LU502916B1 - Method for detecting cycle morphology of cancer cells - Google Patents
Method for detecting cycle morphology of cancer cells Download PDFInfo
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- LU502916B1 LU502916B1 LU502916A LU502916A LU502916B1 LU 502916 B1 LU502916 B1 LU 502916B1 LU 502916 A LU502916 A LU 502916A LU 502916 A LU502916 A LU 502916A LU 502916 B1 LU502916 B1 LU 502916B1
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Abstract
The invention provides a method for detecting the cycle morphology of cancer cells, and relates to the field of biology. Where, the method comprises: extracting four portions of culture medium containing a certain number of cancer cells from the culture medium, and respectively bottling in different culture bottles, adding cell cycle blockers into three of the culture bottles and treating for 24 hours so that the three portions of cancer cells stay in G1 phase, G2 phase and M phase respectively; while the cancer cells in the other culture flask evolved normally, then cutting a plurality of infrared slides into the size of 1cm×1cm respectively, putting into a sterile culture dish in an array arrangement mode after disinfection, and laminating one slide on each infrared slide; dripping culture solutions cultured in each culture bottle containing a certain cell concentration into different slides, and performing synchronous radiation infrared micro-spectral imaging on the culture solutions on each slide to obtain four copies of concentration distribution of functional groups and different growth states of cancer cells. Thereby recognizing cancer cells in essence.
Description
METHOD FOR DETECTING CYCLE MORPHOLOGY OF
CANCER CELLS
The invention relates to the field of biology, and in particular to a method for detecting the cycle morphology of cancer cells.
At present, the classification of cancer cells is roughly divided into four types: squamous cell carcinoma, adenocarcinoma, undifferentiated carcinoma and carcinoma that cannot be classified by cancer cells. In further classification, that is, well-differentiated cancer and poorly differentiated cancer. This distinction is very rough and does not distinguish the nature of cancer cells. It is only classified according to the appearance of cancer cells with different shapes and changes seen under the microscope.
However, the morphology of cancer cells varies in each cell cycle. Such classification is obviously insufficient for understanding and treating cancer cells, which is the bottleneck of cancer treatment. Therefore, there is an urgent need to design a method that can detect the cycle morphology of cancer cells to recognize cancer cells in nature.
The invention provides a method for detecting the cycle morphology of cancer cells.
The invention is realized as the following:
A method for detecting the cycle morphology of cancer cells, comprises:
Extracting four portions of culture medium containing a certain number of cancer cells from the culture medium, and respectively bottling in different culture bottles, adding cell cycle blockers into three of the culture bottles and treating for 24 hours so that the three portions of cancer cells stay in G1 phase, G2 phase and M phase HUS02916 respectively; cutting a plurality of infrared slides into the size of lecmxlcm respectively, putting into a sterile culture dish in an array arrangement mode after disinfection, and laminating one slide on each infrared slide; dripping culture solutions cultured in each culture bottle containing a certain cell concentration into different slides, and performing synchronous radiation infrared micro-spectral imaging on the culture solutions on each slide to obtain four copies of concentration distribution of functional groups and different growth states of cancer cells.
Where, in a preferred embodiment of the present invention, the method for detecting the cycle morphology of cancer cells can also be technically characterized in that:
Further, the culture medium is RPMI-1640 culture medium containing 10% heat-inactivated foetal bovine serum and 1% penicillin-streptomycin-L- glutamine.
Further, the culture medium is stored at 37°C under a humid atmosphere containing 5% CO».
Further, before the step of dropping the culture solution onto the slide, further comprising:
Adding a cell suspension agent to each of the culture bottles to suspend cancer cells from the culture bottles.
Further, the cell suspending agent is trypsin or ethylenediamine tetra acetic acid.
Further, after the step of adding a cell suspending agent into each culture bottle, further comprising:
Sucking the culture solution containing the certain cell concentration from each culture bottle and dropping it into different culture holes of the cell culture plate;
Where, the culture solution dripped on the slide is sucked from the culture hole.
