CN106405086A - Lung cancer screening kit - Google Patents
Lung cancer screening kit Download PDFInfo
- Publication number
- CN106405086A CN106405086A CN201610837837.0A CN201610837837A CN106405086A CN 106405086 A CN106405086 A CN 106405086A CN 201610837837 A CN201610837837 A CN 201610837837A CN 106405086 A CN106405086 A CN 106405086A
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- Prior art keywords
- txk
- reagent
- lung cancer
- expression
- kit
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Classifications
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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
- G01N33/57423—Specifically defined cancers of lung
-
- 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
- G01N33/573—Immunoassay; Biospecific binding assay; Materials therefor for enzymes or isoenzymes
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
- G01N2333/90—Enzymes; Proenzymes
- G01N2333/91—Transferases (2.)
- G01N2333/912—Transferases (2.) transferring phosphorus containing groups, e.g. kinases (2.7)
Abstract
The invention discloses a lung cancer screening kit. The lung cancer screening kit comprises an optional reagent for detecting TXK (resting lymphocy te kinase) expression level. The invention further discloses an application of the reagent for detecting the TXK expression level in preparation of a lung cancer screening reagent. On the basis of the detection of the TXK expression level, the kit can judge the lung cancer risk of a to-be-detected person, can be used for clinical auxiliary diagnosis for lung cancer, provides effective reference for a patient to take related treatment measures or make decisions, and has good clinical application prospect.
Description
Technical field
The present invention relates to a kind of screening lung cancer kit.
Background technology
Lung cancer is one of modal malignant tumour in the world, and its M ＆ M is in ascendant trend year by year, at present
The incidence of disease occupies first place in the world, and seriously threatens human health and life.
The cause of disease complexity of lung cancer is it is considered that influence factor includes：1. smoking；2. environmental pollution：As haze, indoor dress
Repair；3. bad life style：As poor in eating habit, life stress is big；4. chronic lung disease：As pulmonary tuberculosis, pneumoconiosis, silicosis,
Chronic bronchitis；5. human body internal factor：As familial inheritance, immunity function reduction, endocrine function imbalance etc..
Meanwhile, lung cancer is a kind of disease be good at and hiding, and often develops into late period in disease and just shows clinical symptoms, 70
～80% patients with lung cancer when being diagnosed to be with Lung Cancer Symptoms be in, late period, cancer cell has spread, and misses and most preferably controls
More opportunity, five year survival rate is low.For the patients with lung cancer of early stage, through timely treatment be greatly improved 5 years of patient and more than
Survival rate and life quality.The therefore early diagnosis of lung cancer and to carry out effective examination most important.
The examination of lung cancer, refers to do not have the related indication crowd of lung cancer to carry out routine physical examination those, before symptom
Find lung cancer in time.If lung cancer molecular marker can be found, for pointing out clinician's early stage that patient is taken with correlation
Remedy measures or decision-making have great importance.
TXK, also known as RLK (Resting lymphocy te kinase), is non-receptor protein tyrosine kinase Tec family
One of member, main expression in T cell, participates in signal transduction in the T cell of cytokine activation, plays in immunological regulation
Important function.Have no the report in Expressions in Lung Cancer situation for the TXK at present.
Content of the invention
In order to solve the above problems, inventor studies in detail to lung cancer it was found that TXK can be used as its molecule mark
Will thing.Wherein, expression in lung tissue for the TXK is proportionate with lung cancer.Therefore, by detecting the table of TXK in lung tissue
Reach level, the risk that person to be checked suffers from lung cancer can be predicted.
Accordingly, the invention provides a kind of screening lung cancer kit, and the reagent of the expression of detection TXK is in preparation
Purposes in screening lung cancer reagent.
The screening lung cancer kit of the present invention, it includes the optional reagent for detecting TXK expression.
Wherein, described reagent is the reagent for detecting TXK expression in lung tissue.
Wherein, the reagent of described detection TXK expression is method for immunohistochemical detection reagent.
Wherein, the reagent of described detection TXK expression is Western Blot or ELISA detection method reagent.
Present invention also offers purposes in preparing screening lung cancer reagent for the reagent of detection TXK expression.
Wherein, described reagent is the reagent for detecting TXK expression in lung tissue.
Wherein, the reagent of described detection TXK expression is method for immunohistochemical detection reagent.
