TY - CONF A1 - Kage, Daniel A1 - Hoffmann, Katrin A1 - Resch-Genger, Ute T1 - FLiMFlow – Recent achievements in flow cytometry with lifetime detection and lifetime-encoded beads N2 - Flow cytometry is a widespread technique in biological research and clinical applications. Two different directions are currently of importance in development of new methods in this field. Whereas analyses in research become increasingly complex and require a growing number of simultaneously detectable codes and fluorescent labels, also low-cost methods and portable devices are needed in routine application. Lifetime encoding could present an alternative to common spectral multiplexing. On the one hand, it addresses the need for additional codes by combining spectral and lifetime multiplexing. On the other hand, using only lifetime encoding could help to reduce instrument costs by keeping the number of excitation sources and detectors low. Here, we report on our recent progress in employing dye-stained lifetime-encoded polymer microparticles as a model system for lifetime encoding in flow cytometry. The discrimination of two lifetime codes was achieved with two bead sets. Moreover, the simultaneous detection of a spectrally different ligand fluorescence signal could be demonstrated. T2 - FLiMFlow meeting CY - Pisa, Italy DA - 28.09.2016 KW - Flow cytometry KW - Fluorescence KW - Life sciences KW - Lifetime encoding KW - Polymer particles PY - 2016 AN - OPUS4-38239 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Kage, Daniel A1 - Hoffmann, Katrin A1 - Resch-Genger, Ute T1 - Luminophores for time-resolved flow cytometry N2 - For the application of time-resolved detection in the framework of flow cytometry, suitable luminescence lifetime code carriers are required. Here we report on our achievements concerning strategies to increase the accessible range of lifetime values and to realize continuously tunable lifetimes. To that end, we investigated polymer (PMMA) microbeads stained with mixtures of dyes exhibiting different fluorescence decay kinetics. At the expense of spectrally varying decay kinetics, it is possible to modify the lifetime by changing the dye concentration ratio. Moreover, semiconductor quantum dots incorporated into polymer beads were studied as alternative luminophores outperforming organic dyes with respect to long luminescence lifetimes, flexible choice of excitation wavelength and narrow spectral emission width. Our experiments demonstrate that lifetime adaption with dye mixing is basically feasible and semiconductor quantum dots represent promising candidates for long-lifetime codes. T2 - FLiMFlow meeting CY - Münster, Germany DA - 03.03.2017 KW - Flow cytometry KW - Fluorescence KW - Life sciences KW - Lifetime encoding KW - Polymer particles PY - 2017 AN - OPUS4-39307 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Kage, Daniel A1 - Fischer, Linn A1 - Hoffmann, Katrin A1 - Wittkamp, M. A1 - Ameskamp, J. A1 - Göhde, W. A1 - Thiele, T. A1 - Schedler, U. A1 - Resch-Genger, Ute T1 - Stained microparticles for lifetime encoding - Flow cytometry application & photophysics N2 - Multiparametric analyses involving optical techniques like flow cytometry are at the core of studying complex systems in biological research and diagnostic applications. However, for fluorescence-based techniques, the number of reporters distinguishable in spectral multiplexing is limited by spectral overlap and requires a multitude of excitation light sources and detection Systems. Intensity encoding often used for bead assays suffers from problems regarding dye concentration control and excitation light intensity fluctuations. An alternative is luminescence lifetime encoding, particularly to minimize instrument costs. Here, we report on dye-stained polymer microparticles for lifetime encoding in flow cytometry with different organic dyes. This ranges from studies of the impact of parameters like dye loading concentration and particle diameter on fluorescence decay behavior to the demonstration of lifetime code reading and simultaneous ligand fluorescence signal detection with single-wavelength excitation in a flow. T2 - DPG Spring Meeting 2017 CY - Dresden, Germany DA - 19.03.2017 KW - Flow cytometry KW - Fluorescence KW - Life sciences KW - Lifetime encoding KW - Polymer particles PY - 2017 AN - OPUS4-40056 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Kage, Daniel A1 - Hoffmann, Katrin A1 - Wittkamp, M. A1 - Ameskamp, J. A1 - Göhde, W. A1 - Thiele, T. A1 - Schedler, U. A1 - Resch-Genger, Ute T1 - Dye-stained lifetime-encoded polymer microbeads for application in time-resolved flow cytometry N2 - Flow cytometry is a standard analytical tool for biological research and in medical applications. There are different requirements triggering recent device and method development depending on the desired field of application. One trend is governed by the need for an increasing number of simultaneously detectable codes, i.e., fluorescent labels. The other one focuses on cost-effective methods and development of miniaturized, portable devices. Fluorophore encoding is usually based on spectral encoding. However, this