Method for detecting specific mRNA in cell adopting molecular probe
A technology of molecular probes and cells, applied in the direction of measuring devices, analytical materials, material excitation analysis, etc., can solve the problems of inability to distinguish samples, limited sensitivity, detection errors, etc., and achieve high accuracy, strong measurement stability, and easy operation easy effect
Inactive Publication Date: 2016-05-25
TSINGHUA UNIV
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However, although the fluorescence intensity can distinguish the difference in the fluorophore concentration of the sample, the sensitivity is limited, and it cannot distinguish the samples with small difference in the fluorescence intensity.
At the same time, the detection method of fluorescence intensity will also be affected by factors such as excitation light intensity, detector gain, optical path and sample light loss, sample fluorophore concentration, photobleaching and other factors, resulting in detection errors
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[0011] The method for detecting intracellular mRNA provided by the present invention using a molecular probe will be further described in conjunction with specific examples below.
[0012] The present invention provides a kind of method adopting molecular probe to detect intracellular specific mRNA, it comprises the following steps:
[0013] S1, providing a first culture medium mixed with molecular probes, the molecular probes include reporter DNA modified with fluorescent molecules;
[0014] S2, placing the cells in a culture dish containing a second culture medium;
[0015] S3, when the cultured and grown cells occupy 60%-80% of the bottom area of the culture dish, suck off the second culture medium in the culture dish;
[0016] S4, putting the first culture medium into the culture dish, and after the cells continue to culture for 2-6 hours, suck off the first culture medium;
[0017] S5, using a laser confocal microscope to image the cell, using a single photon counter ...
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Abstract
The invention relates to a method for detecting specific mRNA in a cell adopting a molecular probe. The method comprises the following steps: providing a first culture medium doped with the molecular probe, wherein the molecular probe comprises a report DNA modified by fluorescent molecules; putting the cell into a culture dish with a second culture medium for cultivation; when the cells account for 60 to 80 percent of the bottom of the culture dish, sucking away the second culture medium in the culture dish, then putting the first culture medium into the culture dish for further culturing the cells for 2 to 6 hours, and then sucking away the first culture medium; imaging the cells adopting a laser confocal microscopy, and measuring and calculating the lifetime of fluorescence displayed in the cells adopting a single-photon-counter to obtain a fluorescence lifetime image, and detecting the information whether the specific mRNA exists in the cell according to the fluorescence lifetime image.
Description
technical field [0001] The invention relates to a method for detecting specific mRNA in cells, in particular to a method for detecting specific mRNA in cells by means of molecular probe fluorescence lifetime imaging. Background technique [0002] Cancer is an important cause of human death worldwide, so early detection and treatment of cancer has become an important challenge in the fields of medicine and biological research. Researchers have demonstrated that endogenous cancer-associated mRNAs are important indicators of cancer and that their expression levels can reveal information about cancer progression and prognosis. Therefore, identification and identification of intracellular cancer-associated mRNAs can provide important information for early diagnosis and treatment of cancer. At present, there are many methods to identify and identify mRNA, such as Northern blot, microarray analysis technology, and RT-PCR technology. However, these methods have cumbersome steps, r...
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Login to View More IPC IPC(8): G01N21/64
CPCG01N21/6486
Inventor 周明史婧
Owner TSINGHUA UNIV