Real-time Fluorescence Microscopy
Real-time Fluorescence Microscopy - Related Videos
16:10
A TIRF Microscopy Technique for Real-time, Simultaneous Imaging of the TCR and its Associated Signaling Proteins
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Cited by 13 •
2012
The compartmentalization of proteins either within the plasma membrane or into intracellular locations is one regulatory mechanism that can greatly influence signaling outcomes; hence, to understand signaling it is important to study the spatial and temporal behavior of the proteins involved. We describe here a TIRF microscopy based system to study signal transduction in T cells, but is broadly applicable.
06:31
Real-time Imaging of Plant Cell Surface Dynamics with Variable-angle Epifluorescence Microscopy
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Cited by 10 •
2015
The goal of this protocol is to demonstrate how to monitor fluorescently-tagged protein dynamics on plant cell surfaces with variable-angle epifluorescence microscopy, showing blinking dots of GFP-tagged PATROL1, a membrane trafficking protein, in the cell cortex of the stomatal complex in Arabidopsis thaliana.
04:23
Real-Time Analysis of Resident T-cell Migration in Different Tumor Regions Using Confocal Microscopy
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2025
This video describes the imaging of resident tumor-infiltrating CD8 T cells labeled with fluorescently coupled antibodies within human lung tumor slices. This technique permits real-time analyses of CD8 T cell migration using confocal microscopy.
06:37
Multispectral Real-time Fluorescence Imaging for Intraoperative Detection of the Sentinel Lymph Node in Gynecologic Oncology
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Cited by 41 •
2010
Fluorescence imaging is a promising innovative modality for image-guided surgery in surgical oncology. In this video we describe the technical procedure for detection of the sentinel lymph node using fluorescence imaging as showcased in gynecologic oncologicy. A multispectral fluorescence camera system, together with the fluorescent agent indocyanine green, is applied.
10:34
FRET Microscopy for Real-time Monitoring of Signaling Events in Live Cells Using Unimolecular Biosensors
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Cited by 29 •
2012
Förster resonance energy transfer (FRET) microscopy is a powerful technique for real-time monitoring of signaling events in live cells using various biosensors as reporters. Here we describe how to build a customized epifluorescence FRET imaging system from commercially available components and how to use it for FRET experiments.
FAQs
Fluorophores first absorb excitation light and then emit light at a longer wavelength. Optical filters separate this emitted signal from the excitation illumination, allowing the microscope to detect the labeled structure or molecule with greater specificity. This separation is essential for distinguishing the target fluorescence from the illumination used to activate it during repeated imaging.
Time-lapse acquisition records successive images rather than relying on a single observation. Comparing these images reveals how a labeled target changes, moves, or redistributes over time. This temporal information can connect molecular or structural changes with events such as cell division, migration, invasion, or altered behavior during treatment, which fixed images alone cannot show.
Fixed-sample imaging provides a snapshot of structures at one selected time, whereas real-time fluorescence microscopy follows labeled targets as events unfold. The time-resolved view can show the sequence and timing of biological changes, helping researchers relate intracellular activity to later changes in cell behavior. This distinction is particularly valuable when cancer-related processes are dynamic.
A typical sequence begins by identifying the structure or molecule to be fluorescently labeled, directing excitation light onto the specimen, and collecting the longer-wavelength emission through optical filters. The microscope then repeats image acquisition over time to create a time-lapse record. In living cells or tissues, this workflow preserves the temporal context needed to examine ongoing biological events.
In cancer research, the method can track cell division, migration, invasion, and intracellular signaling as they occur. These observations show not only where labeled components are located, but also how their changes accompany tumor-cell behavior. Such visual evidence helps investigators examine disease mechanisms in a dynamic context rather than interpreting cellular activity from isolated, fixed snapshots.
Researchers can observe labeled cancer cells while an anticancer treatment is being studied and follow changes in cellular behavior over time. Imaging may reveal altered division, migration, invasion, or intracellular signaling after treatment. These time-resolved observations support evaluation of therapeutic effects and can help investigate treatment resistance by linking molecular activity with persistent or changing tumor-cell behavior.
The technique connects fluorescently observed molecular activity with behavior at the tumor-cell level. By following events in living cells or tissues, researchers can relate intracellular signaling or structural changes to processes relevant to disease mechanisms. This connection supports studies that seek to interpret how cellular dynamics contribute to tumor progression, therapeutic effects, and resistance.