CN111100930B

CN111100930B - Grading model for detecting benign and malignant degree of pancreatic tumor and application thereof

Info

Publication number: CN111100930B
Application number: CN201811259835.3A
Authority: CN
Inventors: 成彤; 周宁
Original assignee: Lisen Imprinting Diagnosis Technology Wuxi Co ltd
Current assignee: Lisen Imprinting Diagnosis Technology Wuxi Co ltd
Priority date: 2018-10-26
Filing date: 2018-10-26
Publication date: 2024-02-09
Anticipated expiration: 2038-10-26
Also published as: CN111100930A; WO2020082642A1

Abstract

The invention relates to a grading model for detecting pancreatic tumors and application thereof, wherein the grading model is used for grading the change of imprinted genes in pancreatic tumors by calculating the deletion expression quantity of imprinted genes, the abnormal copy number expression quantity of imprinted genes and the total expression quantity of imprinted genes. The detection model and the device of the invention show the appearance of imprinting missing on tissue and cell samples of pancreatic tumor patients in an intuitive way, objectively, intuitively, early and accurately detect the change of imprinting (trace) genes by the method of marking imprinting genes in situ, and can provide a quantized model, thereby greatly contributing to diagnosis of pancreatic tumors.

Description

Grading model for detecting benign and malignant degree of pancreatic tumor and application thereof

Technical Field

The invention relates to the field of biotechnology, relates to the field of gene diagnosis, relates to a grading model and application thereof, relates to a grading model for detecting benign and malignant degrees of pancreatic tumors and application thereof, and in particular relates to a grading model of a group of imprinting genes in detecting benign and malignant degrees of pancreatic tumors and a device formed by the grading model.

Background

Pancreatic cancer is a malignant tumor with high malignant degree, the survival rate of 5 years is less than 5 percent, and the pancreatic cancer is one of malignant tumors with the fastest progress and worst prognosis. About 33.8 tens of thousands of people worldwide are diagnosed as pancreatic cancer, 33.0 tens of thousands of deaths, 6.6 tens of thousands of new cases per year in China, and 6.4 tens of thousands of deaths, and the incidence rate thereof is also increasing year by year.

Early diagnosis of pancreatic cancer is more difficult, and 80% of pancreatic cancer patients are already advanced at the time of diagnosis and cannot undergo surgery. Early pancreatic cancer has symptoms similar to pancreatitis, is atypical, and is usually easily ignored by patients. Because of the special anatomical location of the pancreas, it presents difficulties in ultrasound examination, and the pancreas also does not have a direct conduit to the outside of the body, and cannot be examined endoscopically. The existing pancreatic cancer tumor markers also have the problem of low sensitivity and specificity. There is a need for a more sensitive and accurate detection means to increase the early diagnosis rate of pancreatic cancer.

Traditional pathology makes a judgment on the benign or malignant diagnosis of cells based on the relationship of cell size, morphology, wettability and surrounding cell tissue. It has a great limitation in the discovery of early changes in cells (cancers), and thus a method for diagnosing cancer at the cellular molecular level has become a research hotspot at one time. With the continuous intensive research in the field of molecular biology, more and more molecular detection techniques are being applied to cancer diagnosis.

Cancer is the generation of uncontrolled cell growth/division caused by epigenetic changes and genetic variations that accumulate over time. Traditional pathological diagnosis makes a judgment on benign and malignant pancreatic tumors based on the size, morphology and structural variation of cells and tissues. With the development and penetration of molecular biology, more and more molecular detection techniques are applied to pancreatic cancer detection. From the analysis of the progression of cancer, changes in molecular level (epigenetic and genetic) are far ahead of variations in cell morphology and tissue structure. Molecular biological assays are therefore more sensitive to early detection of cancer.

Genomic imprinting is one way of gene regulation in epigenetics. It is characterized in that by methylating alleles from specific parents, one gene is expressed while the other is put into a gene silencing state. This kind of gene is called a blot (marker) gene. A print deletion is an epigenetic change in which the print gene demethylation causes the activation of the silent state allele and the initiation of gene expression. Numerous studies have shown that this phenomenon (print loss) is common to all types of cancer and occurs earlier in time than changes in cell and tissue morphology. Meanwhile, in healthy cells, the proportion of imprinting missing is extremely low, in sharp contrast to cancer cells. Therefore, the methylation state of the imprinted gene can be used as a pathological marker, and the abnormal state of the cells can be analyzed by a specific molecular detection technology.

For the above reasons, current pancreatic cancer diagnosis requires new detection systems and detection models that resolve the molecular marker changes that exist at the cellular level of pancreatic cancer based on patient biopsy samples, thereby providing more accurate prognosis and diagnostic information.

Disclosure of Invention

Aiming at the defects and actual demands of the prior art, the invention provides a grading model for detecting the benign and malignant degree of pancreatic tumors and application thereof, wherein the detection device and the model are used for intuitively observing the change of imprinting (trace) genes of the pancreatic tumors at the single cell and tissue level in early stage so as to judge the benign and malignant degree of the pancreatic tumors.

In order to achieve the above purpose, the invention adopts the following technical scheme:

in a first aspect, the present invention provides a imprinted gene classification model for pancreatic tumors, which classifies the expression state of imprinted genes by calculating the total expression amount of imprinted genes, the deletion expression amount of imprinted genes, and the variation of the copy number abnormal expression amount of imprinted genes in pancreatic cancer;

wherein the imprinting gene is any one or the combination of at least two of Z1, Z5, Z10, Z11 or Z16, the imprinting gene Z1 is Gnas, the imprinting gene Z5 is Mest, the imprinting gene Z10 is Gatm, the imprinting gene Z11 is Grb10, and the imprinting gene Z16 is Snrpn/Snurf.

In the present invention, the inventors found that the diagnostic sensitivity to pancreatic cancer can be up to 86.7% or more by calculating the imprinted gene deletion expression level and the imprinted gene copy number abnormal expression level of any one of the imprinted genes Z1, Z5, Z10, Z11 and Z16 in pancreatic tumors.

According to the present invention, if only one imprinted gene is detected by preliminary detection, any one of Z1, Z5, Z10, Z11 and Z16, preferably any one of Z1, Z11 or Z16, and more preferably Z11 or Z16, can be detected.

In the present invention, the inventors found that if one Z1 imprinting gene was detected alone, the diagnostic sensitivity to pancreatic cancer could be 88.9%, if one Z5 imprinting gene was detected alone, the diagnostic sensitivity to pancreatic cancer could be 86.7%, if one Z10 imprinting gene was detected alone, the diagnostic sensitivity to pancreatic cancer could be 86.7%, if one Z11 imprinting gene was detected alone, the diagnostic sensitivity to pancreatic cancer could be 91.1%, and if one Z16 imprinting gene was detected alone, the diagnostic sensitivity to pancreatic cancer could be 91.1%.

According to the invention, the method for calculating the imprinting gene by the model comprises the following steps: if a combination of two imprinted genes of the imprinted genes is detected, the combination may be any two of Z1, Z5, Z10, Z11 and Z16, preferably a combination of Z1 and Z10, a combination of Z5 and Z11, a combination of Z10 and Z16 or a combination of Z11 and Z16.

In the present invention, the inventors found that sensitivity can be further improved by calculating the total expression amount of two or more imprinted genes, the amount of expressed imprinted gene deletion and the amount of abnormally expressed imprinted gene copy number, that the diagnostic sensitivity to pancreatic cancer can be 93.3% or more by detecting the combination of any two imprinted genes among imprinted genes Z1, Z5, Z10, Z11 and Z16, that the diagnostic sensitivity to pancreatic cancer can be 97.8% or more by detecting the combination of Z1 and Z10, the combination of Z11 and Z16, and that the diagnostic sensitivity to pancreatic cancer can be 99.0% or more by detecting the combination of Z5 and Z11, and the combination of Z10 and Z16.

According to the invention, the imprinted gene further comprises any one or a combination of at least two of Z2, Z3, Z4, Z6, Z8, Z9, Z12, Z13, Z14 or Z15; the marking gene Z2 is Igf2, the marking gene Z3 is Peg10, the marking gene Z4 is Igf2r, the marking gene Z6 is Plagl1, the marking gene Z8 is Dcn, the marking gene Z9 is Dlk1, the marking gene Z12 is Peg3, the marking gene Z13 is Sgce, the marking gene Z14 is Slc38a4, and the marking gene Z15 is Diras3.

