immunostaining
Immunostaining is a method of confirming the localization of an antigen expression using an antibody. An antibody against an antigen is called a primary antibody. An antibody against the primary antibody that is labeled with a fluorescent dye or an enzyme such as peroxidase or alkaline phosphatase is called a secondary antibody. Tissues and cells are treated with the primary antibody, and then with the secondary antibody. The primary antibody may be labeled with a fluorescent dye. In this case, a secondary antibody is not required. Some antibodies can recognize live cells, but in general, immunostaining is performed after fixing the cells. When the antigen is expressed in the cytoplasm, the cell must be permeabilized. In addition, depending on the immobilization method, activation treatment may be required because the structure of the antigen changes.
When performing immunostaining of cultured cells, the cells are usually fixed, but live cells are also stained. Generally, dishes or plates used for cell culture may be used, but a large vessel such as the 6 well require a large amount of antibody. In addition, when performing observation using confocal, plastic culture vessels are not suitable, so chamber slides or glass bottom plates are used.
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