A rapid detection kit for circulating tumor cells

A detection kit and technology for tumor cells, applied in the field of rapid detection kits for circulating tumor cells, to achieve the effects of easy mastery by clinical personnel, objective results, and reduced burden

Active Publication Date: 2019-01-25
武汉海吉力生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] In order to solve the defects of the existing circulating tumor cell detection technology, the present invention provides a detection kit for simultaneously extracting circulating tumor cells from peripheral blood

Method used

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  • A rapid detection kit for circulating tumor cells
  • A rapid detection kit for circulating tumor cells
  • A rapid detection kit for circulating tumor cells

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] Embodiment 1: Kit composition and usage method.

[0035] (1) The components of the kit mainly include:

[0036] ① Oncoquick tumor cell density gradient separation solution 4ml (Geriner, Germany);

[0037] ② 10 porous membrane separation tubes with a volume of 15mL (Wuhan Haijili Biotechnology Co., Ltd.);

[0038] ③ 10 centrifuge tubes with a volume of 5 mL (Eppendorf, Germany);

[0039] ④ 488 Mouse Anti-CK7 / CK8 (BD, USA);

[0040] ⑤ BB515MouseAnti-Human CD326 antibody (BD, USA);

[0041] ⑥ Anti-Human CD325 (N-Cadherin) PE (eBioscience, USA).

[0042] (2) The specific detection process is as follows (see figure 1 , kit detection process flow chart):

[0043] ①Blood collection 5mL whole blood EDTA anticoagulant (stored at 4 degrees and pre-cooled; test within 24 hours)

[0044] ② Take 5 mL of peripheral blood from tumor patients and add it into a centrifuge tube with separation solution, and centrifuge at 1200 g for 25 minutes. After centrifugation, the peripher...

Embodiment 2

[0049] Example 2: Detection and analysis of simulated samples of healthy human peripheral blood mixed with tumor cell lines

[0050] In this example, the peripheral blood of healthy people was mixed with gradiently diluted MCF-7 cells, and it was used as a simulated sample for detection and analysis, so as to evaluate the tumor cell enrichment efficiency of the kit and the recovery rate of the kit, etc. Performance, detailed as follows:

[0051] 1. Sample preparation

[0052] The MCF-7 cells in the culture dish were digested and made into a single cell suspension, counted with a red blood cell counting plate, and diluted to an appropriate concentration with PBS. Add different concentrations of MCF-7 cells to 5 mL of healthy human peripheral blood anticoagulated with EDTA, and the concentrations are 800 / tube, 400 / tube, 100 / tube, 20 / tube, 5 / tube, 0 pcs / tube. 5 sets of repetitions were set for each concentration.

[0053] 2. Sample detection

[0054] (1) Gently invert the simu...

Embodiment 3

[0066] Example 3: Extraction and Analysis of Individual Samples of Circulating Tumor Cell DNA

[0067] 1. Sample preparation

[0068] The peripheral blood of 100 healthy people, the peripheral blood of 79 patients with breast cancer, the peripheral blood of 80 patients with lung cancer, and the peripheral blood of 30 patients with other primary tumors were selected as test samples (attached image 3 ).

[0069] 2. Sample detection:

[0070] The experimental process is as in Example 2.

[0071] 3. Test results: see attached Figure 4 .

[0072] The median value of normal population detection is: 1.8;

[0073] The median detection value of the tumor patient population is: 7.9;

[0074] There was a significant difference between the two groups (P<0.0001).

[0075] Table 2. Positive coincidence rate table of tumor patient population

[0076]

[0077] When 5 is selected as the cutoff value, it is obtained from the ROC curve model: the sensitivity is 73.8%, and the specif...

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Abstract

The invention discloses a rapid detection kit for circulating tumor cells. The rapid detection kit comprises a cell density gradient separating medium, a porous diaphragm separator tube, a centrifuge tube with a volume of 5 mL, epidermal tumor marker antibodies and mesenchymal tumor marker antibodies. The kit employs a density gradient centrifugation method to remove background interference of most erythrocytes and leucocytes in blood and enrich circulating tumor cells in the blood; then epidermal and mesenchymal tumor marker antibodies, altogether four fluorescent tumor marker antibodies, are used for fluorescence labeling of the circulating tumor cells; and a general flow cytometer is used for detecting and counting the circulating tumor cells. The kit provided by the invention takes the epidermal-mesenchymal transformation process of the tumor cells into consideration, increases captured mesenchymal circulating tumor cells, improves the positive coincident rate of detection, and is high in accuracy, rapid, convenient, and low in cost and time consumption; and detection counting is automatically accomplished by the flow cytometer, so good repeatability is realized.

Description

technical field [0001] The invention relates to the technical field of tumor prognosis detection, in particular to a rapid detection kit for circulating tumor cells. Background technique [0002] In my country, the incidence of malignant tumors is increasing year by year, and now it has become a public health problem that seriously affects people's health. Due to the fact that the biological characteristics of the tumor have not yet been fully clarified, in addition to traditional imaging and pathological methods, there is currently a lack of simple, rapid, and non-invasive methods for evaluating the prognosis of tumor surgery or radiotherapy and chemotherapy in clinic, and An effective means of tracking the patient's relapse and disease progression. [0003] Circulating tumor cells (CTCs) are tumor cells that are shed from the primary tumor or metastases and enter the peripheral blood circulation. They have biological characteristics such as tumor-specific antigens and gen...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/574G01N15/14
CPCG01N15/14G01N33/57484G01N2015/1486
Inventor 张志立黄俊郭骁赵平锋卢彦羽张全胡树军
Owner 武汉海吉力生物科技有限公司
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