CN107727859B - A kind of serum I FN- γ and LncRNAs composition is used to prepare the purposes of cancer diagnosis reagent box - Google Patents
A kind of serum I FN- γ and LncRNAs composition is used to prepare the purposes of cancer diagnosis reagent box Download PDFInfo
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- CN107727859B CN107727859B CN201710926251.6A CN201710926251A CN107727859B CN 107727859 B CN107727859 B CN 107727859B CN 201710926251 A CN201710926251 A CN 201710926251A CN 107727859 B CN107727859 B CN 107727859B
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/574—Immunoassay; Biospecific binding assay; Materials therefor for cancer
- G01N33/57407—Specifically defined cancers
- G01N33/57446—Specifically defined cancers of stomach or intestine
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/574—Immunoassay; Biospecific binding assay; Materials therefor for cancer
- G01N33/57484—Immunoassay; Biospecific binding assay; Materials therefor for cancer involving compounds serving as markers for tumor, cancer, neoplasia, e.g. cellular determinants, receptors, heat shock/stress proteins, A-protein, oligosaccharides, metabolites
Abstract
The invention discloses the purposes that a kind of serum I FN- γ and LncRNAs composition is used to prepare cancer diagnosis reagent box.When serum I FN- γ diagnosis differentiation gastric cancer and healthy volunteer is used alone, area (AUC) is 0.636 under ROC curve.It is a discovery of the invention that AUC can be improved to 0.793 or 0.805 when distinguishing gastric cancer and healthy volunteer using serum NO level NHSAT089447 or NONHSAT041499 and serum I FN- γ Combining diagnosis；When distinguishing gastric cancer and healthy volunteer using serum NO level NHSAT089447 and NONHSAT041499 and serum I FN- γ Combining diagnosis, AUC be can be improved to 0.921.One skilled in the art will appreciate that area is between 1.0 and 0.5 under ROC curve, in the case where AUC > 0.5, AUC illustrates that diagnosis effect is better closer to 1.AUC has lower accuracy in 0.5-0.7, and AUC has certain accuracy in 0.7-0.9, and AUC has high accuracy at 0.9 or more.Therefore, serum I FN- γ can combine the diagnostic kit that serum NO level NHSAT089447 and/or NONHSAT041499 are used to prepare diagnosis of gastric cancer.
Description
Technical field
The invention belongs to gene diagnosis fields, are related to the exploitation of diagnostic kit, and in particular to a kind of serum I FN- γ and
LncRNAs composition is used to prepare the purposes of cancer diagnosis reagent box.
Background technique
Gastric cancer occupies malignant tumour the 2nd in China's disease incidence, and death toll accounts for cancer occurrence numbers 14.33%, occupies the 3rd, only
Inferior to lung cancer and liver cancer.It can be seen that gastric cancer is the emphasis for the treatment of and prevention of tumour.As tumor markers are widely used in clinic, more
Come more scholars the study found that tumor markers play an important role during prevention and control of cancer.IFN-γ is to participate in being immunized
The important cytokine of adjusting is generally secreted by the T lymphocyte activated, can influence the growth of tumour cell.IFN-γ exists
Tumor patient Level of Expression of Retinoic Acid is obviously abnormal.Based on this, has multinomial research using IFN-γ as the marker of diagnosing tumor.
Serum I FN- γ is used for diagnosis of gastric cancer (serum I FN- γ and tumor markers by existing research
Joint-detection is to the diagnostic value of gastric cancer, biomedical engineering and clinical the 2nd phase of volume 20 in March, 2016), still, individually make
Accuracy with serum I FN- γ diagnosis of gastric cancer is lower, and clinical value is low.
Summary of the invention
The purpose of the present invention is to provide serum I FN- γ and LncRNAs compositions to be used to prepare cancer diagnosis reagent box
Purposes, to improve the accuracy of clinical diagnosis gastric cancer.