In the invention, the inventor finds that on the basis of using the Z1, Z5, Z10, Z11 and Z16 genes for detection, Z2, Z3, Z4, Z6, Z8, Z9, Z12, Z13, Z14 and Z15 genes are additionally arranged for joint diagnosis, so that not only is the detection accuracy improved, but also the occurrence of false positives can be further avoided by adding other probes for auxiliary diagnosis, the detection accuracy can be further improved, and the accurate classification and judgment of all pancreatic tumor samples can be realized.

According to the invention, the method for calculating the imprinting gene by the model comprises the following steps: combinations of the imprinted genes were calculated, and combinations of the Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15, and Z16 genes were calculated.

In the invention, the imprinted gene is deleted, two red/brown marks exist in the nucleus after the cells are subjected to hematoxylin staining, the imprinted gene copy number abnormality is that more than two red/brown marks exist in the nucleus after the cells are subjected to hematoxylin staining, and the copy number abnormality is caused by abnormal gene replication of cancer cells, so that the gene appears as triploid or even higher polyploid during expression.

In the present invention, the imprinting gene and the imprinting gene are the same concept, and the same meaning is expressed, and substitution is possible.

Preferably, the formulas for calculating the total expression amount of the imprinted gene, the deletion expression amount of the imprinted gene, and the copy number abnormal expression amount of the imprinted gene are as follows:

total expression = (b+c+d)/(a+b+c+d) ×100%;

normal imprinted gene expression amount = b/(b+c+d) ×100%;

imprinted gene deleted gene expression amount (LOI) =c/(b+c+d) ×100%;

gene expression level (CNV) of the imprinted gene copy number abnormality=d/(b+c+d) ×100%;

Wherein a is a cell nucleus in which no marker exists in the cell nucleus and the imprinted gene is not expressed after the cell is subjected to hematoxylin staining; b is a cell nucleus with a red/brown mark in the cell nucleus and a marking gene after the cell is subjected to hematoxylin staining; c is a cell nucleus with two red/brown marks in the cell nucleus and marking the gene deletion after the cell is subjected to hematoxylin staining; and d is a cell nucleus with more than two red/brown marks in the cell nucleus and abnormal imprinted gene copy number after the cell is subjected to hematoxylin staining.

In the invention, the hematoxylin-stained marker is selected from red or brown, and the stained marker with other colors can be used for calculating the expression level of the imprinting gene, the deletion expression level of the imprinting gene and the abnormal expression level of the copy number of the imprinting gene.

In the invention, the probe is used for judging whether the imprinting gene exists, the imprinting gene is missing or the copy number is abnormal in each cell nucleus through in-situ hybridization and a Hemotoxy (hematoxylin) cell nucleus staining amplification signal under a 40-multiplied or 60-multiplied microscope, and the tumor benign and malignant degree of the sample is judged through calculating the gene expression quantity of the imprinting gene missing gene and the gene expression quantity of the imprinting gene copy number abnormality. Since the sections were only 10 μm, approximately 20% of the nuclei seen under the microscope were incomplete nuclei, i.e. there was a possibility of partial false negatives.

According to the present invention, the total imprinted gene expression level, the imprinted gene deletion expression level, and the imprinted gene copy number abnormal expression level are classified into five different levels, and at least 1200 cells are counted in the most positive region of sample expression by each probe, and five different levels are respectively classified for the imprinted gene deletion expression level, the imprinted gene copy number abnormal expression level, and the imprinted gene total expression level of fifteen imprinted genes of Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15, and Z16.

According to the present invention, the five different grades of the imprinted gene deletion expression level, the imprinted gene copy number abnormal expression level and the total expression level for Z1 and Z11 are:

level 0: any one of or a combination of at least two of a imprinting gene deletion expression amount of the imprinting genes Z1 and Z11 of less than 15%, a imprinting gene copy number abnormal expression amount of the imprinting genes Z1 and Z11 of less than 1.5%, or a total expression amount of the imprinting genes Z1 and Z11 of less than 25%;

stage I: the imprinted gene deletion expression amount of the imprinted genes Z1 and Z11 is 15-20%, the imprinted gene copy number abnormal expression amount of the imprinted genes Z1 and Z11 is 1.5-3%, or the total expression amount of the imprinted genes Z1 and Z11 is any one or a combination of at least two of 25-35%;

Stage II: the imprinted gene deletion expression amount of the imprinted genes Z1 and Z11 is 20-25%, the imprinted gene copy number abnormal expression amount of the imprinted genes Z1 and Z11 is 3-5%, or the total expression amount of the imprinted genes Z1 and Z11 is any one or a combination of at least two of 35-45%;

class III: the imprinted gene deletion expression amount of the imprinted genes Z1 and Z11 is 25-30%, the imprinted gene copy number abnormal expression amount of the imprinted genes Z1 and Z11 is 5-8%, or the total expression amount of the imprinted genes Z1 and Z11 is any one or a combination of at least two of 45-55%;

grade IV: any one or a combination of at least two of the imprinted gene deletion expression amount of the imprinted genes Z1 and Z11 being more than 30%, the imprinted gene copy number abnormal expression amount of the imprinted genes Z1 and Z11 being more than 8%, or the total expression amount of the imprinted genes Z1 and Z11 being more than 55%;

in the present invention, the imprinted gene deletion expression level, the imprinted gene copy number abnormal expression level and the total expression level of the imprinted genes Z1 and Z11 are independent of each other.

According to the present invention, the five different grades of the imprinted gene deletion expression level, the imprinted gene copy number abnormal expression level and the total expression level for Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z12, Z13, Z14, Z15 and Z16 are divided into:

Level 0: the imprinted genes Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z12, Z13, Z14, Z15 and Z16 have a imprinted gene deletion expression level of less than 11%, the imprinted genes Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z12, Z13, Z14, Z15 and Z16 have an imprinted gene copy number abnormal expression level of less than 1.5%, or the imprinted genes Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z12, Z13, Z14, Z15 and Z16 have a total expression level of less than 20% or a combination of at least two;

stage I: the imprinted genes Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z12, Z13, Z14, Z15 and Z16 have a deletion expression level of 11-15%, the imprinted genes Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z12, Z13, Z14, Z15 and Z16 have an abnormal expression level of 1.5-2.5% of the imprinted gene copy number or the imprinted genes Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z12, Z13, Z14, Z15 and Z16 have a total expression level of 20-30% or a combination of at least two;

stage II: the imprinted genes Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z12, Z13, Z14, Z15 and Z16 have a deletion expression level of 15 to 20%, the imprinted genes Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z12, Z13, Z14, Z15 and Z16 have an abnormal expression level of 2.5 to 4% of the imprinted gene copy number or the imprinted genes Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z12, Z13, Z14, Z15 and Z16 have a total expression level of 30 to 40% or a combination of at least two thereof;

Class III: the imprinted genes Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z12, Z13, Z14, Z15 and Z16 have a deletion expression level of 20 to 25%, the imprinted genes Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z12, Z13, Z14, Z15 and Z16 have an abnormal expression level of 4 to 6% of the imprinted gene copy number or the imprinted genes Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z12, Z13, Z14, Z15 and Z16 have a total expression level of 40 to 50% or a combination of at least two;

grade IV: the imprinted genes Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z12, Z13, Z14, Z15 and Z16 have a imprinting gene deletion expression level of more than 25%, the imprinted genes Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z12, Z13, Z14, Z15 and Z16 have an imprinting gene copy number abnormal expression level of more than 6% or the imprinted genes Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z12, Z13, Z14, Z15 and Z16 have a total expression level of more than 50% or a combination of at least two;

in the present invention, the imprinted gene deletion expression level, the imprinted gene copy number abnormal expression level and the total expression level of the imprinted genes Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z12, Z13, Z14, Z15 and Z16 are independent of each other.

In a second aspect, the present invention provides an apparatus for detecting benign and malignant pancreatic tumor, comprising the following units:

(1) Sampling unit: obtaining a sample to be tested;

(2) Probe design unit: designing a specific primer according to the imprinting gene sequence;

(3) And a detection unit: performing in situ hybridization on the probe in the step (2) and a sample to be detected;

(4) Analysis unit: microscopic imaging analysis of the expression of the imprinted gene;

wherein the analysis unit judges the benign and malignant degree of the pancreatic tumor by calculating the total expression amount of the imprinted gene, the abnormal expression amount of the imprinted gene deletion and the abnormal expression amount of the imprinted gene copy number through the model described in the first aspect, thereby judging the benign and malignant degree of the pancreatic tumor by the grades of the abnormal expression amount of the imprinted gene deletion, the abnormal expression amount of the imprinted gene copy number and the total expression amount.

