Kit for detecting HER2 different sites, ER, and PR of circulating tumor cells and application thereof

A technology of tumor cells and sites, applied in measurement devices, instruments, fluorescence/phosphorescence, etc., can solve the problems of inability to identify the expression state and easy degradation of RNA

Inactive Publication Date: 2019-01-18
北京莱尔生物医药科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In this study, reverse transcription PCR was used to detect HER2, ER, and PR. RNA is easily degraded, and it is impossible to identify different sites of HER2 and the expression status of ER and PR in single cells.

Method used

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  • Kit for detecting HER2 different sites, ER, and PR of circulating tumor cells and application thereof
  • Kit for detecting HER2 different sites, ER, and PR of circulating tumor cells and application thereof
  • Kit for detecting HER2 different sites, ER, and PR of circulating tumor cells and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0077] Materials: smears of negatively enriched blood samples, and BT-474 cell smears for control cells.

[0078] Experimental steps:

[0079] 1. Draw 3.5ml of peripheral blood into an ACD (sodium citrate) anticoagulant tube. use The human peripheral blood leukocyte depletion kit negatively enriches tumor cells and fixes them on glass slides;

[0080] 2. Wash slides with CYP1 for 3 minutes x 3 times, 100-150 μL each time, to ensure that the entire sample area is covered;

[0081] 3. Absorb the excess liquid on the slide, add CYPP for 5 minutes, CYP1 as above and wash the slides for 3 minutes x 1 time; absorb excess liquid, add 200 μl ice acetone:methanol (7:3) for 5 minutes, CYP1 wash the slides for 3 min x 3 times , to absorb excess water;

[0082] 4. Add 100-150 μl of blocking solution to block at room temperature for 25-30 minutes. Absorb excess blocking solution, add 100 μl of diluted HER2 antibody-1, HER2 antibody-2, ER antibody, PR antibody and CD45 antibody, and i...

Embodiment 2

[0089] Materials: 1 tube of appropriate amount of anticoagulated blood, which is enriched by membrane filtration and then detected for protein

[0090] Experimental steps:

[0091] 1. Take an appropriate amount of peripheral blood and put it into a blood collection tube containing anticoagulant, and shake it slightly to mix.

[0092] 2. Add the suspension to the membrane filtration separation tumor cell technology device, and slowly pass through the filter and the filter membrane.

[0093] 3. After the filtration is completed, continue to add 50ml of 0.01M PBS to the membrane filtration device, wash the cell suspension attached around the tube wall into the membrane filtration device, and let it pass through the filter and membrane;

[0094] 4. Fix the cells on the filter membrane;

[0095] 5. Perform the same operation as in Example 1 to detect the protein.

Embodiment 3

[0097] Materials: 1 tube of appropriate amount of anticoagulated blood, which is enriched by microfluidic method and then detected for protein

[0098] Experimental steps:

[0099] 1. The appropriate amount of blood drawn is enriched using microfluidic chips of various principles.

[0100] 2. After enrichment, the samples were subjected to protein immunofluorescence detection.

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PUM

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Abstract

The invention relates to a kit for detecting HER2 different sites, ER, and PR of circulating tumor cells and an application thereof. The detection principle of the method of the invention is as follows: firstly, enriching circulating tumor cells and other rare cells, and then detecting expression of a target protein in cells in the enriched cells by an immunofluorescence detection method, according to the principle of antigen-antibody reaction. The kit is capable of fluorescently labeling target cells and leukocyte common antigen CD45, and screening target protein-positive and CD45-negative cells, thereby interpreting and counting specific protein-positive circulating tumor cells in the blood.

Description

Technical field: [0001] The invention relates to a medical detection method, in particular to the detection of circulating tumor cells in blood, in particular to different sites of human epidermal growth factor receptor 2 (HER2), estrogen receptor (ER), progesterone receptor (PR) The protein immunofluorescence detection kit is suitable for detecting HER2, ER, and PR antigens in different body fluid samples, and also relates to the use of the kit in detecting HER2, ER, and PR. Background technique: [0002] At present, the incidence of cancer is relatively high, which seriously threatens human health. Some cancers are difficult to detect early and are often diagnosed at an advanced stage, such as lung cancer, colorectal cancer, ovarian cancer, and pancreatic cancer. Correspondingly, the 5-year survival rate of advanced cancer is significantly lower than that of early-stage cancer, and early detection, early diagnosis and precise treatment can significantly prolong the surviv...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/64G01N33/574
CPCG01N21/6486G01N33/57484G01N2021/6495G01N2333/70535G01N2333/70528G01N2333/70557
Inventor 郭素杰郭志敏樊晓婷
Owner 北京莱尔生物医药科技有限公司
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