CN103823066B - For lung cancer early metaphase quick diagnosis reagent kit and its detection method - Google Patents
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- 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
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2800/00—Detection or diagnosis of diseases
- G01N2800/12—Pulmonary diseases
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Abstract
The invention discloses one kind to be used for lung cancer early metaphase quick diagnosis reagent kit, and the capture antibody includes galactose agglutinin, cytokeratin fragment antigen 21 1 and monoclonal antibody made from human aspartyl's B-hydroxylase difference；The kit also includes detection antibody, and the detection antibody includes the polyclonal antibody after galactose agglutinin, cytokeratin fragment antigen 21 1 and HRP marks made from human aspartyl's B-hydroxylase difference.Beneficial effects of the present invention are as follows：With high sensitivity accuracy it is strong the characteristics of, its sensitivity and accuracy reach more than 90%, can effectively reduce in addition testing cost, reduce operating procedure and reduce detection error.
Description
Technical field：
The present invention relates to the kit field of cancer diagnosis detection, is specially that one kind is used for lung cancer early metaphase quick diagnosis
Kit and its detection method.
Background technology：
Lung cancer is death rate highest cancer in global range.This treatment of cancer is difficult, and the lung cancer of transfer is especially refractory.
About 75% patients with lung cancer has been metastatic or late period when making a definite diagnosis, and its 5 years survival rates are only 6%.Come over the past several decades,
Although the remedy measures such as chemotherapy, radiotherapy and operation have a quick development, 5 years survival rates of lung cancer are still relatively low.Mainly
Because can also be screened without a kind of safe method to it, early stage disease --- the best opportunity for the treatment of, just and
The possibility of Shi Faxian patients' lung cancer is very little.
There is an urgent need to a kind of effective screening strategy at present.When CT scan is unclear, blood testing can help doctor to make
Preferably judge.Asymptomatic patient is screened using CT scan, it is both expensive and dangerous.This screening has very high vacation sun
Property, some of patients will receive unnecessary lung's biopsy.
Tumor markers (tumor markers, TM) refer to tumour occur and breeding in, by tumour cell in itself
Synthesis, release, or tumour cell is reacted and a kind of material of the presence of caused signal tumor and growth by body.These materials
The level either occurred in cancer patient is not present in normal adult and is significantly higher than normal person.Tumor-marker quality testing at present
Survey technology is considered as the unique channel of the asymptomatic micro- stove tumour of early detection, this detection technique can prior to X-ray, ultrasound,
The PEs such as CT, MRI or PET-CT find tumour.Available for the examination of people at highest risk's malignant tumour, diagnosing tumor is with differentiating
Diagnosis, assess the effect for the treatment of, prediction or monitoring tumor recurrence or transfer.At present, the pulmonary cancer diagnosis kit that hospital occurs is all
It is to detect some common tumor markers, sensitivity and accuracy are all relatively low.Because of mainly selected tumor marker individual event
Detection often has significant limitation, it is difficult to meets the requirement of Rapid＆Early diagnosis.
At present, in the market also comes out without the rapidly and efficiently diagnostic kit for lung cancer, seriously affects lung cancer early stage
It was found that and treatment.
The content of the invention：
The purpose of the present invention is to be directed to insufficient existing for above-mentioned existing pulmonary cancer diagnosis, there is provided a kind of high sensitivity, accurate
Property is strong and easy to use is efficiently used for lung cancer early metaphase quick diagnosis reagent kit.
Another object of the present invention is to provide a kind of detection method for lung cancer early metaphase quick diagnosis reagent kit.
In order to realize the object of the invention, the present invention adopts the technical scheme that：For lung cancer early metaphase fast diagnosis reagent
Box, including capture antibody, the capture antibody include galactose agglutinin and cytokeratin fragment antigen 21-1（CYFRA21-
1）Monoclonal antibody made from respectively.
The capture antibody also includes human aspartyl's B-hydroxylase（HAAH）Obtained monoclonal antibody.
The kit also includes detection antibody, and the detection antibody includes galactose agglutinin, cytokeratin fragment
Antigen 21-1 (CYFRA21-1) and human aspartyl's B-hydroxylase（HAAH）It is polyclonal after HRP marks made from respectively
Antibody.
The capture antibody is by galactose agglutinin, cytokeratin fragment antigen 21-1 (CYFRA21-1) and the mankind
Aspartyl B-hydroxylase（HAAH）Carrier for expression of eukaryon is cloned into, and the expression of albumen is realized in mammalian cell,
Corresponding antigen is obtained after purification, the corresponding monoclonal antibody that the antigen immune mammal is obtained.