In the invention, the imprinted gene is deleted, two red/brown marked cell nuclei exist in the cell nuclei after the cells are subjected to hematoxylin staining, the imprinted gene copy number abnormality is the condition that more than two red/brown marked cell nuclei exist in the cell nuclei after the cells are subjected to hematoxylin staining, and the copy number abnormality is caused by abnormal gene replication of cancer cells, so that the gene appears as triploid or even higher polyploid when expressed.

In the invention, the hematoxylin-stained marker is selected from red or brown, and the stained marker with other colors can be used for calculating the total expression quantity of the imprinted gene, the deletion expression quantity of the imprinted gene and the abnormal expression quantity of the imprinted gene copy number.

The detection device is used for intuitively observing the change of the imprinting (trace) genes of pancreatic tumors at the early stage under the cellular and tissue level so as to judge the benign and malignant degree of the tumors, and provides the most favorable treatment opportunity for early-stage pancreatic tumor patients.

According to the invention, the sample to be tested in step (1) is derived from human tissue and/or cells.

In the invention, the sample to be detected can be selected according to the needs of a person skilled in the art as long as the RNA is subjected to timely fixation, and the sample to be detected comprises any one or a combination of at least two of paraffin sections and puncture biopsy samples of tissues.

The paraffin section of the tissue comprises the specific operation steps of obtaining a human tumor tissue sample, fixing the tissue sample by 10% neutral formalin in time, embedding the tissue sample by paraffin, cutting the tissue sample into 10 mu m thick, and preparing a tissue sheet by using a positively charged slide; since only 10 μm thick, a part of the nuclei are incomplete under the microscope, so that a partially false negative gene deletion occurs.

The specific operation steps of the biopsy sample are that human cells are obtained through puncturing, and the biopsy sample is fixed by 10% neutral formalin in time.

In the invention, because the puncture biopsy has little harm to a patient, the sampling process is simple, compared with the blood circulation characteristic, the puncture biopsy can be positioned, and the puncture biopsy has special advantages as an experimental sample.

Preferably, the sample to be tested is a puncture biopsy sample.

Preferably, the imprinting genes are Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16, the imprinting gene Z1 is Gnas, the imprinting gene Z2 is Igf2, the imprinting gene Z3 is Peg10, the imprinting gene Z4 is Igf2r, the imprinting gene Z5 is Mest, the imprinting gene Z6 is Plagl1, the imprinting gene Z8 is Dcn, the imprinting gene Z9 is Dlk1, the imprinting gene Z10 is Gatm, the imprinting gene Z11 is Grb10, the imprinting gene Z12 is Peg3, the imprinting gene Z13 is Sgce, the imprinting gene Z14 is Slc38a4, the imprinting gene Z15 is dig 3, and the imprinting gene Z16 is snn/Snurf.

In the invention, the imprinting genes Z1 (Gnas), Z2 (Igf 2), Z3 (Peg 10), Z4 (Igf 2 r), Z5 (Mest), Z6 (Plagl 1), Z8 (Dcn), Z9 (Dlk 1), Z10 (Gatm), Z11 (Grb 10), Z12 (Peg 3), Z13 (Sgce), Z14 (Slc 38a 4), Z15 (dirac 3) and Z16 (Snrpn/Snurf) are expressed in normal tumor cell tissues to different degrees, and the expression quantity and imprinting state are obviously changed when malignant lesions occur.

In the present invention, the design probes were designed based on the imprinted genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16, that is, gnas, igf2, peg10, igf2r, test, plagl1, dcn, dlk1, gatm, grb10, peg3, sgce, slc38a4, diras3 and Snrpn/Snurf, specifically, a sequence within the intron of each gene was selected as a probe, and a specific probe was designed by Advanced Cell Diagnostics company.

Preferably, the in situ hybridization employs an RNAscope in situ hybridization method.

Preferably, the RNAscope in situ hybridization method uses a single-channel or multi-channel chromogenic kit or a single-channel or multi-channel fluorescent kit, preferably a single-channel red/brown chromogenic kit or a multi-channel fluorescent kit.

In the invention, the multi-channel coloring kit or the multi-channel fluorescent kit comprises two or more than two channels of coloring kits or fluorescent kits, and the two channels of coloring kits or the multi-channel fluorescent kits can use two imprinting gene probes or the joint expression of imprinting genes and other genes or even the comprehensive expression of a plurality of imprinting genes and non-imprinting genes.

According to the present invention, the formulas for calculating the total expression amount of the imprinted gene, the deletion expression amount of the imprinted gene and the copy number abnormal expression amount of the imprinted gene in the model are as follows:

total expression = (b+c+d)/(a+b+c+d) ×100%;

normal imprinted gene expression amount = b/(b+c+d) ×100%;

imprinted gene deleted gene expression amount (LOI) =c/(b+c+d) ×100%;

In the invention, the probe is used for judging whether the imprinting gene exists, the imprinting gene is missing or the copy number is abnormal in each cell nucleus through in-situ hybridization and a Hemotoxy (hematoxylin) cell nucleus staining amplification signal under a 40-multiplied or 60-multiplied microscope, and the tumor benign and malignant degree of the sample is judged through calculating the total imprinting gene expression quantity, the imprinting gene missing gene expression quantity and the gene expression quantity with abnormal imprinting gene copy number. Since the sections were only 10 microns, approximately 20% of the nuclei seen under the microscope were incomplete nuclei, i.e. there was a possibility of partial false negatives.

According to the present invention, the imprinted gene deletion expression level, imprinted gene copy number abnormal expression level and total expression level are classified into five different levels.

The five different grades are that at least 1200 cells are counted in a region where each probe of the sample expresses most positively, and the imprinted gene deletion expression amounts, imprinted gene copy number abnormality expression amounts, and total expression amounts of fifteen imprinted genes for Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15, and Z16 are respectively divided.

The five different grades of the deletion expression quantity of the imprinting genes, the abnormal expression quantity of the copy number of the imprinting genes and the total expression quantity aiming at Z1 and Z11 are divided into:

The five different grades of the deletion expression quantity of the imprinting genes, the abnormal expression quantity of the copy number of the imprinting genes and the total expression quantity of the imprinting genes for Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z12, Z13, Z14, Z15 and Z16 are divided into:

According to the present invention, the judging of benign and malignant degrees of pancreatic tumors is classified into benign tumors, pancreatic cancer potential, early-stage pancreatic cancer, medium-stage pancreatic cancer and advanced-stage pancreatic cancer;

preferably, the result of the judgment of the degree of benign or malignant pancreatic tumor is that the imprinting gene deletion expression level of not more than 1 imprinting gene among imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 and the imprinting gene copy number abnormal expression level are both less than level I or that the imprinting gene copy number abnormal expression level of not more than 1 imprinting genes among imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is level I and that the imprinting gene copy number abnormal expression level of not more than 1 imprinting genes among imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is level I;

preferably, the result of the judgment of the degree of benign or malignant pancreatic tumor is that the imprinting gene copy number abnormal expression amount of at least 2 imprinting genes of the imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is level I, the imprinting gene deletion expression amount of not more than 1 imprinting gene of the imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is level I, or the imprinting gene copy number abnormal expression amount of at least 2 imprinting genes of the imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is level II, and the imprinting gene copy number abnormal expression amount of not more than 1 imprinting genes of the imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z15 and Z16 is level II, the imprinting gene copy number of the imprinting gene is not more than one of the pancreatic tumor;

Preferably, the result of the judgment of the degree of benign or malignant pancreatic tumor is that the amount of defective expression of the imprinted gene of at least 2 imprinted genes among the imprinted genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is class II, or that the amount of defective expression of the imprinted gene of at least 2 imprinted genes among the imprinted genes Z1, Z2, Z3, Z4, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is class III, and the amount of defective expression of at least 1 imprinted gene among the imprinted genes is class III, and the amount of defective expression of at least one of early stage of the imprinted genes among the imprinted genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z11, Z13, Z14 and Z16 is class III, and the amount of defective expression of at least one of the imprinted genes among the imprinted genes of Z1, Z16 is class III;

preferably, the result of the judgment of the degree of benign or malignant pancreatic tumor is that the imprinting gene copy number abnormal expression amount of at least 2 imprinting genes of the imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is class III, the imprinting gene deletion expression amount of not more than 1 imprinting gene of the imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is class III, or the imprinting gene copy number abnormal expression amount of at least 2 imprinting genes of the imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is class IV, and the imprinting gene copy number abnormal expression amount of not more than 1 imprinting genes of the imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z15 and Z16 is class IV, and the imprinting gene copy number of not more than one of the imprinting genes of the pancreatic tumor is class IV;

Preferably, the result of the determination of the degree of benign or malignant pancreatic tumor is that the imprinting gene deletion expression level of at least 2 imprinting genes among imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is IV-level or that the imprinting gene copy number abnormal expression level of at least 2 imprinting genes among imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is IV-level, and the pancreatic cancer is advanced.

