Issues to be noted in antigen repair The following issues should also be noted during operation: 1. Natural cooling should be paid attention to after heat treatment. 2. Prevent the chips from drying during heat treatment. 3. The appropriate antigen repair method should be selected according to different antibodies. 4. The temperature and time used for the detection of the same batch of antigen should be consistent. 5. Pay attention to the changes of morphological structure (generally the cytoplasm will be obviously damaged after high temperature repair, which has a greater impact on the nucleus, and there will be nuclear rupture and hematoxylin to light staining, etc .; and the cells that are hydrolyzed by enzymes The plasma damage varies according to the length of digestion, but the nuclear damage is lighter). 6. If the antigen repair cannot be achieved in the commonly used buffers, or the localization of the antigen after the repair changes, you can use some uncommon buffers or add some chelating agents, such as EDTA to improve the repair of certain antigens . There are many digestive enzymes used in immunoenzymatic histochemical antigen repair. The type of enzyme selected, the concentration used, pH, digestion time, and temperature all depend on the tissue fixation. The nature of the detected antigen varies with the type of tissue. , Should be determined through preliminary experiments. The principle of using enzyme digestion: trypsin and proteinase K are generally used for intracellular antigens, such as Keratin, CEA and GFAP. Pepsin is mainly used for the detection of interstitial antigens. Such as fibronectin, Laminin and various types of collagen. In general, the digestion time is proportional to the length of fixed tissue sections. On the premise that no results can be obtained after enzyme digestion and heat repair, the method of combining antigen repair and enzyme digestion sometimes results in more satisfactory results. The combination of proteinase K and microwave is generally used, but it should be noted that no matter what kind of antigen detection may occur in the nucleus, a false positive reaction. Therefore, it must be compared.
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