Immunofluorescence is one of many techniques in biomedical research and diagnosis. It uses the sensitivity and selectivity of fluorescence to analyze biological tissues. The basic reaction of immunology is antigen-antibody reaction. when antigen-antibody reaction occurs, as long as one factor is known, the other factor can be determined, due to the high specificity of the reaction. Immunofluorescence technology is to label the fluorescent pigment that does not affect the activity of antigen and antibody on the antibody (or antigen), combine it with its corresponding antigen (or antibody), and show a specific fluorescence reaction under the fluorescence microscope. Hence, immunofluorescence technique can be used to detect proteins and other biomolecules in cells and tissues.

Immunofluorescence of cancer cells.Figure 1.Immunofluorescence of cancer cells.

Scientists can use immunofluorescence technology to carry out various laboratory tests and observations. Each test and observation provide a new perspective for the samples currently being analyzed. Researchers can apply this technique to fixed or fresh samples to provide diversified analysis and accurate results. CD BioSciences can provide you with customized services based on immunofluorescence technology. If you have any needs, please feel free to contact us.

Detection Techniques for Immunofluorescence

Fluorescence Microscopy

The basic function of fluorescence microscope is to transfer the excitation energy to the fluorescent dye in the sample to be studied, separate the weak emission fluorescence from the excitation light, and send it to the detector, where the contrast image is generated. The specificity and high sensitivity of fluorescence microscopy allow the study of cellular and subcellular structures in biological tissues with high resolution. It provides three-dimensional spatial information and temporal changes of spatial information in living cells.

Epifluorescence microscopyFigure 2.Epifluorescence microscopy (Fritschy, et al., 2001).

Laser Scanning Confocal Microscopy

Confocal microscope is a mature tool for biomedical research, which can provide improved optical microscopic imaging of fluorescent dye-labeled cells. Its main application is to perform high-resolution analysis of subcellular components in three dimensions. The main difference from traditional epifluorescence microscopes is the imaging method. The former provides an image of the entire specimen, while the confocal microscope focuses on a light spot that scans the specimen line by line.

Flow Cytometry

Flow cytometry is a technique used to measure the physical or chemical properties of cells and other biological particles when they pass through a measuring device in a single row. The measurement of immunofluorescence is a special application of flow cytometry. It is particularly useful for cell sorting based on the presence or absence of fluorescent signals, and can precisely define and quantify heterogeneity in cell populations.

Flow cytometryFigure 3.Flow cytometry (Fritschy, et al., 2001).

Applications of Immunofluorescence

  • Detection of circulating autoantibodies
  • Detection of antigens in cells and cell membranes
  • Detection of immune complexes in biopsy material
  • Detection of circulating immune cells
  1. Fritschy, Jean‐Marc, and Wolfgang Härtig. "Immunofluorescence." e LS (2001).
  2. Odell, Ian D., and Deborah Cook. "Immunofluorescence techniques." The Journal of investigative dermatology 133.1 (2013): e4.

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