Immunofluorescence kit for detecting PD-L1 expression of peripheral blood circulating tumor cells of prostate cancer patient and detection method
A PD-L1 and peripheral blood circulation technology, applied in tumor/cancer cells, cell dissociation methods, animal cells, etc., can solve the problem that tissue samples cannot evaluate the real-time dynamic status of PD-L1 in patients, false negatives, false positives in detection methods, etc. problems, to avoid edge effects, reduce cell loss, and achieve good stability
Inactive Publication Date: 2020-08-18
SHANDONG FIRST MEDICAL UNIV & SHANDONG ACADEMY OF MEDICAL SCI
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[0008] Aiming at the disadvantages of detecting patients with advanced tumor or recurrent prostate cancer in real-time or repeated puncture to obtain tissue samples in the prior art, and thus not being able to evaluate the real-time dynamic status of PD-L1 in patients, and the existing detection methods are prone to false positives and false negatives, this paper The invention provides an immunofluorescence kit and detection method for detecting the expression of PD-L1 in the peripheral blood of patients with prostate cancer: the circulating tumor cells (CTCs) in the peripheral blood of patients with advanced prostate cancer are separated by using a membrane filtration device, and the immunofluorescent Detection of PD-L1 expression on CTC by fluorescence technique
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Embodiment 1
[0050] 1. Use a membrane filtration device to separate and obtain CTCs in the peripheral blood of advanced or recurrent prostate cancer patients whose tissue samples cannot be obtained, and determine whether CTCs exist:
[0051] Collect 5ml of fasting blood from the median vein of the elbow for 8-12 hours, dilute the peripheral blood with 45ml of diluent, and then add 3ml of 4% paraformaldehyde to fix the diluted blood sample for 10 minutes;
[0052] At fixed intervals, assemble the membrane filtration device: as attached figure 1 , figure 2 , image 3 As shown, the filtering device is composed of a filter 3, a filter membrane 7, a blood sample container 2, a waste liquid tank 5, and an iron stand 1;
[0053] Wet the filter 3 with 10ml of PBS, and then add the fixed peripheral blood sample into the blood sample container 2 of the membrane filtration device, so that it is naturally filtered by gravity, and the CTC is trapped on the filter membrane 7;
[0054] The diameter o...
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Abstract
The invention discloses an immunofluorescence kit for detecting PD-L1 expression of peripheral blood circulating tumor cells of a prostate cancer patient and a detection method. The kit comprises diluent, a decolorizing solution, a staining solution A, a staining solution B, methanol, 2% PFA, 10% goat serum, mouse anti-CK, rat anti-CD45 and rabbit anti-PD-L1, a fluorescently-labeled goat anti-mouse, a fluorescently-labeled goat anti-rat, a fluorescently-labeled goat anti-rabbit secondary antibody suspension and a DAPI tablet. The detection method comprises the steps: peripheral blood collection, membrane filtration separation and dyeing, the judgment of whether CTC exists or not, and the detection of the PD-L1 expression condition of the peripheral blood CTC through an immunofluorescence method. According to the method provided by the invention, the PD-L1 expression condition of a patient with advanced or recurrent prostate cancer can be detected without acquiring a tissue specimen through needle biopsy, and the real-time dynamic detection can be realized by utilizing a minimally invasive technology.
Description
technical field [0001] The invention provides an immunofluorescence kit and a detection method for detecting the expression of PD-L1 in peripheral blood circulating tumor cells of prostate cancer patients, and belongs to the technical field of molecular biology. Background technique [0002] Worldwide, the incidence of prostate cancer ranks second among male malignancies, and cancer-specific death ranks sixth. According to the 2015 China Cancer Registration Annual Report, the incidence of prostate cancer in China ranked 9th among malignant tumors in 2011, and the incidence and mortality of prostate cancer in urban men ranked 6th and 9th among male malignant tumors, respectively. The difficulty in the treatment of prostate cancer is metastatic prostate cancer, especially castration resistance after endocrine therapy. At present, the treatment methods for metastatic prostate cancer after castration resistance are limited, and all conventional treatment methods (radiotherapy, c...
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Login to View More IPC IPC(8): G01N33/574C12N5/09
CPCC12N5/0694C12N2509/10G01N33/57434G01N33/57484G01N2333/70532
Inventor 李胜王焕昇李浩高德海
Owner SHANDONG FIRST MEDICAL UNIV & SHANDONG ACADEMY OF MEDICAL SCI