In a third aspect, the present invention provides a model as described in the first aspect or a device as described in the second aspect for use in the manufacture of a medicament or kit for pancreatic tumour detection and/or treatment.

According to the present invention, judging the benign and malignant degree of pancreatic tumor is classified into benign tumor, pancreatic cancer potential, early pancreatic cancer, medium pancreatic cancer and late pancreatic cancer;

according to the present invention, the result of judging the benign and malignant degree of pancreatic tumor is that the imprinting gene deletion expression level of not more than 1 imprinting genes among imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 and the imprinting gene copy number abnormal expression level are both less than level I or that the imprinting gene copy number abnormal expression level of not more than 1 imprinting genes among imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is level I and that the imprinting gene copy number abnormal expression level of not more than 1 imprinting genes among imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is level I;

Compared with the prior art, the invention has the following beneficial effects:

(1) The detection model and the device of the invention show the appearance of imprinting missing on the sample of pancreatic tumor patients in an intuitive way, objectively, intuitively, early and accurately detect the change of imprinting (trace) genes by the method of marking imprinting genes in situ, and can provide a quantized model, thereby greatly contributing to the diagnosis of pancreatic tumor;

(2) The detection device can obtain the judgment of the benign and malignant degree of the pancreatic tumor by puncturing cells before the operation of the pancreatic tumor patient, thereby providing a basis for the operation and the accurate treatment, and being a revolutionary breakthrough in the field of cell molecules for diagnosing the pancreatic tumor;

(3) The invention can sensitively detect early pancreatic cancer, can detect samples beginning to generate canceration from pancreatic precancerous lesions patients such as pancreatic intraepithelial neoplasia, chronic pancreatitis, intraductal Papillary Myxoma (IPMN), myxocystic tumor and the like through the combined detection of imprinting genes, has little harm to the patients by pancreatic puncture and simple material obtaining process, is used for early general investigation and follow-up after cancer operation, particularly for follow-up for suspected recurrent patients, can strive for time and makes great contribution to saving the lives of the patients;

(4) The technology of the invention can be used for detecting tumors of pancreatic exocrine parts, can also be used for detecting benign and malignant tumors of islet neuroendocrine tumors, has important guiding significance for accurate judgment of pancreatic cancer types and selection of treatment methods, and greatly reduces postoperative recurrence of pancreatic cancer.

(5) The detection method is different from an immunohistochemical method, false positive and other negative effects are reduced, and moreover, the target medicine or technical method for silencing, eliminating and rearranging the gene through the discovered pancreatic tumor related imprinted gene deletion site can be used for guiding later treatment and medication.

Drawings

FIG. 1 is a pathological section of pancreatic cancer of a hematoxylin stained cell nucleus according to the present invention, wherein a is that after the cell is hematoxylin stained, there is no marker in the cell nucleus and the imprinted gene is not expressed; the step b is that after the cells are subjected to hematoxylin staining, a red/brown mark exists in the nuclei of the cells, and a mark gene exists; the step c is that after the cells are subjected to hematoxylin staining, two red/brown marks exist in the nuclei of the cells, and the imprinted genes are deleted; the step d is that after the cells are subjected to hematoxylin staining, more than two red/brown marks exist in the nuclei of the cells, and the number of copies of the imprinted genes is abnormal;

Fig. 2 (a) is the expression state of 15 genes in the pathological section of the 0-grade pancreatic tumor, fig. 2 (b) is the expression state of 15 genes in the pathological section of the I-grade pancreatic cancer, fig. 2 (c) is the expression state of 15 genes in the pathological section of the II-grade pancreatic cancer, fig. 2 (d) is the expression state of 15 genes in the pathological section of the III-grade pancreatic cancer, and fig. 2 (e) is the expression state of 15 genes in the pathological section of the IV-grade pancreatic cancer;

FIG. 3 (a) shows the intensities of the imprinting deletions of the imprinting genes Z1, Z5, Z10, Z11 and Z16 on pancreatic cancer, FIG. 3 (b) shows the intensities of the copy number abnormalities of the imprinting genes Z1, Z5, Z10, Z11 and Z16 on pancreatic cancer, FIG. 3 (c) shows the intensities of the total expression amounts of the imprinting genes Z1, Z5, Z10, Z11 and Z16 on pancreatic cancer, FIG. 3 (d) shows the intensities of the imprinting deletions of the imprinting genes Z2, Z3, Z4, Z6, Z8, Z9, Z12, Z13, Z14 and Z15 on pancreatic cancer, FIG. 3 (e) shows the intensities of the copy number abnormalities of the imprinting genes Z2, Z3, Z6, Z9, Z12, Z13, Z14 and Z15 on pancreatic cancer, FIG. 3 (f) shows the total expression amounts of the imprinting genes Z2, Z3, Z4, Z6, Z13, Z14 and Z15 on pancreatic cancer, wherein the total expression amounts of the genes CNI show the copy number abnormalities of the imprinting genes;

FIG. 4 (a) shows the intensities of the imprinting deletion, the copy number abnormality and the total expression level of the imprinting gene Z1, FIG. 4 (b) shows the intensities of the imprinting deletion, the copy number abnormality and the total expression level of the imprinting gene Z5, FIG. 4 (c) shows the intensities of the imprinting deletion, the copy number abnormality and the total expression level of the imprinting gene Z10, FIG. 4 (d) shows the intensities of the imprinting deletion, the copy number abnormality and the total expression level of the imprinting gene Z11, FIG. 4 (e) shows the intensities of the imprinting deletion, the copy number abnormality and the total expression level of the imprinting gene Z16, FIG. 4 (f) shows the intensities of the imprinting deletion, the copy number abnormality and the total expression level of the imprinting gene Z2, FIG. 4 (g) shows the intensities of the imprinting deletion, the copy number abnormality and the total expression level of the imprinting gene Z3, FIG. 4 (i) is the intensity of the imprinting deletion, the copy number abnormality and the total expression amount of the imprinting gene Z6, FIG. 4 (j) is the intensity of the imprinting deletion, the copy number abnormality and the total expression amount of the imprinting gene Z8, FIG. 4 (k) is the intensity of the imprinting deletion, the copy number abnormality and the total expression amount of the imprinting gene Z9, FIG. 4 (l) is the intensity of the imprinting deletion, the copy number abnormality and the total expression amount of the imprinting gene Z12, FIG. 4 (m) is the intensity of the imprinting deletion, the copy number abnormality and the total expression amount of the imprinting gene Z13, FIG. 4 (n) is the intensity of the imprinting deletion, the copy number abnormality and the total expression amount of the imprinting gene Z14, FIG. 4 (o) is the intensity of the imprinting deletion, the copy number abnormality and the total expression amount of the imprinting gene Z15, LOI is the gene expression quantity of the imprinting gene deletion, CNV is the gene expression quantity of the imprinting gene with abnormal copy number, and TE is the total expression quantity of the imprinting gene;

FIG. 5 (a) shows the distribution range and the classification standard of the marking loss and the copy number abnormality in the 45-case pancreatic cancer pathological section, FIG. 5 (b) shows the distribution range and the classification standard of the marking loss and the copy number abnormality in the 45-case pancreatic cancer pathological section, FIG. 5 (c) shows the distribution range and the classification standard of the marking loss and the copy number abnormality in the 45-case pancreatic cancer pathological section, FIG. 5 (d) shows the distribution range and the classification standard of the marking loss and the copy number abnormality in the 45-case pancreatic cancer pathological section, FIG. 5 (e) shows the marking loss and the copy number abnormality in the 45-case pancreatic cancer pathological section, FIG. 5 (f) shows the marking loss and the distribution range and the copy number abnormality in the 45-case pancreatic cancer pathological section, FIG. 5 (g) shows the marking loss and the copy number abnormality in the 45-case pancreatic cancer pathological section, FIG. 5 (j) shows the marking loss and the distribution range and the copy number abnormality in the pancreatic cancer pathological section, and the marking abnormality in the marking loss of the pancreatic cancer pathological section, FIG. 5 (j) shows the marking loss and the marking loss, FIG. 5 (k) shows the distribution range and the classification standard of the imprinting gene Z9 applied to 45 pancreatic cancer pathological sections, the imprinting gene Z12 applied to 45 pancreatic cancer pathological sections, the imprinting gene Z13 applied to 45 pancreatic cancer pathological sections, the imprinting gene Z14 applied to 45 pancreatic cancer pathological sections, the imprinting gene Z15 applied to 45 pancreatic cancer pathological sections, the imprinting gene Z expressed by LOI, the imprinting gene CNV expressed by the imprinting gene with abnormal copy number, and the TE expressed by the total imprinting gene;

Detailed Description

The technical means adopted by the invention and the effects thereof are further described below by the specific embodiments in combination with the accompanying drawings, but the invention is not limited to the examples.

