Multiphoton Microscopy in the Biomedical Sciences X, 2010
ABSTRACT We report on the construction of a highly flexible system for advanced biological imagin... more ABSTRACT We report on the construction of a highly flexible system for advanced biological imaging, where all the following imaging techniques are integrated into the same microscope: Coherent anti-Stokes Raman scattering (CARS), two photon excitation fluorescence (TPEF), second harmonic generation (SGH), sum frequency generation (SFG), fluorescence lifetime imaging (FLIM) and differential interference contrast (DIC). The system employs a Nd:YVO4 laser as pump (7 ps, 1064 nm), and two tunable OPOs (6 ps, 700 - 1000 nm). Our microscope comprises a heater stage and perfusion cell for imaging of live cells, and features an atomic force microscope (AFM) which enables optical imaging at 10 nm resolution. Multimodal imaging of breast cancer cells and tissue will be demonstrated as well as imaging of anticancer drugs in living cells.
ABSTRACT We have used a versatile and powerful microscope[1] for multi-modal biomedical imaging o... more ABSTRACT We have used a versatile and powerful microscope[1] for multi-modal biomedical imaging on which we combine Coherent Anti-Stokes Raman Scattering (CARS) with Two Photon Excitation Fluorescence (TPEF) using a Nd: YVO4 pump laser. We acquired 2PEF, CARS, and phase contrast images of Multilamellar Vesicles (MLVs) and Giant Unilamellar Vesicles (GUVs), as well as Raman spectra of the constituent lipids. A wide range of peptides are harmful to cells by altering the structure of the biological membranes. This effect depends on the composition of the membrane and the chemical structure of the peptide. The peptide we studied is the beta amyloid Aβ which is a major component of the amyloid plaques deposited on neuronal membranes of Alzheimer's disease (AD) patients. AD is neurodegenerative disorder in which the hallmark symptoms include cognitive decline and dementia[2] and is characterized by the formation of extracellular amyloid fibrils on the neuronal membranes of the brain. Many questions still remain unanswered concerning the destabilization of cellular ionic homeostasis due to pores formed during the interactions of lipid membranes with peptides. In this project, biomimics of cell membranes are used. The structures that best mimic the plasma membranes are MLVs or GUVs. These vesicles are formed using the gentle hydration technique[3] or the electroformation technique[4] respectively and are composed of phospholipids such as DOPC, DPPC, D62PPC and their binary mixtures. The MLVs and GUVs imaging by CARS and TPEF microscopy not only permits the direct imaging of the leakage phenomenon caused by the toxic peptide (Aβ) on the lipid bilayer, but also records simultaneously the lateral structure of the bilayer and peptide distribution in the plane across the membrane.
We report on the construction of a highly flexible system for advanced biological imaging, where ... more We report on the construction of a highly flexible system for advanced biological imaging, where all the following imaging techniques are integrated into the same microscope: Coherent anti-Stokes Raman scattering (CARS), two photon excitation fluorescence (TPEF), second harmonic generation (SGH), sum frequency generation (SFG), fluorescence lifetime imaging (FLIM) and differential interference contrast (DIC). The system employs a Nd:YVO4 laser as pump (7 ps, 1064 nm), and two tunable OPOs (6 ps, 700 - 1000 nm). Our microscope comprises a heater stage and perfusion cell for imaging of live cells, and features an atomic force microscope (AFM) which enables optical imaging at 10 nm resolution. Multimodal imaging of breast cancer cells and tissue will be demonstrated as well as imaging of anticancer drugs in living cells.
Adipose-derived stem cells (ADSCs) are adult stem cells isolated from lipoaspirates. They are a g... more Adipose-derived stem cells (ADSCs) are adult stem cells isolated from lipoaspirates. They are a good candidate for autologuous cell therapy and tissue engineering. For these applications, label-free imaging could be critical to assess noninvasively the efficiency of stem cell (SC) differentiation. We report on the development and application of a multimodal microscope to monitor and quantify ADSC differentiation into osteoblasts and adipocytes.
Tip-enhanced optical spectroscopy is an approach that holds a good deal of promise for the nanosc... more Tip-enhanced optical spectroscopy is an approach that holds a good deal of promise for the nanoscale characterisation of matter. Tip-enhanced Raman spectroscopy (TERS) has been demonstrated on a variety of samples: inorganic, organic and biological. Imaging using TERS has been shown for carbon nanotubes due to their high scattering efficiency. There are a number of compelling motivations to consider alternative approaches for biological samples; most importantly, the potential for heat damage of biomolecules and long acquisition times. These issues may be addressed through the development of tip-enhanced coherent anti-Stokes Raman scattering microscopy.
