Figure 1. Antigen retrieval and antigen destruction immunohistochemistry methods.<\/figcaption><\/figure>\nHigh Temperature and High Pressure Antigen Retrieval Method<\/span><\/p>\nAfter deparaffinizing the tissue section into water, pour the 0.01mol\/L citrate buffer solution (pH 6.0, antigen retrieval solution) 1000-1500mL into the electric pressure cooker, and place the tissue section on the high temperature plastic section rack.\u00a0Gently put it into the electric pressure cooker, the tissue section must be under the liquid surface, close the pressure valve of the pan, plug in the power source, and adjust the timer to 180s.\u00a0When the pressure of the electric pressure cooker reaches the expected value, it will be maintained for 2 minutes.\u00a0After that, turn off the power and open the air jet valve.\u00a0After the air deflation is complete, open the lid of the electric pressure cooker to cool to room temperature.\u00a0Rinse with distilled water 3 times, and then wash with PBS solution 3 times, 3min each time.\u00a0After washing, add the primary antibody diluted 1:100, incubate at 37\u00b0C for 2h, wash out the unbound antibody with PBS solution, and repeat the washing 3 times, 3 min each time.\u00a0After washing is completed, add the secondary antibody, incubate at 37\u00b0C for 30 min, and then continue to wash with PBS solution for 3 times, 3 min each time.\u00a0Finally, DAB was added for color development, and hematoxylin counterstaining was performed.\u00a0After dehydration, the sections were sealed with gum and finally observed.<\/p>\n
Microwave Antigen Retrieval<\/p>\n
After the tissue section is deparaffinized and put into water, the antigen retrieval solution 0.01mol\/L citrate buffer solution (pH value 6.0) 100-150mL is poured into the microwave box, and the tissue section is placed on the high-temperature-resistant plastic section rack.\u00a0Put it into the microwave box, close the lid, put it in the microwave oven, first treat it on high heat for 5 minutes, and continue to treat it on medium heat for 10 minutes to keep the liquid temperature in the container at about 98 \u2103.\u00a0Stop the fire, take out the container, and cool to room temperature.\u00a0Rinse with distilled water 3 times, and then wash with PBS solution 3 times, 3min each time.\u00a0After washing, add the primary antibody diluted 1:100, incubate at 37\u00b0C for 2h, wash out the unbound antibody with PBS solution, and repeat the washing 3 times, 3 min each time.\u00a0After washing is completed, add the secondary antibody, incubate at 37\u00b0C for 30 min, and then continue to wash with PBS solution for 3 times, 3 min each time.\u00a0Finally, DAB was added for color development, and hematoxylin counterstaining was performed.\u00a0After dehydration, the sections were sealed with gum and finally observed.<\/p>\n
Boiling Antigen Retrieval Method<\/p>\n
After the tissue section is deparaffinized and put into water, the antigen retrieval solution 0.01mol\/L citrate buffer solution (pH value 6.0) 1000-1500ml is poured into the boiling pot, and the tissue section is placed on the high temperature plastic section rack. Gently put it into the boiling pot, the tissue section must be below the liquid surface, count for 30 minutes after boiling, turn off the heat, and cool to room temperature. Rinse with distilled water 3 times, and then wash with PBS solution 3 times, 3min each time. After washing, add the primary antibody diluted 1:100, incubate at 37\u00b0C for 2h, wash out the unbound antibody with PBS solution, and repeat the washing 3 times, 3 min each time. After washing is completed, add the secondary antibody, incubate at 37\u00b0C for 30 min, and then continue to wash with PBS solution for 3 times, 3 min each time. Finally, DAB was added for color development, and hematoxylin counterstaining was performed. After dehydration, the sections were sealed with gum and finally observed.<\/p>\n