EXAMPLE 1 imprinted Gene analysis of pancreatic cancer

The detection method of the imprinting gene comprises the following steps:

(1) Obtaining tissue cell sections (10 microns) of pancreatic cancer, placing the sections into 10% neutral formalin solution for fixation to prevent RNA degradation, wherein the fixation time is 24 hours, paraffin embedding (FFPE), the slides need positive charges to be removed from the slides, and the sections are baked for more than 3 hours in a 40 ℃ oven;

(2) Dewaxing according to a sample treatment method of RNASCope, sealing the endogenous peroxidase activity in the sample, enhancing the permeability and exposing RNA molecules;

(3) Designing a probe: designing a specific primer according to the imprinting gene sequence;

the design probes were designed based on imprinting genes Z1 (Gnas), Z2 (Igf 2), Z3 (Peg 10), Z4 (Igf 2 r), Z5 (test), Z6 (Plagl 1), Z8 (Dcn), Z9 (Dlk 1), Z10 (Gatm), Z11 (Grb 10), Z12 (Peg 3), Z13 (Sgce), Z14 (Slc 38a 4), Z15 (Diras 3) and Z16 (Snrpn/Snuf), and a sequence was selected as a probe within the intron of each gene, and the specific probes were designed by Advanced Cell Diagnostics.

(4) Carrying out RNA SCope in situ hybridization on the probe in the step (3) and a sample to be detected through a kit;

(5) Signal amplification and hematoxylin staining, and analyzing the expression condition of the imprinted gene by using a microscope for imaging;

the formula for calculating the total expression quantity of the imprinting genes, the deletion expression quantity of the imprinting genes and the abnormal expression quantity of the copy number of the imprinting genes in the model is as follows:

total expression = (b+c+d)/(a+b+c+d) ×100%;

normal imprinted gene expression amount = b/(b+c+d) ×100%;

imprinted gene deleted gene expression amount (LOI) =c/(b+c+d) ×100%;

as shown in fig. 1, a, b, c, d, a is a cell nucleus with no marker in the cell nucleus and no expressed imprinted gene after the cell is subjected to hematoxylin staining; b is a cell nucleus with a red/brown mark in the cell nucleus and a marking gene after the cell is subjected to hematoxylin staining; c is a cell nucleus with two red/brown marks in the cell nucleus and marking the gene deletion after the cell is subjected to hematoxylin staining; and d is a cell nucleus with more than two red/brown marks in the cell nucleus and abnormal imprinted gene copy number after the cell is subjected to hematoxylin staining.

From FIGS. 2 (a) -2 (e), it can be seen that the proportion of cells with a loss of imprint (two signal points in the nucleus) and an abnormality in copy number (three or more signal points in the nucleus) gradually increases with the increase in malignancy in the samples from grade 0 to grade IV.

EXAMPLE 2 imprinted Gene analysis of puncture biopsy samples

The biopsy sample was obtained by taking out suspicious lesion tissue by puncturing, and fixing the tissue with 10% neutral formalin solution for 24 hours or longer, and the other detection method was the same as in example 1.

As can be seen from fig. 3 (a) -3 (f), the sensitivity of each gene Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15, Z16 to pancreatic cancer or the intensity and status of the absence of the imprint corresponding to pancreatic cancer expression are different.

Specifically, the sensitivity of each imprinting gene to pancreatic cancer is as shown in fig. 4 (a) -4 (o), and as can be seen from fig. 4 (a) -4 (e), the imprinting deletion, copy number abnormality and expression amount increase of imprinting gene Z1 are obviously increased in the malignant potential stage, slightly decreased in the early pancreatic cancer stage, rapidly increased to a very high level in the metaphase pancreatic cancer stage and stably maintained in the late pancreatic cancer stage; the imprinting deletion and the expression quantity increase of the imprinting gene Z5 are rapidly increased in the malignant potential stage, stably maintained in the early pancreatic cancer stage, continuously increased to a very high level in the middle pancreatic cancer stage, continuously maintained in the late pancreatic cancer stage, rapidly increased in the malignant potential stage, stably maintained in the early and middle pancreatic cancer stages, and gradually increased to a very high level in the late pancreatic cancer stage; the imprinting deletion of the imprinting gene Z10 begins to appear in the malignant potential stage, does not rise obviously in early pancreatic cancer, rises to a very high level gradually in middle and late pancreatic cancer, and the imprinting deletion, the copy number abnormality and the expression quantity increase of the imprinting gene Z10 begin to appear in the malignant potential stage and rise to a very high level gradually in the development process of early to late pancreatic cancer; the imprinting deletion, copy number abnormality and expression amount increase of the imprinting gene Z11 rapidly increase in the malignant potential stage, slow down the increase in the early pancreatic cancer stage, rapidly increase in the middle pancreatic cancer stage and maintain a high level in the late pancreatic cancer; the imprinting deletion and the increase of the expression quantity of the imprinting gene Z16 begin to appear in the malignant potential stage, gradually rise to a very high level in early-stage pancreatic cancer and metaphase pancreatic cancer, continue to maintain stability until the late-stage pancreatic cancer stage, begin to appear in the abnormal malignant potential stage of the copy number of the imprinting gene Z16, rapidly rise in early-stage pancreatic cancer, and slow down in the middle-stage pancreatic cancer and the late-stage pancreatic cancer, and reach a very high level;

As can be seen from fig. 4 (f) -4 (o), the imprinting deletion and copy number abnormality of the imprinting gene Z2 began to appear at the stage of intermediate pancreatic cancer, rapidly increased to a higher level at the stage of advanced pancreatic cancer, and the increase in the expression level of the imprinting gene Z2 rapidly increased to a higher level at the stage of intermediate pancreatic cancer, and remained stable at the stage of advanced pancreatic cancer; the imprinting deletion and the expression quantity increase of the imprinting gene Z3 begin to appear in the stage of the middle-stage pancreatic cancer, rapidly rise to a higher level in the stage of the late-stage pancreatic cancer, and the copy number abnormality of the imprinting gene Z3 begins to appear in the stage of the early-stage pancreatic cancer, remains stable in the stage of the middle-stage pancreatic cancer, and rapidly rise to a very high level until the stage of the late-stage pancreatic cancer; the imprinting deletion of the imprinting gene Z4 rapidly rises in early pancreatic cancer stages, continues to rise to higher levels in mid-and late-stage pancreatic cancer, the copy number abnormality of the imprinting gene Z4 begins to appear in mid-stage pancreatic cancer stages, rapidly rises to higher levels in late-stage pancreatic cancer stages, the increase in the expression level of the imprinting gene Z4 begins to appear in early pancreatic cancer stages, rapidly rises to higher levels in mid-stage pancreatic cancer stages, and continues to be maintained in late-stage pancreatic cancer stages; the imprinting deletion and copy number abnormality of the imprinting gene Z6 begin to appear in the imprinting deletion stage, gradually rise to a very high level in the progress of early-stage to late-stage pancreatic cancer, and the increase in the expression level of the imprinting gene Z6 begins to appear in the early-stage pancreatic cancer stage, and rapidly rise to a very high level in the middle-stage and late-stage pancreatic cancer stages; the imprinting deletion, copy number abnormality and expression amount increase of the imprinting gene Z8 are rapidly increased in the stage of metaphase pancreatic cancer and are maintained stable in the stage of late pancreatic cancer; the imprinting deletion, copy number abnormality and expression amount increase of the imprinting gene Z9 are rapidly increased in the stage of metaphase pancreatic cancer and are maintained stable in the stage of late pancreatic cancer; the imprinting deletion of the imprinting gene Z12 begins to appear in early pancreatic cancer stages, rises rapidly to higher levels in mid-pancreatic cancer stages, remains stable in late pancreatic cancer stages, and the copy number abnormality and the increase in the expression amount of the imprinting gene Z12 begin to appear in early pancreatic cancer stages, and rises gradually to higher levels in mid-and late pancreatic cancer stages; the imprinting deletion and the increase of the expression quantity of the imprinting gene Z13 begin to appear in early pancreatic cancer stages, rapidly rise to high levels in intermediate pancreatic cancer stages, remain stable in late pancreatic cancer stages, and the abnormality of the copy number of the imprinting gene Z13 begins to appear in early pancreatic cancer stages, and gradually rise to high levels in intermediate and late pancreatic cancer stages; the imprinting deletion and copy number abnormality of the imprinting gene Z14 begin to appear in the malignant potential stage, rapidly rise in the early pancreatic cancer stage, slow down in the rising speed in the middle pancreatic cancer stage, rapidly rise to a higher level in the late pancreatic cancer stage, and increase in the expression level of the imprinting gene Z14 begins to appear in the early pancreatic cancer stage, slowly rise in the middle pancreatic cancer stage, and rapidly rise to a higher level in the late pancreatic cancer stage; the imprinting deletion of the imprinting gene Z15 begins to appear in early pancreatic cancer stages, gradually rises to higher levels in mid-and late-stage pancreatic cancer stages, the copy number abnormality of the imprinting gene Z15 begins to appear in malignant potential stages, gradually rises to higher levels in the progress of early to late-stage pancreatic cancer, the increase in the expression level of the imprinting gene Z15 begins to appear in early pancreatic cancer stages, rapidly rises to higher levels in mid-stage pancreatic cancer stages, and remains stable in late-stage pancreatic cancer stages.