Further, after the step of respectively filling four parts of the culture solution in different culture bottles, further comprising: respectively carrying out centrifuge concentration treatment on culture solutions in the four culture bottles or washing by 1x phosphate buffered saline with a pH of
74 LU502916
Further, the disinfection mode of the infrared slide is as follows: washing the infrared slide in 70% ethanol, and exposing to ultraviolet radiation overnight after washing.
Further, the cell cycle blocker is 100ug/ml of L-momicin or 50ng/ml of nocodazole.
Further, the certain cell concentration is 10%.
The invention has the beneficial effects that: by extracting four portions of culture medium containing a certain number of cancer cells from the culture medium, and respectively bottling in different culture bottles, adding cell cycle blockers into three of the culture bottles and treating for 24 hours so that the three portions of cancer cells stay in G1 phase, G2 phase and M phase respectively; while the cancer cells in the other culture flask evolved normally, then cutting a plurality of infrared slides into the size of lemx1cm respectively, putting into a sterile culture dish in an array arrangement mode after disinfection, and laminating one slide on each infrared slide; dripping culture solutions cultured in each culture bottle containing a certain cell concentration into different slides, and performing synchronous radiation infrared micro-spectral imaging on the culture solutions on each slide to obtain four copies of concentration distribution of functional groups and different growth states of cancer cells. So as to obtain the morphology of cancer cells in G1 phase, G2 phase, M phase and the evolution process, thereby recognizing cancer cells in essence.
FIG. 1 is a functional group concentration distribution diagram of human lung cancer cells in the first embodiment of the present invention;
FIG. 2 is a diagram of different growth states of human lung cancer cells in the first embodiment of the present invention.
In order to explain the technical scheme of the embodiment of the present invention more clearly, the following drawings that need to be used in the embodiment will be briefly introduced. It should be understood that the following drawings only show some embodiments of the present invention, so they should not 17008918 be regarded as limiting the scope. For those of ordinary skill in the art, other relevant drawings can be obtained according to these drawings without any creative effort.
In order to make the purpose, technical scheme and advantages of the embodiments of the present invention clearer, the technical scheme in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention.
Obviously, the described embodiments are part of the embodiments of the present invention, but not all of them. Based on the embodiment of the present invention, all other embodiments obtained by ordinary technicians in the field without creative labour belong to the scope of protection of the present invention. Therefore, the following detailed description of the embodiments of the invention provided in the drawings is not intended to limit the scope of the claimed invention, but only to represent selected embodiments of the invention. Based on the embodiment of the present invention, all other embodiments obtained by ordinary technicians in the field without creative labour belong to the scope of protection of the present invention.
In the description of the present invention, it is to be understood that the terms "centre", "longitudinal", "transverse", "length", "width", "thickness", "upper", "lower", "front", "rear", "left", "right", "vertical", "horizontal", "top", "bottom", "inner", "outer", "clockwise", "counter-clockwise" and the like indicate an orientation or positional relationship based on that shown in the drawings, It is intended merely to facilitate description of the invention and to simplify the description, and is not intended to indicate or imply that a device or element in question must have a particular orientation, be constructed and operate in a particular orientation, and is therefore not to be construed as limiting the invention.
In addition, the terms "first" and "second" are only used for descriptive purposes, and cannot be understood as indicating or implying relative importance or implicitly indicating the number of indicated technical features. Therefore, the features defined as "first" and "second" may include one or more of these features explicitly or HUS02916 implicitly. In the description of the present invention, "multiple" means two or more, unless otherwise specifically defined.
In the present invention, unless otherwise specified and limited, terms such as 5 "installed", "connected", "connected" and "fixed" should be understood broadly, for example, they can be fixed, detachable or integrated; can be mechanically connected or electrically connected; it can be directly connected or indirectly connected through an intermediate medium, and it can be the internal communication of two elements or the interaction between two elements. For those of ordinary skill in the art, the specific meanings of the above terms in the present invention can be understood according to specific situations.
