Molecular probe and kit for early prediction of metastatic prostate cancer and application of molecular probe
A technology of prostate cancer and molecular probes, applied in the determination/inspection of microorganisms, biochemical equipment and methods, DNA/RNA fragments, etc., can solve the problems of low specificity and sensitivity, hysteresis, and inability to be used repeatedly for a long time etc. to achieve the effects of high sensitivity and specificity, less trauma, and automated detection methods
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Embodiment 1
[0041] An embodiment of the probe set for early prediction of metastatic prostate cancer in the present invention, the probe set for early prediction of metastatic prostate cancer described in this embodiment includes epithelial markers, mesenchymal markers, CD45 probes and LCRMP4 probes ; The epithelial marker is made up of EpCAM probe, CK8 probe, CK18 probe and CK19 probe, and the mesenchymal marker is made up of Vimentin probe and Twist probe; Wherein, the sequence of the EpCAM probe is as follows Shown in SEQ ID NO:1, the sequence of the CK8 probe is shown in SEQ ID NO:2, the sequence of the CK18 probe is shown in SEQ ID NO:3, the sequence of the CK19 probe is shown in SEQ ID NO:4, Vimentin The sequence of the probe is shown in SEQ ID NO:5, the sequence of the Twist probe is shown in SEQ ID NO:6, the sequence of the CD45 probe is shown in SEQ ID NO:7, and the sequence of the LCRMP4 probe is shown in SEQ ID NO :8 shown. The specific sequences of the probes are shown in Tab...
Embodiment 2
[0046] An embodiment of the kit for early prediction of metastatic prostate cancer of the present invention, the kit for early prediction of metastatic prostate cancer described in this embodiment is an in situ hybridization detection kit, which includes: hybridization probes, permeabilization Reagent, digestion solution, hybridization buffer solution, pre-amplification working solution, amplification working solution, chromogenic solution, counterstaining solution and washing solution;
[0047]Wherein, the hybridization probe is the probe set described in Example 1, the LCRMP4 probe, EpCAM probe, CK8 probe, CK18 probe, CK19 probe, Vimentin probe, Twist probe and CD45 probe Both have markers; epithelial markers (EpCAM probe, CK8 probe, CK18 probe and CK19 probe) are labeled with red fluorescence; mesenchymal markers (Vimentin probe and Twist probe) are labeled with green fluorescence; The LCRMP4 probe is labeled with purple fluorescence; the CD45 probe is labeled with blue flu...
Embodiment 3
[0055] In this example, the kit described in Example 2 was used to detect the expression level of LCRMP4 by using the RNA in situ hybridization technique of CTCs to predict metastatic prostate cancer. In this example, the process of detecting the expression level of LCRMP4 by using the RNA in situ hybridization technique of CTCs is as follows figure 1 shown.
[0056] The specific steps are as follows:
[0057] 1. Clinical sample collection and processing
[0058] a. Collect peripheral blood:
[0059] 1. Collect 5 mL of peripheral blood with EDTA anticoagulant blood collection tubes, and collect 2 tubes (10 mL for each patient);
[0060] 2. After blood collection, invert the blood collection tube more than 5 times and mix well, then store it at 4°C, and the storage time should not exceed 4 hours;
[0061] 3. Record the corresponding number of the patient, and use the connector to transfer the blood in the blood collection tube into the sample storage tube (containing red bl...
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