EXAMPLE 3 imprinting Gene analysis of 45 pancreatic tumor samples

Obtaining the tissue of 45 pancreatic cancer patients comprises puncturing biopsy samples, and the detection method is the same as that of the embodiment 1.

As can be seen from FIGS. 5 (a) -5 (o), the ratio of the imprinting deletion and the copy number abnormality of 15 probes in 45 pancreatic tumor tissue samples is distributed from low to high, and according to the distribution trend of different probes, the classification standard shown by the broken line in the graph is calculated, so that the imprinting deletion and the copy number abnormality of each probe can be respectively classified into 5 grades from low to high.

The specific classification is as follows:

as can be seen from fig. 5 (a), for the imprinted gene Z1, any one or a combination of at least two of the imprinted gene deletion expression level of less than 15%, the imprinted gene copy number abnormal expression level of less than 1.5%, or the imprinted gene total expression level of less than 25% is 0 level, any one or a combination of at least two of the imprinted gene deletion expression level of 15 to 20%, the imprinted gene copy number abnormal expression level of 1.5 to 3%, or the imprinted gene total expression level of 25 to 35% is I level, the imprinted gene deletion expression level of 20 to 25%, the imprinted gene copy number abnormal expression level of 3 to 5%, or the imprinted gene total expression level of 35 to 45% is II level, the imprinted gene deletion expression level of 25 to 30%, the imprinted gene copy number abnormal expression level of 5 to 8%, or the imprinted gene total expression level of 45 to 55% is III level, the abnormal expression level of more than 30%, the imprinted gene copy number of greater than 8%, or the combination of any one or a combination of at least two of the imprinted gene expression levels of greater than 55% is IV level;

As can be seen from fig. 5 (b), for the imprinted gene Z5, any one or a combination of at least two of the imprinted gene deletion expression level of less than 15%, the imprinted gene copy number abnormal expression level of less than 1.5%, or the imprinted gene total expression level of less than 25% is 0 level, any one or a combination of at least two of the imprinted gene deletion expression level of 15 to 20%, the imprinted gene copy number abnormal expression level of 1.5 to 3%, or the imprinted gene total expression level of 25 to 35% is I level, the imprinted gene deletion expression level of 20 to 25%, the imprinted gene copy number abnormal expression level of 3 to 5%, or the imprinted gene total expression level of 35 to 45% is II level, the imprinted gene deletion expression level of 25 to 30%, the imprinted gene copy number abnormal expression level of 5 to 8%, or the imprinted gene total expression level of 45 to 55% is III level, the abnormal expression level of more than 30%, the imprinted gene copy number of greater than 8%, or the combination of any one or a combination of at least two of the imprinted gene expression levels of greater than 55% is IV level;

as can be seen from fig. 5 (c), for the imprinted gene Z10, any one or a combination of at least two of the imprinted gene deletion expression level of less than 15%, the imprinted gene copy number abnormal expression level of less than 1.5%, or the imprinted gene total expression level of less than 25% is 0 level, any one or a combination of at least two of the imprinted gene deletion expression level of 15 to 20%, the imprinted gene copy number abnormal expression level of 1.5 to 3%, or the imprinted gene total expression level of 25 to 35% is I level, the imprinted gene deletion expression level of 20 to 25%, the imprinted gene copy number abnormal expression level of 3 to 5%, or the imprinted gene total expression level of 35 to 45% is II level, the imprinted gene deletion expression level of 25 to 30%, the imprinted gene copy number abnormal expression level of 5 to 8%, or the imprinted gene total expression level of 45 to 55% is III level, the abnormal expression level of more than 30%, the imprinted gene copy number of greater than 8%, or the combination of any one or a combination of at least two of the imprinted gene expression levels of greater than 55% is IV level;

As can be seen from fig. 5 (d), for the imprinted gene Z11, any one or a combination of at least two of the imprinted gene deletion expression level of less than 15%, the imprinted gene copy number abnormal expression level of less than 1.5%, or the imprinted gene total expression level of less than 25% is 0 level, any one or a combination of at least two of the imprinted gene deletion expression level of 15 to 20%, the imprinted gene copy number abnormal expression level of 1.5 to 3%, or the imprinted gene total expression level of 25 to 35% is I level, the imprinted gene deletion expression level of 20 to 25%, the imprinted gene copy number abnormal expression level of 3 to 5%, or the imprinted gene total expression level of 35 to 45% is II level, the imprinted gene deletion expression level of 25 to 30%, the imprinted gene copy number abnormal expression level of 5 to 8%, or the imprinted gene total expression level of 45 to 55% is III level, the abnormal expression level of more than 30%, the imprinted gene copy number of greater than 8%, or the combination of any one or a combination of at least two of the imprinted gene expression levels of greater than 55% is IV level;

as can be seen from fig. 5 (e), for the imprinted gene Z16, any one or a combination of at least two of the imprinted gene deletion expression level of less than 15%, the imprinted gene copy number abnormal expression level of less than 1.5%, or the imprinted gene total expression level of less than 25% is 0 level, any one or a combination of at least two of the imprinted gene deletion expression level of 15 to 20%, the imprinted gene copy number abnormal expression level of 1.5 to 3%, or the imprinted gene total expression level of 25 to 35% is I level, the imprinted gene deletion expression level of 20 to 25%, the imprinted gene copy number abnormal expression level of 3 to 5%, or the imprinted gene total expression level of 35 to 45% is II level, the imprinted gene deletion expression level of 25 to 30%, the imprinted gene copy number abnormal expression level of 5 to 8%, or the imprinted gene total expression level of 45 to 55% is III level, the abnormal expression level of more than 30%, the imprinted gene copy number of greater than 8%, or the combination of any one or a combination of at least two of the imprinted gene expression levels of greater than 55% is IV level;

As can be seen from fig. 5 (f), for the imprinted gene Z2, any one or a combination of at least two of the imprinted gene deletion expression level of less than 11%, the imprinted gene copy number abnormal expression level of less than 1.5% or the imprinted gene total expression level of less than 20% is level 0, any one or a combination of at least two of the imprinted gene deletion expression level of 11 to 15%, the imprinted gene copy number abnormal expression level of 1.5 to 2.5% or the imprinted gene total expression level of 20 to 30% is level I, the imprinted gene deletion expression level of 15 to 20%, the imprinted gene copy number abnormal expression level of 2.5 to 4% or the imprinted gene total expression level of 30 to 40% is level II, the imprinted gene deletion expression level of 20 to 25%, the imprinted gene copy number abnormal expression level of 4 to 6% or the imprinted gene total expression level of 40 to 50% is level III, the gene deletion expression level of more than 25%, the imprinted gene copy number abnormal expression level of more than 6% or the combination of at least 50% is level IV;

as can be seen from fig. 5 (g), for the imprinted gene Z3, any one or a combination of at least two of the imprinted gene deletion expression level of less than 11%, the imprinted gene copy number abnormal expression level of less than 1.5% or the imprinted gene total expression level of less than 20% is level 0, any one or a combination of at least two of the imprinted gene deletion expression level of 11 to 15%, the imprinted gene copy number abnormal expression level of 1.5 to 2.5% or the imprinted gene total expression level of 20 to 30% is level I, the imprinted gene deletion expression level of 15 to 20%, the imprinted gene copy number abnormal expression level of 2.5 to 4% or the imprinted gene total expression level of 30 to 40% is level II, the imprinted gene deletion expression level of 20 to 25%, the imprinted gene copy number abnormal expression level of 4 to 6% or the imprinted gene total expression level of 40 to 50% is level III, the gene deletion expression level of more than 25%, the imprinted gene copy number abnormal expression level of more than 6% or the combination of at least 50% is level IV;

