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 Live cell imaging

Visualization of morphological dynamics in hippocampal cell cultures

To visualize morphological dynamics in dendritic processes during synaptogenesis 2D and 3D-time sequences of EGFP-transfected neurons are acquired by confocal-laserscanning microscopy. Neurons, grown for 10 days, display numerous growth cones at distal processes and highly motile filopodia along their proximal dendrites, whereas other dendritic protrusions persist within the observation period of 2 to 3 hours. Interestingly, small, motile filopodial protrusions arise also from the heads of these presumable spine precursors (see: Kreutz et al., 2008).

Mitochondria traffic


Co-transfection of mitochondria with Bassoon reveals a subfraction of labelled organelles that temporarily comigrate during confocal live cell imaging (LSM).

Long period observation of mitochondria in neuronal processes by single photon counting with our Fluorescence Lifetime Imaging Microscope (FLIM) shows fission and fusion as well as complex anterograde and retrograde movements with resting phases and oscillatory behavior. Note, the mitochondria were continuously exposed to laser illumination and recorded for > 10h. (see: Vitali et al. 2011)

Metabolic mapping

Analysis of natural autofluorescence of cells of the immune and neuronal system to predict metabolic states. Taking the advantage of the autofluorescence of NAD(P)H one follows an indicator for intracellular energy production and enzyme activity without manipulating the cells through labeling by transfections or fluorescence dyes. Since NAD(P)H is distributed almost everywhere inside a cell, it can be used to image cells from cell populations up to single cells.

NAD(P)H is an important intracellular electron carrier and plays a key role in cell metabolism. Its fluorescence intensity reflects the intracellular amount and therefore correlates with the metabolic state of a cell. The signal of NAD(P)H can show dynamics as periodic changes through metabolic oscillations or displays aerobiosis and anaerobiosis by decreased and increased fluorescence intensities, respectively.

Furthermore, it is possible to discriminate between protein bound and freely diffusing NAD(P)H by its fluorescence lifetimes. By using a Fluorescence Lifetime Imaging (FLIM) setup equipped with extremely sensitive single photon counting detectors we are investigating the autofluorescence of cell populations and cellular networks to draw up metabolic maps.

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