In the present invention, unless otherwise specified and limited, the "upper" or "lower" of the first feature on the second feature may include the direct contact between the first and second features, or the contact between the first and second features through another feature instead of direct contact. Furthermore, the first feature "above", "on top of" and "onto" the second feature includes that the first feature is directly above and obliquely above the second feature, or only indicates that the horizontal height of the first feature is higher than that of the second feature. "Below", "under" and "at the bottom of" the first features include that the first features are directly below and obliquely below the second features, or only indicate that the horizontal height of the first features 1s smaller than that of the second features.
Embodiment 1
The first embodiment of the invention provides a method for detecting the cycle morphology of cancer cells, the method for detecting the cycle morphology of cancer cells in this embodiment is realized based on synchrotron radiation infrared (SR-IR) microscopic spectrum and SR-IR spectrum imaging technology. Specifically comprising the steps SO1 - S03:
S01, extracting four portions of culture medium containing a certain number of cancer cells from the culture medium, and respectively bottling in different culture bottles, adding cell cycle blockers into three of the culture bottles and treating for 24 HUS02916 hours so that the three portions of cancer cells stay in G1 phase, G2 phase and M phase respectively.
In the concrete implementation, the culture medium is RPMI-1640 culture medium containing 10% heat-inactivated foetal bovine serum and 1% penicillin-streptomycin-L- glutamine (Roswell Park Memorial Institute-1640), and the culture medium is stored at 37°C under a humid atmosphere containing 5% CO». The cell cycle blocker is 100pg/ml of L-momicin or 50ng/ml of nocodazole.
In this embodiment, human lung cancer cell line A549 (CCL-185) is selected as an experimental model, and samples of human lung cancer cells are obtained from the
American Type Culture Collection (ATCC). However, it should be noted that this does not limit the scope of invention of the method for detecting the cycle morphology of cancer cells, and in other embodiments, other cancer cells may be used as experimental models.
Where, G1 phase is the first interval of cell growth, which is mainly characterized by vigorous cell metabolism and the synthesis of various biochemical substances needed by cell growth, such as protein, sugars, lipids, RNA, etc.. The cell volume increases, making preparations for DNA synthesis. Therefore, G1 phase is also called DNA synthesis preparation phase or replication prophase. G2 phase, also known as the late stage of DNA synthesis or mitosis preparation stage, is mainly characterized by the rapid growth of cells and the synthesis of a large number of protein required for mitosis. Some cancers can enter mitosis directly after the completion of DNA replication without G2 phase. M phase, also known as mitosis, refers to the period of nuclear division and cytoplasmic division in the cell cycle after mitosis. It can be divided into interphase, prophase, (prometaphase), metaphase, anaphase and telophase according to the different chromosome morphology and activities. Cell division stage includes prophase, metaphase, anaphase and telophase.
S02, cutting a plurality of infrared slides into the size of lemx1cm respectively, putting into a sterile culture dish in an array arrangement mode after disinfection, and laminating one slide on each infrared slide.
Where, the function of infrared slide is to transmit the infrared light emitted by HUS02916 the outside spectral imager back, so as to realize spectral imaging. Meanwhile, the plurality of infrared slides are cut into a size of 1cmx1cm and arranged in an array arrangement manner so that the growth state diagrams of each sample obtained by the later synchronous radiation infrared spectroscopy imaging are arranged in an array arrangement manner, and the growth state diagrams of each sample are in a square area with a size of 1cm<1cm (as shown in FIG. 2).
In addition, in the concrete implementation, the slide can also be made to a size of lcmx 1 cm.
In the concrete implementation, the disinfection mode of the infrared slide is as follows: washing the infrared slide in 70% ethanol, and exposing to ultraviolet radiation overnight after washing.
S03, dripping culture solutions cultured in each culture bottle containing a certain cell concentration into different slides, and performing synchronous radiation infrared micro-spectral imaging on the culture solutions on each slide to obtain four copies of concentration distribution of functional groups and different growth states of cancer cells.