As can be seen from fig. 5 (h), for the imprinted gene Z4, any one or a combination of at least two of the imprinted gene deletion expression level of less than 11%, the imprinted gene copy number abnormal expression level of less than 1.5% or the imprinted gene total expression level of less than 20% is level 0, any one or a combination of at least two of the imprinted gene deletion expression level of 11 to 15%, the imprinted gene copy number abnormal expression level of 1.5 to 2.5% or the imprinted gene total expression level of 20 to 30% is level I, the imprinted gene deletion expression level of 15 to 20%, the imprinted gene copy number abnormal expression level of 2.5 to 4% or the imprinted gene total expression level of 30 to 40% is level II, the imprinted gene deletion expression level of 20 to 25%, the imprinted gene copy number abnormal expression level of 4 to 6% or the imprinted gene total expression level of 40 to 50% is level III, the gene deletion expression level of more than 25%, the imprinted gene copy number abnormal expression level of more than 6% or the combination of at least 50% is level IV;

as can be seen from fig. 5 (I), for the imprinted gene Z6, any one or a combination of at least two of the imprinted gene deletion expression level of less than 11%, the imprinted gene copy number abnormal expression level of less than 1.5%, or the imprinted gene total expression level of less than 20% is level 0, any one or a combination of at least two of the imprinted gene deletion expression level of 11 to 15%, the imprinted gene copy number abnormal expression level of 1.5 to 2.5%, or the imprinted gene total expression level of 20 to 30% is level I, the imprinted gene deletion expression level of 15 to 20%, the imprinted gene copy number abnormal expression level of 2.5 to 4%, or the imprinted gene total expression level of 30 to 40% is level II, the imprinted gene deletion expression level of 20 to 25%, the imprinted gene copy number abnormal expression level of 4 to 6%, or the imprinted gene total expression level of 40 to 50%, any one or a combination of at least two of the imprinted gene copy number abnormal expression levels of more than 25%, the imprinted gene copy number of more than 6%, or the imprinting level of at least 50% is level IV;

As can be seen from fig. 5 (j), for the imprinted gene Z8, any one or a combination of at least two of the imprinted gene deletion expression level of less than 11%, the imprinted gene copy number abnormal expression level of less than 1.5%, or the imprinted gene total expression level of less than 20% is level 0, any one or a combination of at least two of the imprinted gene deletion expression level of 11 to 15%, the imprinted gene copy number abnormal expression level of 1.5 to 2.5%, or the imprinted gene total expression level of 20 to 30% is level I, the imprinted gene deletion expression level of 15 to 20%, the imprinted gene copy number abnormal expression level of 2.5 to 4%, or the imprinted gene total expression level of 30 to 40% is level II, the imprinted gene deletion expression level of 20 to 25%, the imprinted gene copy number abnormal expression level of 4 to 6%, or the imprinted gene total expression level of 40 to 50%, any one or a combination of at least two of the imprinted gene copy number abnormal expression levels of more than 25%, the imprinted gene copy number of more than 6%, or the imprinting level of at least 50% is level IV;

as can be seen from fig. 5 (k), for the imprinted gene Z9, any one or a combination of at least two of the imprinted gene deletion expression level of less than 11%, the imprinted gene copy number abnormal expression level of less than 1.5%, or the imprinted gene total expression level of less than 20% is level 0, any one or a combination of at least two of the imprinted gene deletion expression level of 11 to 15%, the imprinted gene copy number abnormal expression level of 1.5 to 2.5%, or the imprinted gene total expression level of 20 to 30% is level I, the imprinted gene deletion expression level of 15 to 20%, the imprinted gene copy number abnormal expression level of 2.5 to 4%, or the imprinted gene total expression level of 30 to 40% is level II, the imprinted gene deletion expression level of 20 to 25%, the imprinted gene copy number abnormal expression level of 4 to 6%, or the imprinted gene total expression level of 40 to 50%, any one or a combination of at least two of the imprinted gene copy number abnormal expression levels of more than 25%, the imprinted gene copy number of more than 6%, or the imprinting level of at least 50% is level IV;

As can be seen from fig. 5 (l), for the imprinted gene Z12, any one or a combination of at least two of the imprinted gene deletion expression level of less than 11%, the imprinted gene copy number abnormal expression level of less than 1.5%, or the imprinted gene total expression level of less than 20% is level 0, any one or a combination of at least two of the imprinted gene deletion expression level of 11 to 15%, the imprinted gene copy number abnormal expression level of 1.5 to 2.5%, or the imprinted gene total expression level of 20 to 30% is level I, the imprinted gene deletion expression level of 15 to 20%, the imprinted gene copy number abnormal expression level of 2.5 to 4%, or the imprinted gene total expression level of 30 to 40% is level II, the imprinted gene deletion expression level of 20 to 25%, the imprinted gene copy number abnormal expression level of 4 to 6%, or the imprinted gene total expression level of 40 to 50%, any one or a combination of at least two of the imprinted gene copy number abnormal expression levels of more than 25%, the imprinted gene copy number of more than 6%, or the imprinting level of at least 50% is level IV;

as can be seen from fig. 5 (m), for the imprinted gene Z13, any one or a combination of at least two of the imprinted gene deletion expression level of less than 11%, the imprinted gene copy number abnormal expression level of less than 1.5% or the imprinted gene total expression level of less than 20% is level 0, any one or a combination of at least two of the imprinted gene deletion expression level of 11 to 15%, the imprinted gene copy number abnormal expression level of 1.5 to 2.5% or the imprinted gene total expression level of 20 to 30% is level I, the imprinted gene deletion expression level of 15 to 20%, the imprinted gene copy number abnormal expression level of 2.5 to 4% or the imprinted gene total expression level of 30 to 40% is level II, the imprinted gene deletion expression level of 20 to 25%, the imprinted gene copy number abnormal expression level of 4 to 6% or the imprinted gene total expression level of 40 to 50% is level III, the gene deletion expression level of more than 25%, the imprinted gene copy number abnormal expression level of more than 6% or the combination of at least 50% is level IV;

As can be seen from fig. 5 (n), for the imprinted gene Z14, any one or a combination of at least two of the imprinted gene deletion expression level of less than 11%, the imprinted gene copy number abnormal expression level of less than 1.5%, or the imprinted gene total expression level of less than 20% is level 0, any one or a combination of at least two of the imprinted gene deletion expression level of 11 to 15%, the imprinted gene copy number abnormal expression level of 1.5 to 2.5%, or the imprinted gene total expression level of 20 to 30% is level I, the imprinted gene deletion expression level of 15 to 20%, the imprinted gene copy number abnormal expression level of 2.5 to 4%, or the imprinted gene total expression level of 30 to 40% is level II, the imprinted gene deletion expression level of 20 to 25%, the imprinted gene copy number abnormal expression level of 4 to 6%, or the imprinted gene total expression level of 40 to 50%, any one or a combination of at least two of the imprinted gene copy number abnormal expression levels of more than 25%, the imprinted gene copy number of more than 6%, or the imprinting level of at least 50% is level IV;

as can be seen from FIG. 5 (o), for the above-mentioned imprinted gene Z15, the amount of the imprinted gene deletion expression is less than 11%, the amount of the imprinted gene copy number abnormal expression is less than 1.5% or the amount of the imprinted gene total expression is less than 20%, any one or combination of at least two of the imprinted gene deletion expression is 0 level, the amount of the imprinted gene deletion expression is 11-15%, the amount of the imprinted gene copy number abnormal expression is 1.5-2.5% or the amount of the imprinted gene total expression is 20-30%, any one or combination of at least two of the imprinted gene copy number abnormal expression is 1.5-20%, the amount of the imprinted gene copy number abnormal expression is 2.5-4%, or the amount of at least two of the imprinted gene total expression is 30-40%, the amount of the imprinted gene copy number abnormal expression is 20-25%, the amount of the imprinted gene copy number abnormal expression is 4-6%, or the combination of at least two of the imprinted gene copy number abnormal expression is 40-50%, the amount of the imprinted gene deletion expression is 15-20%, the amount of the imprinted gene copy number is 50%, the amount of the imprint is 50%, and the amount of the imprinted gene copy number is greater than 25%, the amount of the imprinted gene copy number is 50%, the one or the combination of the imprinted gene copy number is greater than 50%.