The present invention is achieved by following technical solution:
IFN-γ and LncRNANONHSAT089447 or LncRNANONHSAT041499, which combine, is used to prepare diagnosis of gastric cancer
Diagnosis composition purposes.
IFN-γ and LncRNANONHSAT089447, LncRNANONHSAT041499, which combine, is used to prepare diagnosis of gastric cancer
The purposes of diagnosis composition.
Preferably, the IFN-γ, LncRNANONHSAT089447, LncRNANONHSAT041499 are serum I FN-
γ、NONHSAT089447、NONHSAT041499。
IFN-γ and LncRNANONHSAT089447 or LncRNANONHSAT041499, which combine, is used to prepare diagnosis of gastric cancer
Diagnostic kit purposes.
IFN-γ and LncRNANONHSAT089447, LncRNANONHSAT041499, which combine, is used to prepare diagnosis of gastric cancer
The purposes of diagnostic kit.
Preferably, the IFN-γ, LncRNANONHSAT089447, LncRNANONHSAT041499 are serum I FN-
γ、NONHSAT089447、NONHSAT041499。
Advantages of the present invention:
When serum I FN- γ diagnosis differentiation gastric cancer and healthy volunteer is used alone, area (AUC) is under ROC curve
0.636.It is a discovery of the invention that being distinguished using serum NO level NHSAT089447 or NONHSAT041499 and serum I FN- γ Combining diagnosis
When gastric cancer is with healthy volunteer, AUC be can be improved to 0.793 or 0.805；Using serum NO level NHSAT089447 and
When NONHSAT041499 and serum I FN- γ Combining diagnosis distinguish gastric cancer and healthy volunteer, AUC be can be improved to 0.921.This
Field technical staff knows that area is between 1.0 and 0.5 under ROC curve, in the case where AUC > 0.5, AUC closer to 1,
Illustrate that diagnosis effect is better.AUC has lower accuracy in 0.5-0.7, and AUC has certain accuracy in 0.7-0.9, and AUC exists
There is high accuracy when 0.9 or more.Therefore, serum I FN- γ can combine serum NO level NHSAT089447 and/or
NONHSAT041499 is used to prepare the diagnostic kit of diagnosis of gastric cancer.
Detailed description of the invention
Fig. 1 is that serum I FN- γ individually diagnoses the ROC curve for distinguishing patients with gastric cancer and healthy volunteer；
Fig. 2 is that serum I FN- γ combines NONHSAT089447 diagnosis differentiation patients with gastric cancer and the ROC of healthy volunteer is bent
Line；
Fig. 3 is that serum I FN- γ combines NONHSAT041499 diagnosis differentiation patients with gastric cancer and the ROC of healthy volunteer is bent
Line；
Fig. 4 is that serum I FN- γ combines serum NO level NHSAT089447 and serum NO level NHSAT041499 diagnosis differentiation gastric cancer trouble
The ROC curve of person and healthy volunteer.
Specific embodiment
It is further described technical solution of the present invention combined with specific embodiments below.
One, experimental material
Serum Obtained From Advance Gastric Cancer sample is derived from patient after the attached middle large hospital Surgery for Gastric Carcinoma of Southeast China University.All patients are preoperative
Do not receive the treatment such as radiotherapy, chemotherapy and anti-tumor drug, there is complete clinical and pathological data.Patients with gastric cancer
138 (gastric cancer group), wherein women 63, male 75；Age 35-75 years old, average age 63.3 years old.
Healthy volunteer's serum specimen 55 (Normal group), wherein women 26, male 29；Age 35-70 years old,
Average age 61.5 years old.
Age and sex composition no significant difference between gastric cancer group and Normal group.
IFN-γ detection Enzyme Linked Immunoadsorbent Assay (ELISA) kit is purchased from Raybiotech company.
Two, experimental method
1, serum I FN- γ measuring method
IFN-γ detection uses ELISA method, illustrates strict implement according to kit.
