Fluorescence Lifetime Imaging Solutions

Leica Microsystems is at the cutting edge of today’s fluorescence lifetime imaging advances. Our systems make lifetime imaging faster and easier to use than ever before, bringing its advantages to every day confocal imaging experiments.

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Our experts on solutions for FLIM are happy to help you with their advice.

Boost confocal imaging with fluorescence lifetime information

Fluorescence lifetime imaging microscopy (FLIM) is an imaging technique that makes use of the inherent properties of fluorescent dyes. Aside from having characteristic emission spectra, each fluorescent molecule has a characteristic lifetime that reflects how long the fluorophore spends in the excited state before emitting a photon. Lifetime analysis provides you with information in addition to your standard fluorescence intensity measurements.

A Guide to FLIM

The lifetime contrast of sepal of silverberry allows to separate different structure within the sample. The intensity image (yellow image) doesn't show any specific structure while the FastFLIM (rainbow scale image) and the lifetime fitted images (RGB, three component image) give insigth into the different part of the sample. The end of the video shows the three structures determined by the lifetime fit split into three channels (RGB).

Single slice taken from a zebrafish heart showing the ventricle with an injury in the lower area. Image shows the nuclei of all cells (blue), nuclei of the cardiomyocyte (heart muscle cells, green) and the proliferating cells (red). Image courtesy of Laura Peces-Barba Castaño, Max Planck Institute for Heart and Lung Research.
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Improve quality

Fluorescence lifetime imaging provides additional information that can help you improve the quality of your confocal imaging. It can be particularly useful to discriminate fluorescence probes that have overlapping fluorescent emission spectra, or to eliminate unwanted background fluorescent signals.

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Functional imaging

FLIM is a crucial application for measuring and quantifying imaging data. As lifetime information is independent of fluorophore concentration, it is extremely well suited for functional imaging. Functional imaging goes beyond the traditional recording of the location and concentration of molecular species and enables further investigation of molecular function, interactions, and their environment.

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Novel imaging applications

FLIM enables novel imaging applications, such as the use of biosensors to study the cell’s microenvironment or imaging based on the intrinsic fluorescent properties of your sample. Learn more about FLIM and its imaging applications with the SP8 FALCON system by referring to this application note.

Setting the standard for lifetime imaging

STELLARIS 8 is a forward-looking system, offering all the benefits of the STELLARIS 5 core system plus added functionality thanks to an extended spectrum WLL and specialized detector options of the Power HyD detector family.

Leica Microsystems solutions for FLIM enable you to harness the power of fluorescence lifetime to extract a new dimension of information from every sample with your confocal experiment. Investigate cellular physiology and explore cellular dynamics with FALCON. Get immediate access to functional information such as metabolic status, pH and ion concentration with the potential of Tausense in STELLARIS.

With our simplified, integrated platforms, you can gain new perspectives on your experiments, allowing you to harness new answers to your experimental questions.

Intensity

TauSeparation

Mammalian cells expressing LifeAct-GFP (manufactured by ibidi GmbH) and labeled with a MitoTracker Green. Acquisition with one detector, separation performed by TauSeparation.

FLIM imaging with STELLARIS confocal platform

Confocal microscopy relies on the effective excitation of fluorescence probes and the efficient collection of photons emitted from the fluorescence process. One aspect of fluorescence is the emission wavelength (spectral characteristics of a fluorophore). Another one, very powerful albeit less explored, is the fluorescence lifetime (the time a fluorophore stays in the excited state). Information based on fluorescence lifetime adds an additional dimension to confocal experiments, revealing information about the fluorophore microenvironment and enabling multiplex of spectrally overlapping species.

Fast & fully integrated lifetime imaging technology

With the release of the fast and fully integrated lifetime imaging technology – the SP8 FALCON – Leica Microsystems set the benchmark for the future of lifetime imaging. Leica Microsystems received the prestigious R&D 100 Award for the SP8 FALCON system. This award recognizes the solution’s innovative approach to lifetime imaging.

FLIM Microscope STELLARIS 8 FALCON

A novel concept in fluorescence lifetime imaging enabling video-rate confocal FLIM

SP8 FALCON (FAst Lifetime CONtrast) is a fast and completely integrated fluorescence lifetime imaging microscopy (FLIM) confocal platform. SP8 FALCON delivers video-rate FLIM with pixel-by-pixel quantification, thanks to a novel concept for measuring fluorescence lifetimes built on fast electronics and sensitive spectral hybrid detectors. Photon arrival times are recorded at count rates typical for standard confocal imaging. The system has ultra-short dead time and powerful built-in algorithms for data acquisition and analysis. The deep integration of FLIM into the confocal platform provides easy access to complex FLIM experiments.

Live 3D FLIM on a patient-derived organoid. Monitoring a cancer-derived increase of signal transduction activity in live organoids with an ERK FRET-FLIM biosensor. Functional information showing FRET efficiency of the ERK biosensor. Higher FRET efficiency indicates higher ERK activity.

Adding Dimensions to Multiplex Molecular Imaging

Molecular imaging of living specimens offers a means to draw upon the growing body of high-throughput molecular data to better understand the underlying cellular and molecular mechanisms of complex events ranging from embryonic development to disease processes. However, imaging approaches are challenged by unavoidable tradeoffs between spatial resolution, temporal resolution, field of view and the limited photon budget.

