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Method for separating enriched target cells from tissues

A technology for separating, enriching, and targeting cells. It is applied in animal cells, tumor/cancer cells, vertebrate cells, etc. It can solve the problems of incomplete and incomplete collection, and achieve the effect of improving sensitivity.

Inactive Publication Date: 2011-09-07
GD TECH INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

After pathological sections, research and analysis of specific cells, DNA / RNA collection and analysis of single cells have become common in clinical research and applications, but the collection of DNA / RNA of single cells from sliced ​​tissues is incomplete and not comprehensive. complete

Method used

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  • Method for separating enriched target cells from tissues
  • Method for separating enriched target cells from tissues
  • Method for separating enriched target cells from tissues

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Using magnetic beads bound to epithelial cell adhesion molecule antibodies to enrich cancer cells treated with 10% dimethyl sulfoxide medium and preserved in liquid nitrogen.

[0031] (1) Add fresh cancer tissue pieces into phosphate buffered solution (PBS), and make them as thin as possible to make a single-cell suspension.

[0032] (2) Centrifuge the single cell suspension at 1000 rpm for 5 minutes, remove the supernatant, add trypsin (Tripsin) for 5 minutes; then add cell culture medium containing 10% calf serum; Centrifuge at a speed of 1000 rpm for 5 minutes, remove the supernatant; finally add calf serum containing 10% DMSO, and transfer to a 2 mL test tube.

[0033] (3) Store the test tube at -80°C for 12 hours, then transfer to a liquid nitrogen tank for storage.

[0034] (4) Take out the required cancer tissue from the liquid nitrogen tank to dissolve the cancer tissue; add 10 times the volume of protein refolding buffer and incubate for 60 minutes; finally ad...

Embodiment 2

[0054] In this embodiment, enriched cancer cells are separated from bone marrow fluid, and cancer cells are detected. The specific steps are as follows:

[0055] (1) Use clinically extracted bone marrow fluid from cancer patients and dilute it 3 times with PBS.

[0056] (2) After mixing, add one-tenth volume of magnetic beads with epithelial cell adhesion molecule antibody, and incubate at 4 degrees or room temperature for 30 minutes to one hour.

[0057] (3) Use a magnet to absorb the target cells bound to the magnetic beads in the tube. Wash three times with PBS buffer.

[0058] (4) Withdraw the magnet to release the target cells bound to the magnetic beads.

[0059] (5) Collect target cells and identify cancer cells.

[0060] The cell identification method is the same as in Example 1. Through the above steps, the following results can be obtained, refer to Figure 7 . from Figure 7 It can be seen that bone marrow cancer cells positive for epithelial cell adhesion mo...

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Abstract

The invention discloses a method for separating enriched target cells from tissues, and belongs to the field of biological science and medicament development. The method comprises the following steps of: preparing the tissues into suspension of single cells, and separating and enriching target cells from the cell suspension by using a positive enrichment method. The method can separate and enrich the target cells from the tissues in different treatment states, and provides enough and high-purity experimental materials for basic research, clinical research, diagnosis, medicament development and the like.

Description

technical field [0001] The invention belongs to the fields of biological science and drug research and development. Background technique [0002] The isolation and enrichment of target cells has a wide range of applications in cytology, molecular biology and immunology. The reported clinical research results of blood circulating cancer cells show that the number of blood circulating cancer cells in cancer patients can be used for early detection, diagnosis, follow-up and prognosis of cancer patients. Since cancer cells exist in very small amounts in the blood of cancer patients, in order to accurately detect the presence and quantity of cancer cells in blood, it is necessary to enrich the cancer cells in blood samples to improve the sensitivity and accuracy of detection. In recent years, the separation and enrichment of blood circulating cancer cells have received more and more scientific research and clinical attention. Taking breast cancer patients as an example, the num...

Claims

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Application Information

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IPC IPC(8): C12N5/07C12N5/09
Inventor 邓亚光邓存常邓伟平
Owner GD TECH INC
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