Circulating tumor cell detection method

Abstract

The invention discloses a circulating tumor cell detection method. The circulating tumor cell detection method is characterized in that highly heterogeneous circulating tumor cells are accurately marked through glucose metabolism engineering independent of tumor cell types, the marked circulating tumor cells are subjected to lossless capture through a biological orthogonal reaction, and the captured circulating tumor cells are subjected to nondestructive release through reversible reaction, so that accurate and nondestructive detection of the circulating tumor cells is realized.

Description

technical field [0001] The invention belongs to the technical field of detection methods for circulating tumor cells, in particular to a detection method for circulating tumor cells based on bio-orthogonal metabolic glycoengineering markers, that is, a method based on metabolic glycoengineering pathway markers, based on bio-orthogonal reaction capture, and based on reversible reaction release "Mark-capture-release" three-stage CTC detection. Background technique [0002] Circulating tumor cells (CTCs) are cancer cells that come off from the tumor site, enter and survive in the circulatory system. The presence of CTC is an early sign of cancer spread and metastasis. The detection of CTC is of great significance for early detection of tumor micrometastasis, evaluation of prognosis and efficacy, and individualized treatment of tumors. [0003] The difficulty and challenge of CTC detection lies in the small number and high heterogeneity of CTCs in the blood of cancer patients. ...

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Problems solved by technology

In principle, the most common detection method based on changes in the physical properties of CTCs is to use the difference in cell size of CTCs compared to white blood cells to sort CTCs and white blood cells through separation processes such as filtration and deterministic lateral displacement. The size of white blood cells coincides, so this method is suitable for the primary screening and enrichment of CTCs, and cannot achieve accurate detection; CTC detection based on tumor genes generally first lyses the sample cells, extracts genetic material, and then uses polymerase chain reaction (PCR) to detect Detect genetic material such as tumor-associated DNR / RNA in it, the cell lysis operation cannot perform cell counting and capture complete CTC cells for downstream application expansion; CTC detection based on cell surface specific antigen is currently the mainstream detection method, mainly using CTC Specific antigens present on the surface, immobilizing the corresponding antibodies on the substrate of the detection device, and capturing CTCs through immune binding. In the process of entering the blood circulation from the primary site of the tumor, it undergoes epithelial-mesenchymal transition (EMT), the surface EpCAM level decreases, the invasion ability increases, and enters the blood circulation, while there is no EpCAM molecule on the surface of leukocytes, so The EpCAM antibody can be used as a capture molecule, immobilized on the surface of the detection device, and the antibody is used to immunologically bind EpCAM on the surface of CTC to capture and isolate CTC. The immune reaction may lead to the apoptosis of the captured CTC, and as mentioned above, CTC is highly heterogeneous According to the classification of its surface marker molecules, there are more than ten subtypes. One or several cell surface antigens can only cover part of the CTC. Although the CTC detection of cell surface marker antigens can provide cell counting results, it is limited in the accuracy of the detection results and the non-destructiveness of the capture process

[0005] The main reason for the lack of accuracy of current CTC detection and the inability to capture / release CTCs non-destructively is that CTCs are highly heterogeneous (heterogeneicty), that is, the differences and diversity of CTCs in different tumor individuals in terms of genes and phenotypes, even in the same patient. CTCs also often show heterogeneity, so a single physical or biochemical characteristic change index only covers part of the CTCs, which limits the accuracy and non-destructiveness of current CTC detection methods

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