Wherein, the reagent of described detection TXK expression is Western Blot or ELISA detection method reagent.
Kit of the present invention passes through to detect the TXK expression with normal portions for the pulmonary abnormalities position of lung exception person, can
To judge the TXK expression difference of lung exception person, and then judge the risk that person to be checked suffers from lung cancer：If the expression of TXK is high,
The risk then suffering from lung cancer is high, if the expression of TXK is low, the risk suffering from lung cancer is low, can be used for the auxiliary diagnosis of clinical lung cancer,
Potential applicability in clinical practice is good.
Obviously, the above according to the present invention, according to ordinary technical knowledge and the customary means of this area, without departing from
Under the premise of the present invention above-mentioned basic fundamental thought, modification, replacement or the change of other various ways can also be made.
The specific embodiment of form by the following examples, remakes further specifically to the above of the present invention
Bright.But this scope being interpreted as the above-mentioned theme of the present invention should not be only limitted to Examples below.All based on the above of the present invention
The technology realized belongs to the scope of the present invention.
Specific embodiment
Embodiment 1TXK expression and the relation of lung cancer
First, experimental technique
1st, clinical data
Choose patients with lung cancer paraffin section 20, far-end compares 20 and (refers to the far-end normal lung tissue paraffin of patients with lung cancer
Section, apart from cancerous tissue 5cm), essential information is shown in Table 1.
Table 1 clinical elementary information
*Note：Other include large cell carcinoma, adenosquamous carcinoma, small cell carcinoma.
2nd, the detection of TXK expression
Paraffin section is compareed to patients with lung cancer paraffin section, remote organization, carries out the SABC with TXK as index
(IHC).
TXK antibody：Purchased from Abcam company, article No. ab37818；
EnVisionTMHRP bis- resists, antibody diluent, DAB developer, antigen retrieval buffers, Wash Buffer, immune group
Change pen：It is purchased from Dako company of Denmark.
Detect the expression of TXK as follows：
(1) close peroxidase：Paraffin section is taken to be dewaxed, dewaxing, (dewaxing process is as follows to aquation：Dimethylbenzene I
10min → dimethylbenzene II 10min → absolute ethyl alcohol 5min → 95% ethanol 5min → 85% ethanol 5min → 75% ethanol 5min)
Afterwards, use 3%H2O2Solution closes endogenous peroxydase, incubated at room 15min in dark place；
(2) distilled water rinsing：Section is placed in distilled water, shaking table is washed 5min/ time, totally three times；
(3) antigen retrieval：Add the sodium citrate solution of Tris-EDTA or pH 6.0 of pH 8.0, in 95 DEG C of water-baths
Middle water-bath 50min, carries out antigen retrieval；
(4) distilled water rinsing：Take out section, after naturally cooling to room temperature, washed three times with distillation, then use Wash
Buffer rinse is once；
(5) incubation one resists：With SABC pen, tissue is enclosed, in circle, dropping TXK antibody diluent is (for TXK antibody
With antibody diluent mixed preparing, the two ratio is 1:100), 4 DEG C of overnight incubation；
(6) distilled water rinsing：With distillation washing three times, Wash Buffer rinse is once；
(7) dropping two resists：Two anti-working solutions of dropping Dako HRP mark (are resisted to be mixed with antibody diluent for HRP bis- and join
System, the two ratio is 1:1), incubated at room 60min；
(8) distilled water rinsing：Distillation washing 3 times, each 5min；
(9) DAB colour developing：Dropping chromogenic substrate DAB, examines under a microscope colour developing situation, terminating reaction in distilled water；
(10) haematoxylin is redyed：Dyeing 2min in haematoxylin dye liquor is put in section, after 1% hydrochloride alcohol differentiation, flowing water
Rinse 10min；
(11) dehydration is transparent：Transparent through 80%, 95%, 100% gradient alcohol dehydration, dimethylbenzene, it is dried in ventilating kitchen；
(12) mounting：With neutral Instant cement mounting, observe under an optical microscope, DP Controller IMAQ system
System adopts figure.After adopting figure, the tumour cell staining power according to patients with lung cancer and remote organization and positive area give SABC
Scoring.
3rd, SABC standards of grading
Tumour cell staining power * positive tumor cell area=0-9 divides,
Result counts：Negative：0 point；Low positive：1-3 divides；High positive>3 points.