2, serum LncRNAs measuring method
200 μ L of serum is taken, extracts total serum IgE using the RNAprepPure kit of Tiangen company.It is answered in experimentation
RNA degradation is reduced, as far as possible to obtain the RNA that purity and quality are higher, pollute without albumen and other impurity.With 1.5% fine jade
The integrality of sepharose electrophoretic analysis RNA, ultraviolet spectrophotometry detect RNA concentration.
Use TIANScript cDNA the first chain synthetic agent box and the general reverse of oligo (dT) 15 of Tiangen company
It records primer and carries out reverse transcription reaction.Reverse transcription total system is 20 μ L:1-5 μ g total serum IgEs, 2 μ L oligo (dT), 15,2 μ L Super
Pure dNTPs mends RNase-Free ddH2O to 14.5 μ L.Reaction tube cools down rapidly on ice after setting 70 DEG C of heating 5min
4 μ L 5X FIRST STRAND BUFFER (containing DTT), 0.5 μ L RNasin, 1 μ L TIANScriptM- are added in 2min
MLV is gently mixed with pipettor, 42 DEG C of warm bath 50min.95 DEG C of heating 5min terminate reaction, and reaction product is standby in -20 DEG C of preservations
With.
According to Genbank objective gene sequence, designed with primer-design software primerpremier 5.0
NONHSAT089447, NONHSAT041499 specific primer, are synthesized by Shanghai Sangon Biotech Company.Internal control primer HS-ACTB (β-
Actin) it is purchased from Shanghai Sangon Biotech Company.Reaction system (20 μ L): 10 2 μ L of μ L, cDNA of SYBR Select Master Mix, on
Swim 1 μ L of primer, downstream primer 1 μ L, ddH2O 6μL.It is detected using the 7500FAST fluorescent quantitative detector of ABI company.
Response parameter: 50 DEG C of 2min；95 DEG C of 2min, 95 DEG C of 15s, 60 DEG C of 15s, 72 DEG C of 1min, totally 45 recycle.Solubility curve parameter:
95 DEG C of 15s, 60 DEG C of 1min, 95 DEG C of 15s, 60 DEG C of 15s.Every part sample replication 3 times, obtain Ct value.With the copy number of ACTB
As correction radix, the opposite internal reference expression quantity of each LncRNAs in each sample is calculated, target LncRNAs in the sample
Relative expression quantity.
NONHSAT089447, NONHSAT041499 specific primer are as follows:
The upstream NONHSAT089447: 5 '-GTAGGGCTTGGATGGACTTT-3 '；
The downstream NONHSAT089447: 5 '-CCGTCAACACATCAGGCCT-3 '；
The upstream NONHSAT041499: 5 '-CGATTTTGGGAACCTCTATC-3 '；
The downstream NONHSAT041499: 5 '-CAACTGCTGGGTGGAGTCTA-3 '.
3, statistical procedures
It is analyzed using 20.0 software of SPSS, is indicated with mean value ± deviation, comparison among groups are examined using t.Evaluation is every
Index application value draws Receiver Operating Characteristics (ROC) curve, and area (AUC) under calculated curve；It is poor with P < 0.05
It is different statistically significant.Each single index and Joint Index are evaluated to the diagnostic value of gastric cancer using ROC curve.
Three, experimental result
1, between two groups Tumor Marker Levels comparison
In gastric cancer group serum IFN-γ level be significantly lower than Normal group, gastric cancer group serum NO NHSAT089447,
The relative expression levels of NONHSAT041499 are obviously higher than Normal group, and difference is statistically significant, and (t value is respectively
1.413,2.034 and 1.935, P < 0.01).Comparison result is as shown in table 1.
