Quick immune histochemical detection reagent for milk gland cancer lymph node metastasis and its detecting method

Abstract

The present invention is one kind of reagent for quick immune histohcemical detection of lymphatic metastasis of mammary cancer and corresponding detecting method. Several kinds of directly marked antibody capable of distinguishing metastatic cancer cell and inherent lymph node cell are mixed into reagent and one-step reacted to stain the metastatic cancer cell in lymph node specifically. The present invention is used in the fast pathological diagnosis of early lymphatic metastasis of mammary cancer, and has raised sensitivity, high accuracy and high specificity.

Description

technical field

[0001] The invention relates to a reagent for lymph node detection of breast cancer, in particular to a rapid immunohistochemical detection reagent for lymph node metastasis of breast cancer. The invention also relates to an immunohistochemical detection method, which is applied to rapid pathological diagnosis of early breast cancer lymph node. Background technique

[0002] Breast cancer lymph node dissection is an essential principle of breast surgery in the past. In recent years, with the advancement of breast disease screening and diagnostic technology, the detection rate of early breast cancer has been increasing. Because the majority of early breast cancers do not have axillary lymph node metastasis, if axillary lymph node dissection is performed uniformly, patients without axillary lymph node metastasis will not benefit from it but will cause unnecessary damage. Therefore, accurate prediction of the presence or absence of axillary...

Examples

Embodiment 1

[0046] [Example 1] Materials: 1. positive marker CK19, EMA, MUC1, mammaglobin antibody; 2. negative control marker VEGFR-3, CD31, CD68 antibody, LCA antibody; 3. horseradish peroxidase (HRP); 4. dextran (Dextron); ⑤ Chromogenic oxidation reaction reagent (0.3%H 2 o 2 ); ⑥ Chromogenic agent (3.3-diaminobenzidine, DAB); ⑦ Chromogenic buffer (0.005MolTris-Hcl).

[0047] Firstly, the working concentration of the antibody was determined by ELISA method.

[0048] Specific method: first dilute the antibody with 0.05M PH=9.6 coating buffer to about 10 μg / ml, add 0.1ml to the polystyrene plate well, overnight at 4°C, and wash 3 times with washing buffer the next day. Enzyme-labeled antibody was diluted sequentially with 1% BSA-PBS solution to 1:100, 1:200, 1:400, 1:800, 1:1600..., respectively added to the reaction wells, two wells for each dilution, 0.1 ml, incubated at 37°C for 1 hour and washed. Then add substrate solution, 0.1ml per well, 37°C for 10-30 minutes. Take 2M H 2 S...

Embodiment 2

[0054] [Example 2] Materials: ① positive marker CK19, EMA, MUC1, mammaglobin antibody; ② negative control marker VEGFR-3, CD31, CD68 antibody, LCA antibody; ③ fluorescent pigment; ④ dextran (Dextron); ⑤ color development Oxidation reagent (0.3% H 2 o 2 ); ⑥ Chromogenic agent (3.3-diaminobenzidine, DAB); ⑦ Chromogenic buffer (0.005MolTris-Hcl).

[0055] Firstly, the working concentration of the antibody was determined by ELISA method.

[0056] Specific method: first dilute the antibody with 0.05M PH=9.6 coating buffer to about 10 μg / ml, add 0.1ml to the polystyrene plate well, overnight at 4°C, and wash 3 times with washing buffer the next day. Enzyme-labeled antibody was diluted sequentially with 1% BSA-PBS solution to 1:100, 1:200, 1:400, 1:800, 1:1600..., respectively added to the reaction wells, two wells for each dilution, 0.1 ml, incubated at 37°C for 1 hour and washed. Then add substrate solution, 0.1ml per well, 37°C for 10-30 minutes. Take 2M H 2 SO 4 , 0.05ml t...

technical field

[0001] The invention relates to a reagent for lymph node detection of breast cancer, in particular to a rapid immunohistochemical detection reagent for lymph node metastasis of breast cancer. The invention also relates to an immunohistochemical detection method, which is applied to rapid pathological diagnosis of early breast cancer lymph node. Background technique

[0002] Breast cancer lymph node dissection is an essential principle of breast surgery in the past. In recent years, with the advancement of breast disease screening and diagnostic technology, the detection rate of early breast cancer has been increasing. Because the majority of early breast cancers do not have axillary lymph node metastasis, if axillary lymph node dissection is performed uniformly, patients without axillary lymph node metastasis will not benefit from it but will cause unnecessary damage. Therefore, accurate prediction of the presence or absence of axillary...

Examples

Embodiment 1

[0046] [Example 1] Materials: 1. positive marker CK19, EMA, MUC1, mammaglobin antibody; 2. negative control marker VEGFR-3, CD31, CD68 antibody, LCA antibody; 3. horseradish peroxidase (HRP); 4. dextran (Dextron); ⑤ Chromogenic oxidation reaction reagent (0.3%H 2 o 2 ); ⑥ Chromogenic agent (3.3-diaminobenzidine, DAB); ⑦ Chromogenic buffer (0.005MolTris-Hcl).

[0047] Firstly, the working concentration of the antibody was determined by ELISA method.

[0048] Specific method: first dilute the antibody with 0.05M PH=9.6 coating buffer to about 10 μg / ml, add 0.1ml to the polystyrene plate well, overnight at 4°C, and wash 3 times with washing buffer the next day. Enzyme-labeled antibody was diluted sequentially with 1% BSA-PBS solution to 1:100, 1:200, 1:400, 1:800, 1:1600..., respectively added to the reaction wells, two wells for each dilution, 0.1 ml, incubated at 37°C for 1 hour and washed. Then add substrate solution, 0.1ml per well, 37°C for 10-30 minutes. Take 2M H 2 S...

Embodiment 2

[0054] [Example 2] Materials: ① positive marker CK19, EMA, MUC1, mammaglobin antibody; ② negative control marker VEGFR-3, CD31, CD68 antibody, LCA antibody; ③ fluorescent pigment; ④ dextran (Dextron); ⑤ color development Oxidation reagent (0.3% H 2 o 2 ); ⑥ Chromogenic agent (3.3-diaminobenzidine, DAB); ⑦ Chromogenic buffer (0.005MolTris-Hcl).

[0055] Firstly, the working concentration of the antibody was determined by ELISA method.

[0056] Specific method: first dilute the antibody with 0.05M PH=9.6 coating buffer to about 10 μg / ml, add 0.1ml to the polystyrene plate well, overnight at 4°C, and wash 3 times with washing buffer the next day. Enzyme-labeled antibody was diluted sequentially with 1% BSA-PBS solution to 1:100, 1:200, 1:400, 1:800, 1:1600..., respectively added to the reaction wells, two wells for each dilution, 0.1 ml, incubated at 37°C for 1 hour and washed. Then add substrate solution, 0.1ml per well, 37°C for 10-30 minutes. Take 2M H 2 SO 4 , 0.05ml t...

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