Dr. Claudia Scalfi-Happ Profile

Dr. Claudia Scalfi-Happ

at Univ Ulm

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Author

Publications (8)

Proceedings Article | 15 July 2015 Paper

Raman and fluorescence microscopy to study the internalization and dissolution of photosensitizer nanoparticles into living cells

Claudia Scalfi-Happ, Rudolf Steiner, Rainer Wittig, Susanna Graefe, Anastasia Ryabova, Victor Loschenov

Proceedings Volume 9537, 953705 (2015) https://doi.org/10.1117/12.2183726

KEYWORDS: Luminescence, Raman spectroscopy, Nanoparticles, Microscopy, Confocal microscopy, Photodynamic therapy, Imaging spectroscopy, Signal detection, Laser scanners, Crystals

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Proceedings Article | 18 June 2013 Paper

Label-free detection of tumor markers in a colon carcinoma tumor progression model by confocal Raman microspectroscopy

Claudia Scalfi-Happ, Angelika Rück, Martin Udart, Carmen Hauser, Christine Dürr, Martin Kriebel

Proceedings Volume 8798, 87980B (2013) https://doi.org/10.1117/12.2032459

KEYWORDS: Raman spectroscopy, Tumors, Imaging spectroscopy, Colon, Confocal microscopy, Tumor growth modeling, In vitro testing, Biological research, Data acquisition, Colorectal cancer

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Proceedings Article | 12 July 2007 Paper

Confocal Raman microscopy for investigation of the level of differentiation in living neuroblastoma tumor cells

Claudia Scalfi-Happ, Andrea Jauss, Olaf Hollricher, Simone Fulda, Carmen Hauser, Rudolf Steiner, Angelika Rück

Proceedings Volume 6630, 66300T (2007) https://doi.org/10.1117/12.729455

KEYWORDS: Raman spectroscopy, Confocal microscopy, Microscopy, Tumors, Chemical analysis, Imaging spectroscopy, Microscopes, Data acquisition, Proteins, Nd:YAG lasers

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The investigation of living cells at physiological conditions requires very sensitive, sophisticated, non invasive methods. In this study, Raman spectral imaging is used to identify different biomolecules inside of cells. Raman spectroscopy, a chemically and structurally sensitive measuring technique, is combined with high resolution confocal microscopy. In Raman spectral imaging mode, a complete Raman spectrum is recorded at every confocal image point, giving insight into the chemical composition of each sample compartment. Neuroblastoma is the most common solid extra-cranial tumor in children. One of the unique features of neuroblastoma cells is their ability to differentiate spontaneously, eventually leading to complete remission. Since differentiation agents are currently used in the clinic for neuroblastoma therapy, there is a special need to develop non-invasive and sensitive new methods to monitor neuroblastoma cell differentiation. Neuroblastoma cells at different degrees of differentiation were analysed with the confocal Raman microscope alpha300 R (WITec GmbH, Germany), using a frequency doubled Nd:YAG laser at 532 nm and 10 mW for excitation. Integration time per spectrum was 80-100 ms. A lateral resolution in submicrometer range was achieved by using a 60x water immersion lens with a numerical aperture of 1,0. Raman images of cells were generated from these sets of data by either integrating over specific Raman bands, by basis analysis using reference spectra or by cluster analysis. The automated evaluation of all spectra results in spectral unmixed images providing insight into the chemical composition of the sample. With these procedures, different cell organelles, cytosol, membranes could be distinguished. Since neuroblastoma cells at high degree of differentiation overproduce noradrenaline, an attempt was made to trace the presence of this neurotransmitter as a marker for differentiation. The results of this work may have applications in the monitoring of molecular changes and distribution of biomolecules and in particular of low molecular weight markers as they occur during the differentiation of neuroblastoma cells.

Proceedings Article | 23 February 2006 Paper

Improving fluorescence diagnosis of cancer by SLIM

Angelika Rück, Frank Dolp, Ingrid Kinzler, Carmen Hauser, Claudia Scalfi-Happ

Proceedings Volume 6089, 608906 (2006) https://doi.org/10.1117/12.644576

KEYWORDS: Luminescence, Semiconductor lasers, Fluorescence lifetime imaging, Rhodamine, Tumors, Cancer, Pulsed laser operation, Microscopes, Bladder cancer, Photodynamic therapy

