorption in the red spectral region and substantial singlet oxygen quantum yields. Both molecules show efficient cellular uptake, but display a different intracellular localisation. Both new photosensitisers exhibit no significant dark-toxicity at concentrations of up to 100 μM. The phototoxicity of the two photosensitisers is strikingly different, with one derivative being 13 times more efficient than the parent PPa and another derivative being 18 times less efficient in SKOV3 ovarian cancer cells. We investigate the reasons behind such drastic differences in phototoxicity using confocal fluorescence microscopy and conclude that intracellular localisation is a crucial factor in the photodynamic efficiency of pheophorbide derivatives. These studies highlight the underlying factors behind creating more potent photosensitisers through synthetic manipulation. © 2010 The Royal Society of Chemistry and Owner Societies.


Patent
PHOTOBIOTICS Ltd | Date: 2014-11-20

The invention provides compounds of Formula I: wherein b, D, R^(1), R^(2), G, R^(a )and R^(b )have meanings given in the description, or pharmaceutically-acceptable salts or solvates, or pharmaceutically functional derivatives thereof. The invention further provides process for conjugating the compounds to carrier molecules and uses of such compounds and conjugates in the treatment of disease.

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