Wherein, the scoring of tumour cell staining power, positive area@As follows：
@
Note：Independently immunohistochemical staining result is scored by two veteran pathologist
4th, interpretation of result
Statistical analysis are carried out using SPSS17.0 cancerous lung tissue group and remote organization's control group.
2nd, experimental result
During cancerous lung tissue is compareed with remote organization, the expression testing result of TXK is shown in Table 2.
The expression of table 2TXK
From table 2, in far-end normal structure, the positive expression rate of TXK is 30%, and High positive expression rate is 5%；Lung
TXK positive expression rate in cancerous tissue is 90%, and High positive expression rate is 65%；TXK significantly raises in cancerous lung tissue, with
Far-end normal structure is compared, and TXK expression difference has statistical significance (P<0.001).
As can be seen from the above results, compared with Ai Pang normal lung tissue, the TXK expression of cancerous lung tissue significantly raises
(P<0.001), illustrate that lung cancer is proportionate with TXK expression, the high expression of TXK can significantly improve the possibility suffering from lung cancer.
Because the TXK level of Ai Pang normal lung tissue can reflect the TXK level of Normal Lung tissue, therefore, it can be checked by detecting
The expression of the TXK of person, by Susceptible population's examination of lung cancer out.
Embodiment 2 present invention detects kit forms and its using method of TXK expression
First, the composition of immunologic combined detection reagent kit
Detection kit (50 person-portion)：
| Component | Volume |
| TXK antibody | 30μl |
| EnVisionTMHRP bis- resists | 100μl |
| Antibody diluent | 10ml |
| DAB developer | 10ml |
| Antigen retrieval buffers | 500ml |
| Wash Buffer* | 500ml |
| SABC pen | 1 |
Note：It is based on 100 μ l that this kit drips all kinds of experiment reagents by every section, and concrete consumption can be according to tissue size
Suitably increase and decrease.
2nd, the using method of kit
By sample lung tissue to be checked, prepare paraffin section, as detection sample, detect the expression of TXK as follows
Level：
(1) close peroxidase：Paraffin section is taken to be dewaxed, dewaxing, (dewaxing process is as follows to aquation：Dimethylbenzene I
10min → dimethylbenzene II 10min → absolute ethyl alcohol 5min → 95% ethanol 5min → 85% ethanol 5min → 75% ethanol 5min)
Afterwards, use 3%H2O2Solution closes endogenous peroxydase, incubated at room 15min in dark place；
(2) distilled water rinsing：Section is placed in distilled water, shaking table is washed 5min/ time, totally three times；
(3) antigen retrieval：Add the sodium citrate solution of Tris-EDTA or pH 6.0 of pH 8.0, in 95 DEG C of water-baths
Middle water-bath 50min, carries out antigen retrieval；
(4) distilled water rinsing：Take out section, after naturally cooling to room temperature, washed three times with distillation, then use Wash
Buffer rinse is once；
(5) incubation one resists：With SABC pen, tissue is enclosed, in circle, dropping TXK antibody diluent is (for TXK antibody
With antibody diluent mixed preparing, the two ratio is 1:100), 4 DEG C of overnight incubation；
(6) distilled water rinsing：With distillation washing three times, Wash Buffer rinse is once；
(7) dropping two resists：Two anti-working solutions of dropping Dako HRP mark (are resisted to be mixed with antibody diluent for HRP bis- and join
System, the two ratio is 1:1), incubated at room 60min；
(8) distilled water rinsing：Distillation washing 3 times, each 5min；
(9) DAB colour developing：Dropping chromogenic substrate DAB, examines under a microscope colour developing situation, terminating reaction in distilled water；
(10) haematoxylin is redyed：Dyeing 2min in haematoxylin dye liquor is put in section, after 1% hydrochloride alcohol differentiation, flowing water
Rinse 10min；
(11) dehydration is transparent：Transparent through 80%, 95%, 100% gradient alcohol dehydration, dimethylbenzene, it is dried in ventilating kitchen；
(12) mounting：With neutral Instant cement mounting, observe under an optical microscope, DP Controller IMAQ system
System adopts figure.After adopting figure, the tumour cell staining power according to patients with lung cancer and remote organization and positive area give SABC
Scoring.
To pulmonary abnormalities person, abnormal position, normal portions tissue can be taken respectively, compare TXK expression, and then evaluate it
Suffer from the possibility of lung cancer, as the auxiliary diagnosis means of clinical lung cancer.
