Fluorescence imaging, or fluorescence, is a widely used method in cellular studies that helps monitor cellular processes and drug actions in living organisms. In fluorescence microscopy, fluorescence is used instead of scattering, absorption, or reflection of light to study the properties of biological or non-biological samples. Fluorescence microscopy refers to any type of microscopy that utilizes fluorescence, whether it be simple microscopy or confocal microscopy. In this technique, light of a specific wavelength is directed onto the sample, resulting in the sample emitting light at a longer wavelength. This emitted light is significantly weaker than the incident light, and for this reason, it is separated through filters. Fluorescence is a type of luminescence where the emission of light occurs after the sample is exposed to electromagnetic waves, and it is referred to as a fluorophore.
Typically, conventional light sources such as mercury and xenon lamps are used in these microscopes. However, some microscopes utilize more complex sources, such as LEDs and laser light. The figure below illustrates the components of fluorescence microscopes, including the light source, filters, and mirrors.
The emission of each color from the sample leads to the production of a color image. Various colors can be generated by combining the emitted colors. For imaging with this microscope, the sample must be fluorescent. There are various techniques available for fluorescently labeling samples. Labeling samples with fluorescent dyes or expressing fluorescent proteins in biological samples are the most common methods. By selectively staining proteins, it is possible to study cellular processes. These dyes can bind to specific parts of molecules within the cell. Among the most well-known dyes are nucleic acid dyes such as DAPI and Hoechst, which are used for imaging the nuclei of cells due to their affinity for DNA. Peptides, toxins, and other drugs are additional dyes that have been derivatized with fluorescent materials. Immunofluorescence is another widely used technique where antibodies are utilized for highly selective binding to antigens for labeling proteins and other molecules in cells. An example of images captured with this microscope is shown in the figure below.
Fluorescence imaging allows researchers to observe and study the dynamics of protein and gene expression and other molecular interactions and processes within cells, making it an invaluable tool in modern research. In the Max imaging facility, fluorescence imaging and DAPI and Hoechst staining, along with other staining techniques, are performed with the highest quality.