The comparison (mean value ± deviation) of Tumor Marker Levels between 1 two groups of table
2, tumor-marker analyte detection gastric cancer ROC curve is analyzed
When serum I FN- γ diagnosis differentiation gastric cancer and healthy volunteer is used alone, area (AUC) is under ROC curve
0.636.Gastric cancer and health are distinguished using serum NO level NHSAT089447 or NONHSAT041499 and serum I FN- γ Combining diagnosis
When volunteer, AUC be can be improved to 0.793 or 0.805；Use serum NO level NHSAT089447 and NONHSAT041499 and serum
When IFN-γ triple combination diagnoses differentiation gastric cancer and healthy volunteer, AUC be can be improved to 0.921, and accuracy rate is up to
92.23%.It can be seen that serum I FN- γ joint NONHSAT089447 or NONHSAT041499 diagnosis of gastric cancer has centainly
Clinical value, serum I FN- γ combine NONHSAT089447 and NONHSAT041499 diagnosis of gastric cancer clinical valence with higher
Value.The ROC curve analysis of two or three indexs combines dualistic logistic regression to operate.
ROC curve analyzes result as shown in table 2 and Fig. 1-4.
2 serum I FN- γ of table is individually and joint NONHSAT089447 and/or NONHSAT041499 diagnostic value compares
One skilled in the art will appreciate that area is between 1.0 and 0.5 under ROC curve, and in the case where AUC > 0.5, AUC
Closer to 1, illustrate that diagnosis effect is better.AUC has lower accuracy in 0.5-0.7, and AUC is fixed in 0.7-0.9 Shi Youyi
True property, AUC have high accuracy at 0.9 or more.Therefore, serum I FN- γ can combine serum NO level NHSAT089447 and/or
NONHSAT041499 is used to prepare the diagnostic kit of diagnosis of gastric cancer.
Claims (6)
1.IFN- γ and LncRNANONHSAT089447 or IFN-γ and LncRNANONHSAT041499, which combine to be used to prepare, to be examined
The purposes of the diagnosis composition of disconnected gastric cancer.
2.IFN- γ and LncRNANONHSAT089447, LncRNANONHSAT041499, which combine, is used to prepare examining for diagnosis of gastric cancer
The purposes of disconnected composition.
3. purposes according to claim 1 or 2, it is characterised in that: the IFN-γ, LncRNANONHSAT089447,
LncRNANONHSAT041499 is serum I FN- γ, NONHSAT089447, NONHSAT041499.
4.IFN- γ and LncRNANONHSAT089447 or IFN-γ and LncRNANONHSAT041499, which combine to be used to prepare, to be examined
The purposes of the diagnostic kit of disconnected gastric cancer.
5.IFN- γ and LncRNANONHSAT089447, LncRNANONHSAT041499, which combine, is used to prepare examining for diagnosis of gastric cancer
The purposes of disconnected kit.
6. purposes according to claim 4 or 5, it is characterised in that: the IFN-γ, LncRNANONHSAT089447,
LncRNANONHSAT041499 is serum I FN- γ, NONHSAT089447, NONHSAT041499.
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| CN105586399A (en) * | 2015-09-07 | 2016-05-18 | 张国新 | Kit for serum/plasma lncRNA marker related to stomach cancer |
| CN106701900B (en) * | 2015-11-16 | 2020-03-20 | 上海市东方医院 | Long-chain non-coding RNA HERC2P3 gene and application thereof in gastric cancer |
| CN106148529B (en) * | 2016-07-13 | 2019-12-06 | 中国人民解放军总医院 | LncRNA marker related to gastric cancer, special detection primer, detection method, kit and application thereof |
| CN106755513B (en) * | 2017-02-08 | 2018-10-19 | 泰山医学院 | A kind of lncRNA markers of gastric cancer |
| CN106868204A (en) * | 2017-05-02 | 2017-06-20 | 北京泱深生物信息技术有限公司 | A kind of biomarker for sdenocarcinoma of stomach diagnosis |
| CN106906305A (en) * | 2017-05-02 | 2017-06-30 | 谷建斌 | Applications of the lncRNA in sdenocarcinoma of stomach diagnosis |
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