We have used a versatile and powerful microscope[1] for multi-modal biomedical imaging on which w... more We have used a versatile and powerful microscope[1] for multi-modal biomedical imaging on which we combine Coherent Anti-Stokes Raman Scattering (CARS) with Two Photon Excitation Fluorescence (TPEF) using a Nd: YVO4 pump laser. We acquired 2PEF, CARS, and phase contrast images of Multilamellar Vesicles (MLVs) and Giant Unilamellar Vesicles (GUVs), as well as Raman spectra of the constituent lipids. A wide range of peptides are harmful to cells by altering the structure of the biological membranes. This effect depends on the composition of the membrane and the chemical structure of the peptide. The peptide we studied is the beta amyloid Aβ which is a major component of the amyloid plaques deposited on neuronal membranes of Alzheimer's disease (AD) patients. AD is neurodegenerative disorder in which the hallmark symptoms include cognitive decline and dementia[2] and is characterized by the formation of extracellular amyloid fibrils on the neuronal membranes of the brain. Many questions still remain unanswered concerning the destabilization of cellular ionic homeostasis due to pores formed during the interactions of lipid membranes with peptides. In this project, biomimics of cell membranes are used. The structures that best mimic the plasma membranes are MLVs or GUVs. These vesicles are formed using the gentle hydration technique[3] or the electroformation technique[4] respectively and are composed of phospholipids such as DOPC, DPPC, D62PPC and their binary mixtures. The MLVs and GUVs imaging by CARS and TPEF microscopy not only permits the direct imaging of the leakage phenomenon caused by the toxic peptide (Aβ) on the lipid bilayer, but also records simultaneously the lateral structure of the bilayer and peptide distribution in the plane across the membrane.
Micro-Raman scattering are used like a probe to determiner the Co substitution process in the sys... more Micro-Raman scattering are used like a probe to determiner the Co substitution process in the system Li{2}O-Ta{2}O{5}-(CoO){2} . We follow the Raman spectra modifications as function of the substitution rate along the line B: Li{1+x}Ta{1-x/2}Co{3x/4}O{3} and confirm the substitution of Co2+ ions on the Li+ site.
ABSTRACT The elaboration of the CdS nanocrystals embedded in a NaCl single crystal has been perfo... more ABSTRACT The elaboration of the CdS nanocrystals embedded in a NaCl single crystal has been performed using the Czochralski method. The optical density spectrum shows a blue shift of the absorption edge towards the high energies. The energy gap has been found to be 2.70eV. The photoluminescence has been assigned to the neutral-donor band (D0, X) and radiative recombinations from deep defect and impurity levels.
Micro-Raman scattering are used like a probe to determiner the Co substitution process in the sys... more Micro-Raman scattering are used like a probe to determiner the Co substitution process in the system Li{2}O-Ta{2}O{5}-(CoO){2} . We follow the Raman spectra modifications as function of the substitution rate along the line B: Li{1+x}Ta{1-x/2}Co{3x/4}O{3} and confirm the substitution of Co2+ ions on the Li+ site.
The LiNbO3 crystals doped with Nd3+, Dy3+ and Er3+ ions were investigated by means of Raman scatt... more The LiNbO3 crystals doped with Nd3+, Dy3+ and Er3+ ions were investigated by means of Raman scattering measurements. Data were compared with results recorded in undoped congruent crystal and highly Mg-doped samples. The doping dependence of frequency and damping in the E(TO1) and the two lowest frequency A1(TO) phonon modes were employed to deduce the incorporation site of rare earth
The defect structure of Mg-doped lithium niobate crystals was studied with varying Mg doping conc... more The defect structure of Mg-doped lithium niobate crystals was studied with varying Mg doping concentration by means of Raman scattering measurements. The Mg content dependences of both the frequency and damping of the two lowest-frequency A1(TO) phonon modes were used to derive the Mg incorporation mechanism in the LiNbO3 lattice. Results indicated that Mg ions are located on the Li site for low doping and then replace both Li and Nb ions for larger concentration.
It has been established by Raman scattering study that the optical damage in nominally pure LiNbO... more It has been established by Raman scattering study that the optical damage in nominally pure LiNbO3 crystals presents an irreversible part. This irreversible fraction of the optical damage is induced only by the high-intensity (I ⩾ 25 kW/cm2) exposition for a long time, and it has accumulative character with exposure time. This irreversible effect has been shown to be related to the optical fatigue of the material. The consequent needle-like domain inversion was considered to cause accumulative deterioration of the material, i.e., a fatigue due to the continuous electric field application, relating to the high space-charge field induced by inhomogeneous high-intensity illumination.