To sum up, kit of the present invention passes through to detect the expression of TXK it can be determined that the pulmonary abnormalities position of lung exception person
With the TXK expression difference of normal portions, and then examination person to be checked suffers from the risk of lung cancer：If the expression of TXK is high, suffer from
The risk of lung cancer is high, if the expression of TXK is low, the risk suffering from lung cancer is low, can be used for the auxiliary diagnosis of clinical lung cancer, for suffering from
Person takes the remedy measures of correlation or decision-making to provide effective foundation, and potential applicability in clinical practice is good.
Claims (8)
1. a kind of screening lung cancer kit it is characterised in that：It includes the optional reagent for detecting TXK expression.
2. kit for screening according to claim 1 it is characterised in that：Described reagent is for detecting TXK in lung tissue
The reagent of expression.
3. kit for screening according to claim 1 and 2 it is characterised in that：The reagent of described detection TXK expression is
Method for immunohistochemical detection reagent.
4. kit for screening according to claim 1 and 2 it is characterised in that：The reagent of described detection TXK expression is
Western Blot or ELISA detection method reagent.
5. purposes in preparing screening lung cancer reagent for the reagent of detection TXK expression.
6. purposes according to claim 5 it is characterised in that：Described reagent is for detecting TXK expression water in lung tissue
Flat reagent.
7. the purposes according to claim 5 or 6 it is characterised in that：The reagent of described detection TXK expression is immune group
Change detection method reagent.
8. the purposes according to claim 5 or 6 it is characterised in that：The reagent of described detection TXK expression is
Western Blot or ELISA detection method reagent.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610837837.0A CN106405086A (en) | 2016-09-21 | 2016-09-21 | Lung cancer screening kit |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610837837.0A CN106405086A (en) | 2016-09-21 | 2016-09-21 | Lung cancer screening kit |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN106405086A true CN106405086A (en) | 2017-02-15 |
Family
ID=57996917
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| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201610837837.0A Pending CN106405086A (en) | 2016-09-21 | 2016-09-21 | Lung cancer screening kit |
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Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2007048978A2 (en) * | 2005-10-28 | 2007-05-03 | Biomerieux Sa | Method for detecting cancer |
| CN102656458A (en) * | 2009-10-26 | 2012-09-05 | 雅培制药有限公司 | Diagnostic methods for determining prognosis of non-small cell lung cancer |
| WO2014071231A1 (en) * | 2012-11-02 | 2014-05-08 | Pharmacyclics, Inc. | Tec family kinase inhibitor adjuvant therapy |
| CN104755474A (en) * | 2012-10-11 | 2015-07-01 | 药品循环公司 | Companion diagnostics for TEC family kinase inhibitor therapy |
| WO2016010961A1 (en) * | 2014-07-15 | 2016-01-21 | Abbvie Inc. | Enzyme occupancy assay |
-
2016
- 2016-09-21 CN CN201610837837.0A patent/CN106405086A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2007048978A2 (en) * | 2005-10-28 | 2007-05-03 | Biomerieux Sa | Method for detecting cancer |
| CN102656458A (en) * | 2009-10-26 | 2012-09-05 | 雅培制药有限公司 | Diagnostic methods for determining prognosis of non-small cell lung cancer |
| CN104755474A (en) * | 2012-10-11 | 2015-07-01 | 药品循环公司 | Companion diagnostics for TEC family kinase inhibitor therapy |
| WO2014071231A1 (en) * | 2012-11-02 | 2014-05-08 | Pharmacyclics, Inc. | Tec family kinase inhibitor adjuvant therapy |
| WO2016010961A1 (en) * | 2014-07-15 | 2016-01-21 | Abbvie Inc. | Enzyme occupancy assay |
Non-Patent Citations (2)
| Title |
|---|
| HSIN-YI WU ET AL.: "Identification of Tyrosine-Phosphorylated Proteins Associated with Lung Cancer Metastasis using Label-Free Quantitative Analyses", 《JOURNAL OF PROTEOME RESEARCH》 * |
| KAZUYA MACHIDA ET AL.: "Characterizing Tyrosine Phosphorylation Signaling in Lung Cancer Using SH2 Profiling", 《PLOS ONE》 * |
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Application publication date: 20170215 |