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Although during the last years, significant progress was made in cancer diagnosis, using either intrinsic or specially designed fluorophores, still problems exist, due to difficulties in spectral separation of highly overlapping probes or in lack of specificity. Many of the problems could be circumvented by focusing on time-resolved methods. In combination with spectral resolved detection (spectral fluorescence lifetime imaging, SLIM) highly sophisticated fluorescence lifetime imaging can be performed which might improve specificity of cell diagnosis. To record lifetime images (τ-mapping) with spectral resolution a setup was realized consisting of a laser scanning microscope equipped with a 16 channel array for time-correlated single photon counting (TCSPC) and a spectrograph in front of the array. A Ti:Saphir laser can be used for excitation or alternatively ps diode lasers. With this system the time- and spectral-resolved fluorescence characteristics of different fluorophores were investigated in solution and in cell culture. As an example, not only the mitochondria staining dye rhodamine 123 could be easily distinguished from DAPI, which intercalates into nucleic acids, but also different binding sites of DAPI. This was proved by the appearance of different lifetime components within different spectral channels. Another example is Photofrin, a photosensitizer which is approved for bladder cancer and for palliative lung and esophageal cancer in 20 countries, including the United States, Canada and many European countries. Photofrin is a complex mixture of different monomeric and aggregated porphyrins. The phototoxic efficiency during photodynamic therapy (PDT) seems to be correlated with the relative amounts of monomers and aggregates. With SLIM different lifetimes could be attributed to various, spectrally highly overlapping compounds. In addition, a detailed analysis of the autofluorescence by SLIM could explain changes of mitochondrial metabolism during Photofrin-PDT.

Proceedings Article | 30 March 2005 Paper

FLIM and SLIM for molecular imaging in PDT

Angelika Rueck, Frank Dolp, Christian Huelshoff, Carmen Hauser, Claudia Scalfi-Happ

Proceedings Volume 5700, (2005) https://doi.org/10.1117/12.590439

KEYWORDS: Luminescence, Fluorescence lifetime imaging, Photodynamic therapy, Semiconductor lasers, Microscopes, Imaging systems, Laser scanners, Molecular imaging, Time resolved spectroscopy, Single photon

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Various problems arising during molecular imaging of different fluoroprobes and metabolites used in photodynamic therapy could be circumvented by focusing on time-resolved detection. For this, an interesting new method seems to be time-correlated single photon counting, where a time-to-amplitude converter determines the temporal position and a scanning interface connected to the scanning unit of a laser microscope determines the spatial location of a signal. In combination with spectral resolved detection (spectral lifetime imaging) the set-up achieves the features of highly sophisticated lifetime imaging systems. The photoactive substance on which 5-ALA PDT is based, is protoporphyrine IX which is synthesized in mitochondria. Alternatively, other metabolites from 5-ALA could be involved. Subcellular differentiation of those metabolites without extensive extraction procedures is not trivial, because of highly overlapping spectral properties. Measuring the fluorescence lifetime on a subcellular level could be a successful alternative. To record lifetime images (τ-mapping) a setup consisting on a laser scanning microscope equipped with detection units for time-correlated single photon counting and ps diode lasers for short-pulsed excitation was implemented. The time-resolved fluorescence characteristics of 5-ALA metabolites were investigated in solution and in cell culture. The lifetimes were best fitted by a biexponential fitting routine. Different lifetimes could be found in different cell compartments. During illumination, the lifetimes decreased significantly. Different metabolites of 5-ALA could be correlated with different fluorescence lifetimes. In addition cells were coincubated with the nuclear staining dye DAPI, in order to investigate the cell cycle. Using appropriate filtering or alternatively spectral lifetime imaging the time-resolved fluorescence of DAPI could be very well distinguished from 5-ALA-metabolites. In contrast to ALA, the lifetime of DAPI, which was best fitted monoexponentially did not change during photobleaching, making this dye a perfect internal standard.

Proceedings Article | 10 September 2004 Paper

Imaging time-resolved fluorescence characteristics of organelle specific fluorophores and photosensitizers using ps pulsed diode lasers and TCSPC techniques

Claudia Scalfi-Happ, Frank Dolp, Florian Forster, Angelika Rueck

Proceedings Volume 5462, (2004) https://doi.org/10.1117/12.545472

KEYWORDS: Luminescence, Rhodamine, Semiconductor lasers, Picosecond phenomena, Laser scanners, Microscopes, Fluorescence lifetime imaging, Time resolved spectroscopy, Microscopy, Photodynamic therapy

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Proceedings Article | 16 October 2003 Paper

Thermal effects in IR-laser-irradiated living cells

Thomas Meier, Angelika Rueck, Claudia Scalfi-Happ, Hubert Hug, Marion Schneider

Proceedings Volume 5142, (2003) https://doi.org/10.1117/12.501158

KEYWORDS: Liquid crystals, Temperature metrology, Fused deposition modeling, Thermal effects, Cell death, Infrared lasers, Absorption, Finite difference methods, Crystals, Tumors

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Proceedings Article | 9 October 2003 Paper

Time-resolved microspectrofluorometry and fluorescence lifetime imaging using ps-pulsed diode lasers in laser scanning microscopes

Angelika Rueck, Frank Dolp, Claudia Scalfi-Happ, Rudolf Steiner, Michael Beil

Proceedings Volume 5139, (2003) https://doi.org/10.1117/12.500456

KEYWORDS: Luminescence, Laser scanners, Fluorescence lifetime imaging, Semiconductor lasers, Microscopes, Rhodamine, Time resolved spectroscopy, Picosecond phenomena, Microscopy, Photodynamic therapy

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