Fluorescence is a molecular phenomenon where a substance or specimen radiates light energy when it’s stuck with light from another source. Some of the light energy radiating during this incident is absorbed by the sample. This happens because the emitted light is typical of lower energy or lower wavelength than the light source.
Many organic and non-organic substances exhibit fluorescence features. This concept is the factor behind the fluorescence microscope, one of the primary tools used by scientists worldwide to study different matter types. If you’re looking to use this type of microscope for research purposes or in an academic setting, you can buy a fluorescence microscope at microscopeinternational.com.
You can also read more below to learn more about its working principle and various uses.
Working Principle Of A Fluorescence Microscope
A fluorescence microscope uses fluorescent light to observe living matter at the point where fluorescent dyes are present. Fluorescent light gives off energy as the atoms emit electrons, thus creating a glow when excited. Fluorescent microscopes utilize tiny and highly focused fluorescent lights to magnify thousands of times than that of normal vision. This process enables scientists to look deep into the microenvironment of most specimens.
To further understand the process, below is the working principle of the microscope:
- Most specimens with cellular components are colorless, so the sample can’t be distinguished immediately under a standard microscope. To make it visible, a researcher needs to stain the specimen with dyes.
- The fluorescent dyes, also known as fluorophores, help absorb excitation light at a given wavelength and then emit light with a longer wavelength to make it visible.
- The emission light will be filtered from the excitation light so the researcher can locate the fluorophores.
- The image produced is based on the second light source, which is used to excite the specimen’s dye.
Components Of A Fluorescence Microscope
A typical fluorescence microscope has the following features:
- Fluorescent dyes: It’s a chemical compound that can re-emit light upon excitation. It contains several combined aromatic groups or cyclic molecules with pi bonds. Many stains are designed for different biological molecules.
- Light Source: The microscope uses four main types of light sources like mercury-vapor lamps, xenon arc lamps with an excitation filter, lasers, and high-power LEDs.
- Excitation Filter: This is typically a bandpass filter where wavelengths absorbed by the dye passes. It’s designed to minimize the excitation of the other sources of fluorescence.
- Dichroic Mirror: It’s a thin film filler used to selectively pass light of a small range of colors while reflecting some unwanted colors.
- Emission Filter: It’s a bandpass filter whose purpose is to let the fluorophore’s emitted wavelengths pass, and blocks any unwanted kind of light outside of the band, like the excitation light. By blocking unwanted light, the specimen can be seen with a dark background.
Uses Of Fluorescence Microscope
Fluorescence Microscopes can be used in research, various fields of medicine, and even in an academic setting to help children learn more about cellular structures.
Fluorescent images are used in biology, chemistry, molecular biology, physiology, and imaging. The techniques used vary according to the intensity of fluorescence microscopy:
- Low-intensity fluorescence microscopy is used for elemental analysis and the study of metabolite functions in living organisms.
- High-intensity fluorescence microscopy is used for detailed biochemical imaging of live or dead cells, for in situ hybridization, and for detecting and localizing gene expression patterns.
Here are also some of the typical applications of fluorescence microscopy you should be aware of:
- Cell Labeling
Fluorescence Microscopes are used to identify and label cell structures with fluorescent dyes. Identifying cell structures are useful in different medical applications. Dyes with high specificity are used in medical diagnostics. Researchers, for instance, use this for analyzing and also tracking viral DNA found in cells. The information is then connected to cellular anti-viral defenses. The information gathered from this procedure is helpful for the future of viral analysis work, as well as categorization.
- Live Cell Imaging
Live cell imaging is where researchers observe the dynamic processes happening inside the cells, tissues, or small organisms. It’s an impressive study where you can see life in motion. Compared to observing fixed cells, live cells provide detailed information on the different cell changes. Many experts have already used this technique to study cell division and the transformation of organelles.
- Protein Characterization
Different dynamic protein interactions are critical for various biological processes. Proteins act as catalysts for many metabolic reactions or responses to stimuli. The techniques used in fluorescence microscopy help provide insights on how protein characterization in cells results in developmental problems and disease transmission.
With additional medical research and studies on vaccines and medicines, researchers need to study live cells and their processes. Fluorescent light microscopes have several advantages over conventional light microscopes, making them a handier tool. It’s the reason why this microscope is widely used for many biological and medical applications. Studying live cells, their processes, and characteristics helps experts learn more about diseases and eradicate them.