The heated antigen retrieval (AR) technology is an important milestone in the history of immunohistochemistry.\u00a0The principles of the three types of antigen retrieval are roughly the same.\u00a0Through the action of high-frequency electromagnetic waves or high temperature and high pressure, the aldehydes formed by the treatment of the cross-linkable fixative formaldehyde in the tissue The chain opens to fully expose the antigen and achieve the purpose of repair.\u00a0By comparing the pretreatment of three different antigen retrieval methods, it is found that the pretreatment using the high temperature and high pressure antigen retrieval method is significantly better than microwave retrieval.\u00a0Because high temperature and high pressure can overcome batch differences and “hot spots” in the use of microwave ovens, it is believed that high temperature and high pressure are more uniform than other heating methods.\u00a0When the electric pressure cooker is fully pressured at 15 Psi, the temperature can reach about 120 \u2103.\u00a0This kind of temperature shows obvious superiority in the de-masking effect of some nuclear antigens.\u00a0More antigens can be exposed, and some false negative and weakly positive specimens can be positively expressed.\u00a0The positive location of the tissue is clear, the background color is light, and the non-specific coloration is weak.\u00a0It can improve the positive detection rate of antigen and antibody, simple operation and short time consumption.\u00a0The positive expression rate is stronger than the microwave antigen retrieval method;\u00a0while the microwave antigen retrieval method, due to the narrow heating range of the microwave oven, the tissue slices receive the microwave radiation unevenly and the amount is small, resulting in the inconsistency of the processing intensity of the tissue slices in different regions.\u00a0Edge effect, resulting in the appearance of different shades of color in different areas.\u00a0At the same time, microwave oven heat repair is prone to the phenomenon of dry slices caused by dry buffer solution, and many disadvantages such as longer repair time and easy tissue peeling;\u00a0while boiling antigen retrieval, the antigen retrieval time is long, the temperature is not easy to control, and it is easy to heat unevenly can lead to poor recovery of the antigen activity of the tissues, thereby affecting the results of immunohistochemical reactions.\u00a0Therefore, the high temperature and high pressure antigen retrieval method is significantly better than the microwave antigen retrieval method and the boiling antigen retrieval method.<\/p>\n","protected":false},"excerpt":{"rendered":"
Immunohistochemistry technology has become an indispensable and important method in the routine work and scientific research of the pathology laboratory.\u00a0It is widely used in clinical pathological diagnosis to help evaluate the degree of tumor proliferation and staging, and has a guiding role in the prognosis and treatment of patients. .\u00a0However, in daily work, pathological specimens […]<\/p>\n","protected":false},"author":1,"featured_media":0,"comment_status":"closed","ping_status":"closed","sticky":false,"template":"","format":"standard","meta":[],"categories":[59],"tags":[80],"aioseo_notices":[],"_links":{"self":[{"href":"https:\/\/www.creative-diagnostics.com\/blog\/index.php\/wp-json\/wp\/v2\/posts\/1303"}],"collection":[{"href":"https:\/\/www.creative-diagnostics.com\/blog\/index.php\/wp-json\/wp\/v2\/posts"}],"about":[{"href":"https:\/\/www.creative-diagnostics.com\/blog\/index.php\/wp-json\/wp\/v2\/types\/post"}],"author":[{"embeddable":true,"href":"https:\/\/www.creative-diagnostics.com\/blog\/index.php\/wp-json\/wp\/v2\/users\/1"}],"replies":[{"embeddable":true,"href":"https:\/\/www.creative-diagnostics.com\/blog\/index.php\/wp-json\/wp\/v2\/comments?post=1303"}],"version-history":[{"count":5,"href":"https:\/\/www.creative-diagnostics.com\/blog\/index.php\/wp-json\/wp\/v2\/posts\/1303\/revisions"}],"predecessor-version":[{"id":1309,"href":"https:\/\/www.creative-diagnostics.com\/blog\/index.php\/wp-json\/wp\/v2\/posts\/1303\/revisions\/1309"}],"wp:attachment":[{"href":"https:\/\/www.creative-diagnostics.com\/blog\/index.php\/wp-json\/wp\/v2\/media?parent=1303"}],"wp:term":[{"taxonomy":"category","embeddable":true,"href":"https:\/\/www.creative-diagnostics.com\/blog\/index.php\/wp-json\/wp\/v2\/categories?post=1303"},{"taxonomy":"post_tag","embeddable":true,"href":"https:\/\/www.creative-diagnostics.com\/blog\/index.php\/wp-json\/wp\/v2\/tags?post=1303"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}
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