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Immunofluorescent kit for detecting circulating tumor cells HER2, ER and PR and application of immunofluorescent kit

A tumor cell, immunofluorescence detection technology, applied in biological testing, fluorescence/phosphorescence, measurement devices, etc., can solve problems such as easy degradation of RNA and inability to identify expression status

Inactive Publication Date: 2019-01-11
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 the expression status of HER2, ER, and PR in single cells cannot be identified.

Method used

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  • Immunofluorescent kit for detecting circulating tumor cells HER2, ER and PR and application of immunofluorescent kit
  • Immunofluorescent kit for detecting circulating tumor cells HER2, ER and PR and application of immunofluorescent kit
  • Immunofluorescent kit for detecting circulating tumor cells HER2, ER and PR and application of immunofluorescent kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0076] Materials: smears of negatively enriched blood samples, and BT-474 cell smears for control cells.

[0077] Experimental steps:

[0078] 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;

[0079] 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;

[0080] 3. Absorb the excess liquid on the slide, add CYPP for 5 minutes, wash the slides with CYP1 as above for 3 minutes × 1 time; absorb excess liquid, add 200 μl of ice acetone:methanol (7:3) for 5 minutes, and wash the slides with CYP1 for 3 minutes × 3 times , to absorb excess water;

[0081] 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, ER antibody, PR antibody and CD45 antibody, and inc...

Embodiment 2

[0090] Materials: 1 tube of appropriate amount of anticoagulated blood, which is enriched by membrane filtration and then detected for protein

[0091] Experimental steps:

[0092] 1. Take an appropriate amount of peripheral blood and put it into a blood collection tube containing anticoagulant, and shake it slightly to mix.

[0093] 2. Add the suspension to the membrane filtration separation tumor cell technology device, and slowly pass through the filter and the filter membrane.

[0094] 3. After the filtration is completed, continue to add 50mI0.01MPBS 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;

[0095] 4. Fix the cells on the filter membrane;

[0096] 5. Perform the same operation as in Example 1 to detect the protein.

Embodiment 3

[0098] Materials: 1 tube of appropriate amount of anticoagulated blood, which is enriched by microfluidic method and then detected for protein

[0099] Experimental steps:

[0100] 1. The appropriate amount of blood drawn is enriched using microfluidic chips of various principles.

[0101] 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 an immunofluorescent kit for detecting circulating tumor cells HER2, ER and PR and an application of the immunofluorescent kit. A detection principle of a method disclosed bythe invention is as follows: firstly, enriching circulating tumor cells and other rare cells, and then, detecting the expression of target proteins in the enriched cells in cells by using an immunofluorescene detection method according to an antigen-antibody reaction principle. According to the kit, a common antigen CD45 of a target cell and a leukocyte is fluorescently labeled, and the cells withpositive target proteins and negative CD45 are screened, so that the circulating tumor cells with positive specific proteins in the blood are interpreted and counted.

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 human epidermal growth factor receptor 2 (HER2), estrogen receptor (ER), progesterone receptor (PR) protein immunofluorescence The 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 survival time of patients...

Claims

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Application Information

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IPC IPC(8): G01N33/574G01N33/68G01N33/533G01N21/64
CPCG01N21/6428G01N33/533G01N33/57496G01N33/6803G01N2800/7028
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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