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Find Relevant Specimen Details from Overviews

Switch from searching image by image to seeing the full sample overview quickly and identifying the important specimen details instantly. Use that knowledge to set up high-resolution image acquisition automatically using templates for slides, dishes, and multi-well plates. Like a GPS for the cells of a specimen, users always have a clear path to high-quality data with the LAS X Navigator software, a powerful navigation tool for the Life Science platforms STELLARIS and THUNDER Imagers. LAS X Navigator enables widefield, stereo, or confocal experiments to be combined with a stage application.

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How to Quantify Changes in the Metabolic Status of Single Cells

Metabolic imaging based on fluorescence lifetime provides insights into the metabolic dynamics of cells, but its use has been limited as expertise in advanced microscopy techniques was needed. Now, STELLARIS 8 DIVE FALCON makes metabolic imaging accessible to every scientist, thanks to the integration of phasor analysis to easily use fluorescence lifetime information.

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Read our latest articles about Fluorescence Lifetime Imaging

The knowledge portal of Leica Microsystems offers scientific research and teaching material on the subjects of microscopy. The content is designed to support beginners, experienced practitioners and scientists alike in their everyday work and experiments.

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Transverse histological cut of a rabbit tongue. 50 Mpixels images (2326 µm x 1739 µm) in 14 x 18 tiles. Lifetime gives an additional contrast that allows to differentiate different structures in histological stainings.

A Guide to Fluorescence Lifetime Imaging Microscopy (FLIM)

The fluorescence lifetime is a measure of how long a fluorophore remains on average in its excited state before returning to the ground state by emitting a fluorescence photon.
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Tubulin - TauInteraction

TauInteraction – Studying Molecular Interactions with TauSense

Fluorescence microscopy constitutes one of the pillars in life sciences and is a tool commonly used to unveil cellular structure and function. A key advantage of fluorescence microscopy resides in the…
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Live-Cell Imaging Techniques

The understanding of complex and/or fast cellular dynamics is an important step for exploring biological processes. Therefore, today’s life science research is increasingly focused on dynamic…
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Fluorescence Lifetime-based Imaging Gallery

Confocal microscopy relies on the effective excitation of fluorescence probes and the efficient collection of photons emitted from the fluorescence process. One aspect of fluorescence is the emission…
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How to Quantify Changes in the Metabolic Status of Single Cells

Metabolic imaging based on fluorescence lifetime provides insights into the metabolic dynamics of cells, but its use has been limited as expertise in advanced microscopy techniques was needed. Now,…
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Adding Dimensions to Multiplex Molecular Imaging

Molecular imaging of living specimens offers a means to draw upon the growing body of high-throughput molecular data to better understand the underlying cellular and molecular mechanisms of complex…
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How FLIM Microscopy helps to detect microplastic pollution

The use of autofluorescence in biological samples is a widely used method to gain detailed knowledge about systems or organisms. This property is not only found in biological systems, but also…
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Development of fluorescence lifetime imaging microscopy (FLIM) and its relevance for functional imaging

Prof. Ammasi Periasamy, Director, Keck Center for Cellular Imaging, University of Virginia, was interviewed by Dr. Giulia Ossato, Product Manager functional imaging, during Leica Microsystems Meets…
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Single-residue bioorthogonal labeling of G protein-coupled receptors (GPCRs) in live cells for quantitative analysis

To perform quantitative studies of membrane bound G protein-coupled receptors (GPCRs) with high-performance microscopy, the proteins must first be labeled with bright and photostable dyes. The…
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primary murine pre B cells expressing the eCALWY-6 Zn2+

Researchers use Functional Imaging to Elucidate the Role of Cellular Zn²⁺ in Human Immunity

An international consortium of researchers has found a new mutation in a gene harboring a reticulum-to-cytoplasm zinc transporter. The gene is called ZIP7 and it is responsible for the early onset of…
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SP8 DIVE Multiphoton Microscope Image Gallery

Today’s life science research focusses on complex biological processes, such as the causes of cancer and other human diseases. A deep look into tissues and living specimens is vital to understanding…
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FLIM FRET - Förster Resonance Energy Transfer

A typical application of FLIM is FLIM-FRET. FRET is a well-established technique to study molecular interactions. It scrutinizes protein binding and estimates intermolecular distances on an Angström…
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FCS - Fluorescence Correlation Spectroscopy

FCS is a fluorescence-based measurement method. Fluorescent molecules passing through a strongly focused, fixed laser beam are excited for fluorescence emission. After passing a confocal pinhole, the…
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Lifetime – a Proper Alternative

„Way too complicated!“ - the notorious feedback when it comes to fluorescence lifetime measurements. This will change now! New technologies and new concepts for data evaluation, all implemented in…
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FCCS - Fluorescence Cross-Correlation Spectroscopy

FCCS (Fluorescence Cross-Correlation Spectroscopy) can be measured using the Leica TCS SP8 FCS system. Similar to FCS , it analyzes fluorescence intensity fluctuations derived from a small…
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What is Photomanipulation?

The term photomanipulation describes a wide range of techniques that enable the microscopist the transition from passive observer to instigator of events by offering a way of interacting with their…
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Testing the Münch Hypothesis of Long Distance Phloem Transport in Plants

Long distance transport in plants occurs in sieve tubes of the phloem. The pressure flow hypothesis introduced by Ernst Münch in 1930 describes a mechanism of osmotically generated pressure…
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Video Talk by Roger Tsien: Fluorescent Protein Indicators

In this talk, Roger Tsien discusses how fluorescent proteins have been turned into indicators for a wide variety of biological molecules, including pH, ions, redox potential, and signaling molecules…
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