approach is hampered by, e.g., spectral crosstalk. Additionally, the sensitivity of fluorescence intensity measurements to fluctuations in excitation light intensity and dye concentration limits the achievable number of detection channels. Moreover, spectral multiplexing typically requires several costly excitation light sources. Lifetime multiplexing and the discrimination between different encoding fluorophores and carrier beads based on their fluorescence decay kinetics could present an innovative alternative. Encoded beads, i.e., beads with lifetime codes corresponding to the surface chemistry, have been employed to evaluate the feasibility of this approach with a custom designed flow cytometer equipped with a pulsed light source and a fast detector for time-resolved measurements in a flow. In a first step, we used steady state and time-resolved photoluminescence measurements for the spectroscopic characterization of micrometer-sized dye-stained PMMA beads. Subsequently, the potential use of these microbeads for flow cytometry applications was analyzed with a prototype flow cytometer with lifetime detection. With our proof-of-concept studies, we could demonstrate that lifetime discrimination and simultaneous readout of a ligand fluorescence signal for analyte quantification is feasible with a set of dye-stained polymer microbeads at single wavelength excitation. These studies are expected to pave the road for new applications of fluorescence lifetime multiplexing in time-domain flow cytometry and bead-based assays in general. T2 - DBS/EBS 2017 CY - Potsdam, Germany DA - 20.03.2017 KW - Flow cytometry KW - Fluorescence KW - Life sciences KW - Lifetime encoding KW - Polymer particles PY - 2017 AN - OPUS4-39996 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Resch-Genger, Ute A1 - Wegner, Karl David A1 - Kalot, G. A1 - Busser, B. A1 - Pliquett, J. A1 - Köster, U. A1 - Koll, J. C. A1 - Denat, F. A1 - Bodio, E. A1 - Goze, C. A1 - Sacey, L. T1 - NIR aza-BODIPY: a new vector for boron neutron capture therapy (BNCT) N2 - Boron neutron capture therapy (BNCT) relies on the activation of 10B by thermal neutrons, which results in small highly energetic particle emission inducing cancer cells damage. However, in order to overcome the limits of the currently used BNCT agents, it is necessary to design new systems, which can specifically accumulate and deliver a sufficient amount of 10B in tumors. In this study, we designed a 10B-BSH-containing aza-BODIPY (aza-SWIR-BSH). It enabled the efficient vectorization of clinically used 10B-BSH to the tumor, resulting in higher therapeutic activity than the 10B-BSH alone. T2 - EMIM 2021 CY - Göttingen, Germany DA - 24.08.2021 KW - Flourescence KW - Optical probe KW - Dye KW - Photophysics KW - Quantum yield KW - Mechanism KW - NIR KW - SWIR KW - Imaging KW - Method KW - Quality assurance KW - BODIPY KW - Boron Neutron Capture Therapy (BNCT) KW - Medicine KW - Life sciences PY - 2021 AN - OPUS4-53731 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Resch-Genger, Ute A1 - Nirmalananthan-Budau, Nithiya A1 - Tavernaro, Isabella A1 - Kläber, Christoph A1 - Kunst, Alexandra T1 - Design, characterization, and application of fluorescent sensor particles N2 - pH and oxygen are amongst the most important and frequently measured analytes in the life and material sciences, indicating, e.g., diseases and corrosion processes. This includes the optical monitoring of pH in living cells for studying cellular internalization pathways, such as phagocytosis, endocytosis, and receptor ligand internalization with the aid of molecular and nanoscale fluorescent sensors. Nanoparticle (NP)-based sensors, that are labeled or stained with a multitude of sensor dyes, have several advantages as compare to conventional molecular probes like enhanced brightness, i.e., amplified signals, ease of designing ratiometric systems by combining analyte sensitive and inert reference dyes, and increased photostability. Moreover, this can enable the use of hydrophobic dyes in aqueous environments. Versatile templates and carriers for the fabrication of nanosensors by the staining and/or labelling with different fluorophores and sensor molecules or surface functionalized NP like silica (SiO2-NP) and polystyrene (PS-NP) particles provide. Here we present the design of a versatile platform of color emissive nanosensors and stimuli-responsive microparticles for the measurement of pH, oxygen, and other targets utilizing both types of matrices and sets of spectrally distinguishable sensor and reference dyes and their characterization and demonstrate the applicability of representative sensor particle for cellular studies. T2 - Vortrag bei dem Projekttreffen MicraGen CY - Copenhagen, Denmark DA - 18.08.2022 KW - Dye KW - Luminescence KW - Quantitative spectroscopy KW - Photophysics KW - Quality assurance KW - Nano KW - Particle KW - Quantum yield KW - Lifetime KW - Standard KW - Integrating sphere spectroscopy KW - Sensor KW - pH KW - Oxygen KW - Microfluidics KW - Cancer KW - Cell KW - Life sciences PY - 2022 AN - OPUS4-57049 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER -