Marker combination and kit for detecting tumor cells in body fluid sample

A detection kit and technology for tumor cells, applied in the field of marker combinations and kits for detection of tumor cells in body fluid samples, can solve the problems of false positives of tumor cells and achieve the effect of improving sensitivity and specificity

Pending Publication Date: 2021-10-26
3D BIOMEDICINE SCI & TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

It mainly solves the problem that the existing technology CellSearch system is prone to false positives in identifying tumor cells

Method used

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  • Marker combination and kit for detecting tumor cells in body fluid sample
  • Marker combination and kit for detecting tumor cells in body fluid sample
  • Marker combination and kit for detecting tumor cells in body fluid sample

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] Example 1: Verification of Telomerase Expression in Tumor Cell Lines

[0046] (1) Centrifuge 1 ml of the MCF-7 cell line (500 g, 5 minutes), discard the supernatant, and add 2 mL of HBSS buffer to resuspend the cells;

[0047] (2) After the above resuspension is counted by a cell counter, take an appropriate amount and drop it on a glass slide, and let it stand for 10 minutes;

[0048] (3) Blot off the solution on the surface of the slide, add 100ul of TERT-FITC antibody with a concentration of 2ug / ml dropwise, and incubate at room temperature for 1h;

[0049] (4) After the incubation, wash the chip 3 times with PBS, drop 100ul of DAPI onto the glass slide, and incubate at room temperature for 10min;

[0050] (5) After the incubation, the chip was washed 3 times with PBS, and imaged with a high-speed fluorescence imaging device.

[0051] Imaging results such as figure 1 As shown, the images of cells with different fluorescence characteristics obtained by scanning sli...

Embodiment 2

[0052] Example 2: Pleural effusion sample test of lung cancer patients

[0053] The testing method of the pleural effusion sample of lung cancer patient comprises the following steps:

[0054] (1) Centrifuge (500g, 5min) 20mL of pleural effusion from lung cancer patients to separate the cells, add 5mL red blood cell lysate (BD company) to lyse in the dark for 5min, centrifuge again (500g, 5min), discard the supernatant and use Hank's balanced salt Solution (HBSS) to resuspend and wash the cells, centrifuge (500g, 5min), discard the supernatant, add 2mL HBSS to resuspend the cells;

[0055] (2) After the above resuspension is counted by a cell counter, take an appropriate amount and drop it on a glass slide, and let it stand for 10 minutes;

[0056] (3) Absorb the solution on the surface of the slide, and drop 100ul antibody mixture onto the slide, wherein the antibody mixture includes: 2ug / ml FITC-labeled TERT antibody, PE-labeled 1ug / ml CK antibody and 2ug / ml ml of APC-labele...

Embodiment 3

[0061] Example 3: Urine Tests for Bladder Cancer Patients

[0062] The method of urine testing in patients with bladder cancer involves the following steps:

[0063] (1) Separate the cells from 20 mL of bladder cancer patient urine (500 g, 5 min), add 5 mL of erythrocyte lysate (BD company) to lyse in the dark for 5 min, centrifuge again (500 g, 5 min), discard the supernatant and equilibrate with Hank Resuspend and wash cells in saline solution (HBSS), centrifuge (500g, 5min), discard supernatant, add 2mL HBSS to resuspend cells;

[0064] (2) After the above resuspension is counted by a cell counter, take an appropriate amount and drop it on a glass slide, and let it stand for 10 minutes;

[0065] (3A) Absorb the solution on the surface of the slide, and drop 100ul antibody mixture onto the slide, wherein the antibody mixture includes: 2ug / ml FITC-labeled TERT antibody, 1ug / ml PE

[0066] Labeled CK antibody and 2ug / ml APC-labeled CD45 antibody, incubated at room temperatur...

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Abstract

The invention discloses a marker combination and a kit for detecting tumor cells in a body fluid sample. The marker combination comprises TERT and CD45, and further comprises CK and a cell nucleus dye. The marker combination can be used for detecting tumor cells in a body fluid sample. The kit provided by the invention comprises the target composition, a red blood cell lysis reagent and a target cell enrichment reagent. The invention provides a group of novel tumor cell recognition markers TERT+ / CK+ / CD45- / DAPI+, wherein TERT is telomerase reverse transcriptase. The method has the advantages that firstly, the detection sensitivity and specificity can be obviously improved; d ansecondly, tumor cells derived from epithelium can be detected. For a patient with urothelial carcinoma, tumor cells and normal cells cannot be distinguished by using a traditional EpCAM+ / CK+ / CD45- / DAPI+ detection method, and the tumor cells and the normal cells can be obviously distinguished by using the TERT+ / CK+ / CD45- / DAPI+ detection method.

Description

technical field [0001] The invention belongs to the technical field of biomedical detection, and in particular relates to a marker combination and a kit for detecting tumor cells in body fluid samples. Background technique [0002] Cancer has a high morbidity and mortality rate worldwide and has seriously threatened human health. Liquid biopsy is a new technology for non-invasive detection of tumors that has emerged in recent years. Due to tumor metastasis, its cells are shed from the original site and enter the circulatory system, and then spread to blood, pleural effusion, cerebrospinal fluid and even urine. The detection of free rare tumor cells and their molecular characteristics from these liquid samples can replace the biopsy of tumor lesions in situ, so it is called liquid biopsy. Because lesion biopsy is traumatic and in most cases it is difficult to perform a second biopsy on patients, liquid biopsy, especially for blood, is very popular in clinic because of its n...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/68G01N33/573G01N33/58G01N33/543
CPCG01N33/68G01N33/6872G01N33/573G01N33/582G01N33/56966G01N33/54326G01N2333/4742G01N2333/9128G01N2333/70589
Inventor 钱祺王晓楠赵小玉李振建张亚楠
Owner 3D BIOMEDICINE SCI & TECH CO LTD
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