A rapid detection method for rare cells based on magnetic microbeads without transfer in low-field NMR

A rare cell and magnetic microbead technology, applied in the field of molecular biology detection, can solve problems such as long centrifugation time, and achieve the effect of facilitating comparison and quality control, strong controllability and accurate detection

Active Publication Date: 2017-01-11
SHANGHAI MAJORBIO BIO PHARM TECH
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  • Abstract
  • Description
  • Claims
  • Application Information

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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  • A rapid detection method for rare cells based on magnetic microbeads without transfer in low-field NMR
  • A rapid detection method for rare cells based on magnetic microbeads without transfer in low-field NMR
  • A rapid detection method for rare cells based on magnetic microbeads without transfer in low-field NMR

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Experimental program
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Embodiment 1

[0036] (1) Sample preparation

[0037] Preparation of samples to be tested:

[0038] 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;

[0039] 1.2 Ficoll density gradient centrifugation method to obtain candidate mixed cell populations

[0040] 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.

[0041] 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).

[0042] 3) After centrifug...

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Abstract

The invention relates to the technical field of molecular biology, and discloses a sample-transfer-free and low-field nuclear magnetic resonance (NMR) rapid rare cell detection method based on magnetic microspheres. The method comprises the steps of (1) respectively preparing a standard sample and a sample to be detected of target rare cell suspension liquid; (2) feeding a NMR contrast agent coupled with a target rare cell specific expression antibody into the standard sample and the sample to be detected respectively, evenly mixing, and then respectively carrying out relaxation time measurement on the standard sample and the sample to be detected by a low-field NMR analyzer immediately; (3) carrying out immune response enrichment on the NMR contrast agent and rare cells in the samples, and then carrying out relaxation time measurement on the standard sample and the sample to be detected by the low-field NMR analyzer again; and (4) calculating the change of relaxation time, and drawing a standard curve to obtain the content of target rare cells in the sample to be detected. After the sample-transfer-free and low-field NMR rapid rare cell detection method is adopted, the rare cells in a biological fluid sample can be simply, conveniently and rapidly detected with high specificity and sensitivity; sample transfer does not occur in the two-time detection, so that high accuracy rate can be obtained.

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 Patents(China)
IPC IPC(8): G01N24/08G01N1/38
Inventor 张祥林
Owner SHANGHAI MAJORBIO BIO PHARM TECH
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