Achim Stangelmayer

Dual Lifetime Referencing (DLR) - A New Scheme for Converting Fluorescence Intensity into a Frequency-Domain or Time-Domain Information

Fluorescence spectroscopy and NMR spectroscopy are probably the most powerful spectroscopies at present albeit with very different (and highly complementary) fields of application. Fluorometry can be based on the intrinsic fluorescence of (bio)molecules or ions, or on the use of fluorescent probes, indicators, or labels Numerous parameters can be measured which include intensity, decay time, polarization, radiative and non-radiative energy transfer, quenching efficiency, and combinations thereof Fluorescence microscopy and imaging are other widely applied techniques, and multi-dimensional and synchronous fluorescence spectroscopy have gained some interest in recent years.

An imaging method for oxygen distribution, respiration and photosynthesis at a microscopic level of resolution

Biological samples are far from homogeneous, with complex compartmentation being the norm. Major physiological processes such as respiration do not therefore occur in a uniform manner within most tissues, and it is currently not possible to image its gradients in living plant tissues. A compact fluorescence ratiometric-based device is presented here, consisting of an oxygen-sensitive foil and a USB (universal serial bus) microscope. The sensor foil is placed on the sample surface and, based on the localized change in fluorescence signal over time, information about the oxygen consumption (respiration) or evolution (photosynthesis) can be obtained. Using this imaging technique, it was possible to demonstrate the spatial pattern of oxygen production and consumption at a c. 20-μm level of resolution, and their visualization in the rhizosphere, stem and leaf, and within the developing seed. The oxygen mapping highlighted the vascular tissues as the major stem sink for oxygen. In the leaf, the level of spatial resolution was sufficient to visualize the gas exchange in individual stomata. We conclude that the novel sensor set-up can visualize gradients in oxygen-consuming and producing processes, thereby facilitating the study of the spatial dynamics of respiration and photosynthesis in heterogeneous plant tissues.

Methodology and Significance of Microsensor-based Oxygen Mapping in Plant Seeds - an Overview.

Oxygen deficiency is commonplace in seeds, and limits both their development and their germination. It is, therefore, of considerable relevance to crop production. While the underlying physiological basis of seed hypoxia has been known for some time, the lack of any experimental means of measuring the global or localized oxygen concentration within the seed has hampered further progress in this research area. The development of oxygen-sensitive microsensors now offers the capability to determine the localized oxygen status within a seed, and to study its dynamic adjustment both to changes in the ambient environment, and to the seeds developmental stage. This review illustrates the use of oxygen microsensors in seed research, and presents an overview of existing data with an emphasis on crop species. Oxygen maps, both static and dynamic, should serve to increase our basic understanding of seed physiology, as well as to facilitate upcoming breeding and biotechnology-based approaches for crop improvement.

Planar Oxygen Sensors for Non Invasive Imaging in Experimental Biology

The presence of molecular oxygen is a sine qua non for aerobic metabolism. In both plant and animal mitochondria, it acts as the terminal electron acceptor for oxidative phosphorylation occurring during cellular respiration, and is necessary for the generation of ATP, the common energy currency within the living cell (Atkinson, 1977; Cooper, 2000). It is the major by-product of photosynthesis in which plant biomass is accumulated by the conversion of carbon dioxide into polymeric compounds. Since respiration and photosynthesis are so fundamental to life on earth, an understanding of the mechanisms underlying oxygen consumption, production and homoeostasis has become a significant field of both biological and biotechnological research (Volkmer et al., 2009). Oxygen (micro-) sensors, which are widely used in the life sciences, are designed to provide a precise measurement of the concentration of oxygen within a localized region of a tissue or an organ (Borisjuk & Rolletschek, 2009). Most of these devices have been based on miniaturized Clark-type electrodes (Revsbech & Jorgensen, 1986), in which oxygen diffuses into the sensor via a permeable membrane, following which its reduction at the cathode generates a measurable electrical current. This approach can deliver a spatial resolution at the low μm scale. Increasingly this technology is being replaced by optical oxygen microsensors (micro-optodes) based on fibre optic materials (Klimant et al., 1995; Rolletschek et al., 2009), in which the concentration is assessed in tapered glass fibres of tip size ~50μm via the dynamic quenching of a luminophore. This approach enjoys several advantages over the electrochemical detection system, as detailed elsewhere (Kuhl & Polerecky, 2008; Rolletschek et al., 2009). Importantly, microsensor-based approaches are invasive, which means that a given biological sample cannot be readily studied over a prolonged time period. Furthermore, the internal structure of most biological samples is far from homogeneous, with complex compartmentation being the norm. As a result, whole tissue measurements can only reflect the mean performance of a tissue, and cannot report variation between distinct compartments. This loss of richness compromises the value of such data for elucidating the biology of the tissue as a whole. At best, conventional sensor systems assess oxygen