Multiphoton Microscopy in the Biomedical Sciences X, 2010
ABSTRACT We report on the construction of a highly flexible system for advanced biological imagin... more ABSTRACT We report on the construction of a highly flexible system for advanced biological imaging, where all the following imaging techniques are integrated into the same microscope: Coherent anti-Stokes Raman scattering (CARS), two photon excitation fluorescence (TPEF), second harmonic generation (SGH), sum frequency generation (SFG), fluorescence lifetime imaging (FLIM) and differential interference contrast (DIC). The system employs a Nd:YVO4 laser as pump (7 ps, 1064 nm), and two tunable OPOs (6 ps, 700 - 1000 nm). Our microscope comprises a heater stage and perfusion cell for imaging of live cells, and features an atomic force microscope (AFM) which enables optical imaging at 10 nm resolution. Multimodal imaging of breast cancer cells and tissue will be demonstrated as well as imaging of anticancer drugs in living cells.
ABSTRACT We have used a versatile and powerful microscope[1] for multi-modal biomedical imaging o... more ABSTRACT We have used a versatile and powerful microscope[1] for multi-modal biomedical imaging on which we combine Coherent Anti-Stokes Raman Scattering (CARS) with Two Photon Excitation Fluorescence (TPEF) using a Nd: YVO4 pump laser. We acquired 2PEF, CARS, and phase contrast images of Multilamellar Vesicles (MLVs) and Giant Unilamellar Vesicles (GUVs), as well as Raman spectra of the constituent lipids. A wide range of peptides are harmful to cells by altering the structure of the biological membranes. This effect depends on the composition of the membrane and the chemical structure of the peptide. The peptide we studied is the beta amyloid Aβ which is a major component of the amyloid plaques deposited on neuronal membranes of Alzheimer's disease (AD) patients. AD is neurodegenerative disorder in which the hallmark symptoms include cognitive decline and dementia[2] and is characterized by the formation of extracellular amyloid fibrils on the neuronal membranes of the brain. Many questions still remain unanswered concerning the destabilization of cellular ionic homeostasis due to pores formed during the interactions of lipid membranes with peptides. In this project, biomimics of cell membranes are used. The structures that best mimic the plasma membranes are MLVs or GUVs. These vesicles are formed using the gentle hydration technique[3] or the electroformation technique[4] respectively and are composed of phospholipids such as DOPC, DPPC, D62PPC and their binary mixtures. The MLVs and GUVs imaging by CARS and TPEF microscopy not only permits the direct imaging of the leakage phenomenon caused by the toxic peptide (Aβ) on the lipid bilayer, but also records simultaneously the lateral structure of the bilayer and peptide distribution in the plane across the membrane.
We report on the construction of a highly flexible system for advanced biological imaging, where ... more We report on the construction of a highly flexible system for advanced biological imaging, where all the following imaging techniques are integrated into the same microscope: Coherent anti-Stokes Raman scattering (CARS), two photon excitation fluorescence (TPEF), second harmonic generation (SGH), sum frequency generation (SFG), fluorescence lifetime imaging (FLIM) and differential interference contrast (DIC). The system employs a Nd:YVO4 laser as pump (7 ps, 1064 nm), and two tunable OPOs (6 ps, 700 - 1000 nm). Our microscope comprises a heater stage and perfusion cell for imaging of live cells, and features an atomic force microscope (AFM) which enables optical imaging at 10 nm resolution. Multimodal imaging of breast cancer cells and tissue will be demonstrated as well as imaging of anticancer drugs in living cells.
Adipose-derived stem cells (ADSCs) are adult stem cells isolated from lipoaspirates. They are a g... more Adipose-derived stem cells (ADSCs) are adult stem cells isolated from lipoaspirates. They are a good candidate for autologuous cell therapy and tissue engineering. For these applications, label-free imaging could be critical to assess noninvasively the efficiency of stem cell (SC) differentiation. We report on the development and application of a multimodal microscope to monitor and quantify ADSC differentiation into osteoblasts and adipocytes.
Tip-enhanced optical spectroscopy is an approach that holds a good deal of promise for the nanosc... more Tip-enhanced optical spectroscopy is an approach that holds a good deal of promise for the nanoscale characterisation of matter. Tip-enhanced Raman spectroscopy (TERS) has been demonstrated on a variety of samples: inorganic, organic and biological. Imaging using TERS has been shown for carbon nanotubes due to their high scattering efficiency. There are a number of compelling motivations to consider alternative approaches for biological samples; most importantly, the potential for heat damage of biomolecules and long acquisition times. These issues may be addressed through the development of tip-enhanced coherent anti-Stokes Raman scattering microscopy.