As shown in FIG. 1 and FIG. 2, the figures show the functional group concentration distribution of human lung cancer cells and the different growth states of human lung cancer cells obtained in this embodiment.
Specifically, multiple groups of cancer cell samples can be dripped for detection, and each group of samples includes four groups of culture solutions in different sample bottles, and each sample is correspondingly attached to a slide, while multiple groups of cancer cell samples can be arranged on the corresponding slide in chronological order, as shown in FIG. 2.
During spectral imaging, the whole sterile culture dish can be put into a synchrotron radiation infrared spectrometer for SR-IR spectral imaging.
It should also be pointed out that the whole operation process is carried out in a sterile environment to ensure that the sample has no biochemical stains.
In summary, the method for detecting the cycle morphology of cancer cells in this embodiment, by extracting four portions of culture medium containing a certain HUS02916 number of cancer cells from the culture medium, and respectively bottling in different culture bottles, adding cell cycle blockers into three of the culture bottles and treating for 24 hours so that the three portions of cancer cells stay in G1 phase, G2 phase and
M phase respectively; while the cancer cells in the other culture flask evolved normally, then cutting a plurality of infrared slides into the size of 1cmx1cm respectively, putting into a sterile culture dish in an array arrangement mode after disinfection, and laminating one slide on each infrared slide; dripping culture solutions cultured in each culture bottle containing a certain cell concentration into different slides, and performing synchronous radiation infrared micro-spectral imaging on the culture solutions on each slide to obtain four copies of concentration distribution of functional groups and different growth states of cancer cells. Thereby recognizing cancer cells in essence. Therefore, the method for detecting the cycle morphology of the cancer cells realizes obtaining the protein and lipid images with spatial resolution in the living human cancer cells of several microns by the synchronous radiation infrared microscopic spectrum and the SR-IR spectrum imaging technology, can achieve the condition without any biochemical stains and fixative, can essentially recognize cancer cells, and can be used as an important technical means of cancer cell screening.
Embodiment 2
The second embodiment of the invention provides a method for detecting the cycle morphology of cancer cells, specifically comprising the steps S11 - S16:
S11, extracting four portions of culture medium containing a certain number of cancer cells from the culture medium, and respectively bottling in different culture bottles, respectively washing the culture solutions in the four culture bottles by using
Ix phosphate buffered saline with a pH of 7.4, and adding fresh culture solutions (free of cancer cells).
In the concrete implementation, a cytometer can be used to measure the number of cancer cells in the culture medium.
Where, the culture medium is RPMI-1640 culture medium containing 10% HUS02916 heat-inactivated foetal bovine serum and 1% penicillin-streptomycin-L- glutamine, and the culture medium is stored at 37°C under a humid atmosphere containing 5%
CO».
It should be noted that, the purpose of washing the culture medium with 1x phosphate buffered saline (1xPBS) with pH7.4 is to release cells from cell cycle blocks.
In addition, in other implementations, when there are too many culture solutions, resulting in low cell concentration, the culture solutions in the four culture bottles can also be concentrated by the centrifuge.
S12, adding cell cycle blockers into three of the culture bottles and treating for 24 hours so that the three portions of cancer cells stay in G1 phase, G2 phase and M phase respectively.
Where, the cell cycle blocker is 100pug/ml of L-momicin or 50ng/ml of nocodazole.
S13, cutting a plurality of infrared slides into the size of lcmx1cm respectively, putting into a sterile culture dish in an array arrangement mode after disinfection, and laminating one slide on each infrared slide.
Where, the disinfection mode of the infrared slide is as follows: washing the infrared slide in 70% ethanol, and exposing to ultraviolet radiation overnight after washing.
S14, adding a cell suspension agent to each of the culture bottles to suspend cancer cells from the culture bottles.
Where, the cell suspending agent is trypsin or ethylenediamine tetra acetic acid, so as to suspend cancer cells from the culture bottle.
S15, sucking the culture solution containing the certain cell concentration from each culture bottle and dropping it into different culture holes of the cell culture plate for later using.