From a comprehensive analysis of the samples of these 45 pancreatic cancer tumors, it can be derived that:

the judging pancreatic tumor benign and malignant degree is divided into benign tumor, pancreatic cancer potential, early pancreatic cancer, medium pancreatic cancer and advanced pancreatic cancer;

the result of the judgment of the benign and malignant degree of pancreatic tumor is that the imprinting gene deletion expression level of not more than 1 imprinting gene among imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 and the imprinting gene copy number abnormal expression level are both less than level I or that the imprinting gene copy number abnormal expression level of not more than 1 imprinting gene among imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is level I and that the imprinting gene deletion expression level of not more than 1 imprinting gene among imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is benign tumor;

the result of the judgment of the benign or malignant degree of pancreatic tumor is that the imprinting gene deletion expression level of at least 2 imprinting genes among imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is level I, the imprinting gene deletion expression level of at least 2 imprinting genes among imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is level I or the imprinting gene copy number abnormality expression level of at least 2 imprinting genes among imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is level II and the imprinting gene copy number of at least 1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z14 and Z16 is level II, the imprinting gene copy number of the imprinting gene is not more than one of pancreatic tumor;

The result of the judgment of the benign or malignant degree of pancreatic tumor is that the imprinting gene deletion expression level of at least 2 imprinting genes among imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is level II, the imprinting gene deletion expression level of not more than 1 imprinting genes among imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is level II or the imprinting gene copy number abnormality expression level of at least 2 imprinting genes among imprinting genes Z1, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is level III and the imprinting gene copy number abnormality expression level of not more than one of early stage genes among imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z14 and Z16 is level III;

the result of the judgment of the benign or malignant degree of pancreatic tumor is that the imprinting gene deletion expression level of at least 2 imprinting genes among imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is class III, the imprinting gene deletion expression level of not more than 1 imprinting genes among imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is class III or the imprinting gene copy number of at least 2 imprinting genes among imprinting genes Z1, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is class IV and the imprinting gene copy number of not more than one of the imprinting genes among imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z14, Z15 and Z16 is class IV;

The result of the determination of the benign and malignant degree of pancreatic tumor is that the imprinting gene deletion expression level of at least 2 imprinting genes among imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is IV or the imprinting gene copy number abnormal expression level of at least 2 imprinting genes among imprinting genes Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16 is IV, and the pancreatic cancer is advanced.

In summary, the detection model and the detection system of the invention show the appearance of imprinting missing on the sample of pancreatic tumor patients in an intuitive way, objectively, intuitively, early and accurately detect the change of imprinting (trace) genes by the method of marking imprinting genes in situ, and can provide a quantitative model, thereby greatly contributing to diagnosis of pancreatic tumors.

The applicant states that the detailed method of the present invention is illustrated by the above examples, but the present invention is not limited to the detailed method described above, i.e. it does not mean that the present invention must be practiced in dependence upon the detailed method described above. It should be apparent to those skilled in the art that any modification of the present invention, equivalent substitution of raw materials for the product of the present invention, addition of auxiliary components, selection of specific modes, etc., falls within the scope of the present invention and the scope of disclosure.

Claims

1. A device for detecting benign and malignant pancreatic tumor, comprising the following elements:

(1) Sampling unit: obtaining a sample to be tested;

(2) Probe design unit: designing a specific probe according to the intron sequence of the imprinted gene;

wherein the imprinting gene comprises a combination of Z1, Z11 and Z16, or a combination of Z1, Z10, Z11 and Z16, or a combination of Z1, Z8, Z9, Z10, Z11 and Z16, the imprinting gene Z1 is Gnas, the imprinting gene Z8 is Dcn, the imprinting gene Z9 is Dlk1, the imprinting gene Z10 is Gatm, the imprinting gene Z11 is Grb10, the imprinting gene Z16 is Snrpn/Snurf,

the imprinted gene further comprises any one or a combination of at least two of Z2, Z3, Z4, Z6, Z12, Z13, Z14 or Z15; wherein the imprinting gene Z2 is Igf2, the imprinting gene Z3 is Peg10, the imprinting gene Z4 is Igf2r, the imprinting gene Z6 is Plagl1, the imprinting gene Z12 is Peg3, the imprinting gene Z13 is Sgce, the imprinting gene Z14 is Slc38a4, the imprinting gene Z15 is Diras3,

The analysis unit judges the benign and malignant degree of the pancreatic tumor by calculating the imprinted gene deletion expression quantity, the imprinted gene copy number abnormal expression quantity and the total expression quantity and by a imprinted gene grading model of the pancreatic tumor, thereby judging the benign and malignant degree of the pancreatic tumor by the grades of the imprinted gene deletion expression quantity, the imprinted gene copy number abnormal expression quantity and the total expression quantity,

the imprinting gene grading model grades the expression state of the imprinting genes by calculating the total expression quantity of the imprinting genes, the deletion expression quantity of the imprinting genes and the variation of the copy number abnormal expression quantity of the imprinting genes in pancreatic tumors;

wherein the imprinted genes Z1, Z8, Z9, Z10, Z11, Z16 and any one or at least two of Z2, Z3, Z4, Z6, Z12, Z13, Z14 or Z15 are subjected to in situ hybridization detection by using a sequence selected from introns of each gene as a probe,

the formulas for calculating the total expression quantity of the imprinting genes, the deletion expression quantity of the imprinting genes and the copy number abnormal expression quantity of the imprinting genes are as follows:

total expression = (b+c+d)/(a+b+c+d) ×100%;

normal imprinted gene expression amount = b/(b+c+d) ×100%;

imprinted gene deleted gene expression amount = c/(b+c+d) ×100%;

Gene expression amount of the imprinted gene copy number abnormality=d/(b+c+d) ×100%;

wherein a is a cell nucleus in which no marker exists in the cell nucleus and the imprinted gene is not expressed after the cell is subjected to hematoxylin staining; b is a cell nucleus with a red/brown mark in the cell nucleus and a marking gene after the cell is subjected to hematoxylin staining; c is a cell nucleus with two red/brown marks in the cell nucleus and marking the gene deletion after the cell is subjected to hematoxylin staining; the d is the cell nucleus with more than two red/brown marks in the cell nucleus after the cell is subjected to hematoxylin staining and the marked gene copy number is abnormal,

the deletion expression level of the imprinting gene, the abnormal copy number expression level of the imprinting gene and the total expression level of the imprinting gene are divided into five different levels;

the five different grades are five different grades respectively divided into a imprinted gene deletion expression level, an imprinted gene copy number abnormal expression level and an imprinted gene total expression level for fifteen imprinted genes of Z1, Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z11, Z12, Z13, Z14, Z15 and Z16;

for pancreatic tumor detection, the five different grades of the deletion expression quantity of the imprinting genes, the copy number abnormal expression quantity of the imprinting genes and the total expression quantity of the imprinting genes aiming at Z1 and Z11 are divided into:

For pancreatic tumor detection, the five different classes of the imprinted gene deletion expression level, the imprinted gene copy number abnormal expression level and the total expression level for Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z12, Z13, Z14, Z15 and Z16 are divided into:

grade IV: the imprinted genes Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z12, Z13, Z14, Z15 and Z16 have a imprinting gene deletion expression level of more than 25%, the imprinted genes Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z12, Z13, Z14, Z15 and Z16 have an imprinting gene copy number abnormal expression level of more than 6% or the imprinted genes Z2, Z3, Z4, Z5, Z6, Z8, Z9, Z10, Z12, Z13, Z14, Z15 and Z16 have a total expression level of more than 50% or a combination of at least two.

2. The device of claim 1, wherein the sample to be tested in step (1) is derived from human tissue and/or cells.

3. The device of claim 2, wherein the sample to be tested is a needle biopsy sample.

4. The apparatus of claim 1, wherein said in situ hybridization employs an RNAscope in situ hybridization method.

5. The device of claim 4, wherein the RNAscope in situ hybridization method uses a single-channel or multi-channel chromogenic kit or a single-channel or multi-channel fluorogenic kit.

6. The device of claim 4, wherein the RNAscope in situ hybridization method uses a single channel red/brown color kit or a multichannel fluorescent kit.

7. The device of any one of claims 1-6, wherein the judging the benign malignancy of pancreatic tumors is classified as benign tumor, pancreatic cancer potential, early-stage pancreatic cancer, mid-stage pancreatic cancer, and late-stage pancreatic cancer;

8. Use of a device according to any one of claims 1-7 for the preparation of a medicament or kit for pancreatic tumour detection.

9. The use according to claim 8, wherein the diagnosis of benign malignancy of pancreatic tumor is classified into benign tumor, pancreatic cancer potential, early pancreatic cancer, intermediate pancreatic cancer and late pancreatic cancer.