We have used a versatile and powerful microscope[1] for multi-modal biomedical imaging on which w... more We have used a versatile and powerful microscope[1] for multi-modal biomedical imaging on which we combine Coherent Anti-Stokes Raman Scattering (CARS) with Two Photon Excitation Fluorescence (TPEF) using a Nd: YVO4 pump laser. We acquired 2PEF, CARS, and phase contrast images of Multilamellar Vesicles (MLVs) and Giant Unilamellar Vesicles (GUVs), as well as Raman spectra of the constituent lipids. A wide range of peptides are harmful to cells by altering the structure of the biological membranes. This effect depends on the composition of the membrane and the chemical structure of the peptide. The peptide we studied is the beta amyloid Aβ which is a major component of the amyloid plaques deposited on neuronal membranes of Alzheimer's disease (AD) patients. AD is neurodegenerative disorder in which the hallmark symptoms include cognitive decline and dementia[2] and is characterized by the formation of extracellular amyloid fibrils on the neuronal membranes of the brain. Many questions still remain unanswered concerning the destabilization of cellular ionic homeostasis due to pores formed during the interactions of lipid membranes with peptides. In this project, biomimics of cell membranes are used. The structures that best mimic the plasma membranes are MLVs or GUVs. These vesicles are formed using the gentle hydration technique[3] or the electroformation technique[4] respectively and are composed of phospholipids such as DOPC, DPPC, D62PPC and their binary mixtures. The MLVs and GUVs imaging by CARS and TPEF microscopy not only permits the direct imaging of the leakage phenomenon caused by the toxic peptide (Aβ) on the lipid bilayer, but also records simultaneously the lateral structure of the bilayer and peptide distribution in the plane across the membrane.
Micro-Raman scattering are used like a probe to determiner the Co substitution process in the sys... more Micro-Raman scattering are used like a probe to determiner the Co substitution process in the system Li{2}O-Ta{2}O{5}-(CoO){2} . We follow the Raman spectra modifications as function of the substitution rate along the line B: Li{1+x}Ta{1-x/2}Co{3x/4}O{3} and confirm the substitution of Co2+ ions on the Li+ site.
ABSTRACT The elaboration of the CdS nanocrystals embedded in a NaCl single crystal has been perfo... more ABSTRACT The elaboration of the CdS nanocrystals embedded in a NaCl single crystal has been performed using the Czochralski method. The optical density spectrum shows a blue shift of the absorption edge towards the high energies. The energy gap has been found to be 2.70eV. The photoluminescence has been assigned to the neutral-donor band (D0, X) and radiative recombinations from deep defect and impurity levels.
Micro-Raman scattering are used like a probe to determiner the Co substitution process in the sys... more Micro-Raman scattering are used like a probe to determiner the Co substitution process in the system Li{2}O-Ta{2}O{5}-(CoO){2} . We follow the Raman spectra modifications as function of the substitution rate along the line B: Li{1+x}Ta{1-x/2}Co{3x/4}O{3} and confirm the substitution of Co2+ ions on the Li+ site.
The LiNbO3 crystals doped with Nd3+, Dy3+ and Er3+ ions were investigated by means of Raman scatt... more The LiNbO3 crystals doped with Nd3+, Dy3+ and Er3+ ions were investigated by means of Raman scattering measurements. Data were compared with results recorded in undoped congruent crystal and highly Mg-doped samples. The doping dependence of frequency and damping in the E(TO1) and the two lowest frequency A1(TO) phonon modes were employed to deduce the incorporation site of rare earth
The defect structure of Mg-doped lithium niobate crystals was studied with varying Mg doping conc... more The defect structure of Mg-doped lithium niobate crystals was studied with varying Mg doping concentration by means of Raman scattering measurements. The Mg content dependences of both the frequency and damping of the two lowest-frequency A1(TO) phonon modes were used to derive the Mg incorporation mechanism in the LiNbO3 lattice. Results indicated that Mg ions are located on the Li site for low doping and then replace both Li and Nb ions for larger concentration.
It has been established by Raman scattering study that the optical damage in nominally pure LiNbO... more It has been established by Raman scattering study that the optical damage in nominally pure LiNbO3 crystals presents an irreversible part. This irreversible fraction of the optical damage is induced only by the high-intensity (I ⩾ 25 kW/cm2) exposition for a long time, and it has accumulative character with exposure time. This irreversible effect has been shown to be related to the optical fatigue of the material. The consequent needle-like domain inversion was considered to cause accumulative deterioration of the material, i.e., a fatigue due to the continuous electric field application, relating to the high space-charge field induced by inhomogeneous high-intensity illumination.
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