Where, the certain cell concentration is the ratio of the number of cells to the volume of culture solution, which is specifically 10% in this embodiment, that is, 10x\
x 10° ml culture medium contains 1x10° cells. HUS02916
S16, Sucking the culture solution with a certain cell concentration cultured in each culture bottle from different culture holes, dropping it on different slides, and performing synchrotron radiation infrared microspectroscopy imaging on the culture solution on each slide to obtain functional group concentration distribution of four cancer cells and different growth states of cancer cells.
The above is only the preferred embodiment of the present invention, and it is not intended to limit the present invention. For those skilled in the art, the present invention can be modified and varied. Any modification, equivalent substitution, improvement, etc. made within the spirit and principle of the present invention shall be included in the scope of protection of the present invention.
Claims (10)
1. A method for detecting the cycle morphology of cancer cells, characterized by comprising: extracting four portions of culture medium containing a certain number of cancer cells from the culture medium, and respectively bottling in different culture bottles, adding cell cycle blockers into three of the culture bottles and treating for 24 hours so that the three portions of cancer cells stay in G1 phase, G2 phase and M phase respectively; cutting a plurality of infrared slides into the size of 1cmx1cm respectively, putting into a sterile culture dish in an array arrangement mode after disinfection, and laminating one slide on each infrared slide; dripping culture solutions cultured in each culture bottle containing a certain cell concentration into different slides, and performing synchronous radiation infrared micro-spectral imaging on the culture solutions on each slide to obtain four copies of concentration distribution of functional groups and different growth states of cancer cells.
2. The method for detecting the cycle morphology of cancer cells according to claim 1, characterized in that, the culture medium is RPMI-1640 culture medium containing 10% heat-inactivated ~~ foetal bovine serum and 1% penicillin-streptomycin-L- glutamine.
3. The method for detecting the cycle morphology of cancer cells according to claim 1, characterized in that, the culture medium is stored at 37°C under a humid atmosphere containing 5% CO».
4. The method for detecting the cycle morphology of cancer cells according to claim 1, characterized in that, before dropping the culture solution onto the slide, further comprising: adding a cell suspension agent to each of the culture bottles to suspend cancer cells from the culture bottles.
5. The method for detecting the cycle morphology of cancer cells according to claim 4, characterized in that, the cell suspending agent is trypsin or ethylenediamine tetra acetic acid. 17008918
6. The method for detecting the cycle morphology of cancer cells according to claim 4, characterized in that, after adding a cell suspending agent into each culture bottle, further comprising: sucking the culture solution containing the certain cell concentration from each culture bottle and dropping it into different culture holes of the cell culture plate; where, the culture solution dripped on the slide 1s sucked from the culture hole.
7. The method for detecting the cycle morphology of cancer cells according to claim 1, characterized in that, after respectively filling four parts of the culture solution in different culture bottles, further comprising: respectively carrying out centrifuge concentration treatment on culture solutions in the four culture bottles or washing by 1x phosphate buffered saline with a pH of
7.4.
8. The method for detecting the cycle morphology of cancer cells according to claim 1, characterized in that, the disinfection mode of the infrared slide is as follows: washing the infrared slide in 70% ethanol, and exposing to ultraviolet radiation overnight after washing.
9. The method for detecting the cycle morphology of cancer cells according to claim 1, characterized in that, the cell cycle blocker is 100ug/ml of L-momicin or 50ng/ml of nocodazole.
10. The method for detecting the cycle morphology of cancer cells according to claim 1, characterized in that, the certain cell concentration is 10%.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| LU502916A LU502916B1 (en) | 2022-10-18 | 2022-10-18 | Method for detecting cycle morphology of cancer cells |
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| Application Number | Priority Date | Filing Date | Title |
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| LU502916A LU502916B1 (en) | 2022-10-18 | 2022-10-18 | Method for detecting cycle morphology of cancer cells |
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| LU502916B1 true LU502916B1 (en) | 2023-04-21 |
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