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Magnetic-bead-based low field NMR (nuclear magnetic resonance) rare cell detection method

A technology of rare cells and magnetic microbeads, which is applied in the field of molecular biology detection, can solve the problems of long centrifugation time, etc., and achieve the effect of long detection time, complicated operation and difficult repeatability

Active Publication Date: 2014-10-29
上海美吉医学检验有限公司
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AI Technical Summary

Problems solved by technology

The disadvantages of this method are: ① longer centrifugation time; ② need to prepare inert gradient medium solution

Method used

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  • Magnetic-bead-based low field NMR (nuclear magnetic resonance) rare cell detection method
  • Magnetic-bead-based low field NMR (nuclear magnetic resonance) rare cell detection method

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Embodiment 1

[0037] (1) Sample preparation

[0038] Preparation of samples to be tested:

[0039] 1.1 Obtain an anticoagulated blood sample (˜10 ml) from a breast cancer model mouse, which contains a mixed cell population suspected to contain target rare cells. Collected specimens should be processed the same day! Keeping time at room temperature away from light should not exceed 24 hours;

[0040] 1.2 Ficoll density gradient centrifugation method to obtain candidate mixed cell populations

[0041] 1) Add an appropriate amount of cell layering solution (prepared from Ficoll solution) to the bottom of sterile centrifuge tube A, then dilute the above anticoagulant test sample with PBS solution (2:1), and gently add On top of the layered liquid, the two form a clear interface.

[0042] 2) Centrifuge horizontally at 400×g for 30 minutes (the increase and decrease of the centrifuge speed should be uniform and stable to ensure the formation of a clear interface).

[0043] 3) After centrifug...

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Abstract

The invention relates to the technical field of molecular biology and discloses a magnetic-bead-based low field NMR (nuclear magnetic resonance) rare cell detection method which comprises the following steps: (1) a standard sample and a to-be-tested sample of a target rare cell suspension are respectively prepared; (2) a nuclear magnetic resonance contrast medium coupled with a specific expression antibody of target rare cells is respectively added, the nuclear magnetic resonance contrast medium is enriched in the rare cells in the samples by immunoreactions, and the unreacted nuclear magnetic resonance contrast medium is removed to obtain the nuclear magnetic resonance contrast medium-enriched standard sample and to-be-tested sample; (3) the relaxation time of the standard sample and the to-be-tested sample are respectively detected by use of a low field nuclear magnetic resonance analyzer, and sterile deionized water or a buffer solution is used as a blank control; and (4) the change amount of the relaxation time is calculated for drawing a standard curve to obtain the content of the target rare cells in the to-be-tested sample. The detection method can be used for convenient, fast, high specificity and high sensitivity detection of low content of rare cells in biological fluid samples.

Description

technical field [0001] The invention relates to molecular biology detection technology, in particular to a detection method for rare cells in biological fluid samples. Background technique [0002] Rare cells refer to some atypical cells in biological fluid samples (including blood, pleural effusion, ascites, urine, cerebrospinal fluid, etc.). has important guiding significance. Therefore, finding an accurate and rapid detection method for rare cells will become an urgent problem to be solved. However, the concentration of rare cells in biological fluids is very low, with a ratio of about 1:10 to non-target cells 7 , cannot be counted with traditional techniques, so a simple, accurate and fast method is urgently needed to solve this difficult problem. [0003] Due to the low content of circulating rare cells in body fluids, effective enrichment steps are required to identify and identify them in subsequent experiments. At present, the methods widely used in the detection...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N24/08G01N1/28
Inventor 张祥林
Owner 上海美吉医学检验有限公司
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