Specific lymphocyte content analysis method and device based on TCR/BCR high-throughput sequencing

A lymphocyte and analysis method technology, applied in the field of gene detection, can solve the problems of detection, high cost, and affecting the accuracy of cancer cells, etc., to achieve accurate reflection and good detection performance

Active Publication Date: 2021-02-05
深圳泛因医学有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] However, by flow cytometry detection, the content of a type of cells with the same surface differentiation antigen is detected, and each cell cannot be accurately distinguished
Moreover, the detection sensitivity of the current common flow cytometry can only reach 1/10000. For samples with a content lower than 1/10000, false negative detection results will be given.
In flow cytometry, data interpretation depends on the operator and the laboratory, and the degree of standardization is low
Most importantly, for cancerous lymphocytes, the expression of their surface antigens will change at different times of cancer occurrence and treatment, which will seriously affect the accuracy of cancer cell detection
[0006] For detection by RQ-PCR, it is necessary to design specific primers and quantitative probes for the TCR/BCR genes of different target cells. If there i

Method used

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  • Specific lymphocyte content analysis method and device based on TCR/BCR high-throughput sequencing
  • Specific lymphocyte content analysis method and device based on TCR/BCR high-throughput sequencing
  • Specific lymphocyte content analysis method and device based on TCR/BCR high-throughput sequencing

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Experimental program
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Effect test

Embodiment 1

[0053] Lymphohematopoietic tumor is a malignant disease that seriously endangers human health caused by malignant clonal proliferation of lymphocytes, including lymphocytic leukemia, myeloma and various lymphomas.

[0054] With the improvement of diagnosis and treatment technology, the diagnosis and treatment of lymphocytic tumors has achieved a qualitative leap. Minimal residual disease (MRD, Minimal residual disease) refers to a small amount of tumor cells remaining in the body after complete remission (CR, Complete Remission) after induction chemotherapy or after bone marrow transplantation. With the continuous improvement of chemotherapy, specific targeted therapy, and hematopoietic stem cell transplantation technology, the treatment effect of lymphocytic tumors is improving day by day. However, recurrence is still a problem that plagues the cure of this type of disease, and minimal residual disease is the main source of recurrence. Regular MRD detection for patients has i...

Embodiment 2

[0131] A specific lymphocyte content analysis kit based on TCR / BCR high-throughput sequencing, including: internal standard sequence, multiple PCR amplification primers and reagent components required for conventional high-throughput sequencing.

[0132] The internal standard sequence is the internal standard sequence of the coding gene of artificially synthesized TCR / BCR; the multiple PCR amplification primers are used for the amplification of the TCR / BCR gene, and the specific sequence is as SEQ ID NO.1-SEQ ID in Example 1 Shown in NO.56.

Embodiment 3

[0134] A specific lymphocyte content analysis device based on TCR / BCR high-throughput sequencing, including a high-throughput sequencing module and a data analysis module. Wherein, the high-throughput sequencing module uses the kit in Example 2, and is performed according to the high-throughput sequencing steps in Example 1; the data analysis module is performed according to the data analysis steps in Example 1.

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Abstract

The invention relates to a specific lymphocyte content analysis method and device based on TCR/BCR high-throughput sequencing, and belongs to the technical field of gene detection. The analysis methodcomprises the following steps of performing high-throughput sequencing: extracting genome DNA of a biological sample to be detected, adding a predetermined amount of artificially synthesized TCR/BCRcoding gene internal standard sequences, and carrying out high-throughput sequencing; and performing data analysis: obtaining high-throughput sequencing data and TCR/BCR gene sequences obtained by analysis, calculating the amplification multiple according to the input internal standard sequences, comparing the TCR/BCR gene sequences of the sample to be detected with antigen receptor gene sequencesA of target lymphocytes, obtaining the number of original antigen receptor gene sequences consistent with the antigen receptor gene sequences A of the target lymphocyte, the number of sequences withsequencing errors, the number of sequences subjected to secondary rearrangement and the number of mutated sequences, obtaining the total number of target lymph sequences, and calculating the number ofthe contained target lymphocytes according to the total number of the target lymph sequences and the amplification multiple, so that the content of each T/B cell can be accurately determined.

Description

technical field [0001] The invention relates to the technical field of gene detection, in particular to a method and device for analyzing the content of specific lymphocytes based on TCR / BCR high-throughput sequencing. Background technique [0002] Adaptive immunity is also called acquired immunity. Specific immune cells mainly include T lymphocytes involved in cellular immunity and B lymphocytes involved in humoral immunity. Adaptive immunity has the characteristics of antigen specificity and immune memory, and its antigen specificity means that different antigens need to be recognized and responded by different lymphocytes. In clinical practice, it is often necessary to determine the content of specific lymphocytes in biological samples (the composition of each component in biological samples is as follows: figure 1 shown), for example, the immune effect of the vaccine can be determined by detecting whether there are specific vaccine-specific immune cells; or the detectio...

Claims

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Application Information

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IPC IPC(8): C12Q1/6869C12M1/34C12M1/00
CPCC12Q1/6869C12Q2531/113C12Q2537/143C12Q2535/122C12Q2537/165C12Q2545/101
Inventor 王谢苏政
Owner 深圳泛因医学有限公司
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