The step of preparation method of the capture antibody includes is as follows：
（1）The preparation of antigen：Galactose agglutinin, cytokeratin fragment antigen 21-1 and human aspartyl
The gene cloning of B-hydroxylase realizes in mammalian cell the expression of albumen to carrier for expression of eukaryon, obtains after purification
Antigen, this antigen can also be used as standard items to use；
（2）Capture the preparation of antibody：By above-mentioned antigen immune mammal, it is anti-for capture to obtain corresponding monoclonal antibody
Body.
The detection antibody is by galactose agglutinin, cytokeratin fragment antigen 21-1 (CYFRA21-1) and the mankind
Aspartyl B-hydroxylase（HAAH）Carrier for expression of eukaryon is cloned into, and the expression of albumen is realized in mammalian cell,
Corresponding antigen is obtained after purification, the corresponding polyclonal antibody that the antigen immune mammal is obtained, to described polyclonal
Antibody carries out HRP marks.
The step of preparation method of the detection antibody includes is as follows：
（1）The preparation of antigen：Galactose agglutinin, cytokeratin fragment antigen 21-1 and human aspartyl
The gene cloning of B-hydroxylase realizes in mammalian cell the expression of albumen to carrier for expression of eukaryon, obtains after purification
Antigen, this antigen can also be used as standard items to use；
（2）Detect the preparation of antibody：By above-mentioned antigen immune mammal, it is anti-for detection to obtain corresponding polyclonal antibody
Body；
（3）HRP is marked：HRP marks are carried out to the polyclonal antibody.
The galactose agglutinin preferably galactose galectin-3 and Galectin-8.
The capture antibody is coated in the hole of microtiter plate in advance, can be simplified step, be improved detection efficiency.
For the preparation method of lung cancer early metaphase quick diagnosis reagent kit, it comprises the following steps：
（1）The preparation of antigen：By galactose agglutinin, cytokeratin fragment antigen 21-1 and human aspartyl
The gene cloning of B-hydroxylase realizes in mammalian cell the expression of albumen to carrier for expression of eukaryon, obtains after purification
Required antigen, wherein, the antigen can also be used as standard items to use.
（2）Capture the preparation of antibody：Above-mentioned antigen immune mammal is obtained into corresponding monoclonal antibody, the Dan Ke
Capture antibody of the grand antibody as this kit.
（3）Detect the preparation of antibody：Above-mentioned antigen immune mammal is obtained into corresponding polyclonal antibody, to described more
Clonal antibody carries out HRP marks.Present invention detection antibody has mark function, can save labelled antibody and addition mark is anti-
The operating procedure of body, it is effective cost-effective, operating procedure is reduced, reduces detection error.
（4）Capture antibody coating：
1. the capture antibody that concentration is 1 μ g/ml is coated with microtitration by carbonate/bicarbonates buffer solution (pH9.6)
The hole of plate；2. covers microtiter plate and is incubated overnight at 4 DEG C；3. discards coating buffer（Carbonate/bicarbonate buffer solution
The capture antibody of dilution）, and wash microtiter plate twice with cleaning solution, 200 μ l PBST are added in micropore every time（Phosphoric acid
Salt Tween buffer）, the gently whipping microtiter plate above tank, cleaning solution is removed, microtiter plate is patted on paper handkerchief,
Remove remaining drop, dry be put in it is standby in 4 DEG C of environment.
（5）Closing：200 μ l Block buffers are added per hole（1.2%BSA/PBS）, close remaining albumen in coating hole
Matter binding site.
For the detection method of lung cancer early metaphase quick diagnosis reagent kit, it comprises the following steps：
（1）The preparation of antigen：By Galectin-3, HAAH and CYFRA21-1 gene clonings to carrier for expression of eukaryon, and
The expression of albumen is realized in mammalian cell, obtains required antigen after purification；
（2）Capture the preparation of antibody：Above-mentioned antigen immune mammal is obtained into corresponding monoclonal antibody, the Dan Ke
Capture antibody of the grand antibody as this kit；
（3）Detect the preparation of antibody：Above-mentioned antigen immune mammal is obtained into corresponding polyclonal antibody, to described more
Clonal antibody carries out HRP marks；
（4）Capture antibody coating：
1. the capture antibody that concentration is 1 μ g/ml is coated with microtitration by carbonate/bicarbonates buffer solution (pH9.6)
The hole of plate；
2. covers microtiter plate and is incubated overnight at 4 DEG C；
3. discards coating buffer（The capture antibody of carbonate/bicarbonate buffer solution dilution）, and washed with cleaning solution micro
Titer plate adds 200 μ l PBST in micropore twice, every time（Phosphate Tween buffer）, gently whipping is micro- above tank
Measure titer plate, remove cleaning solution, microtiter plate is patted on paper handkerchief, removes remaining drop, dry be put in it is standby in 4 DEG C of environment
With.
（5）Closing
1. adds 200 μ l Block buffers in each hole of microtiter plate（1.2%BSA/PBS）, close in coating hole
Remaining protein binding site；
2. covers microtiter plate and is incubated 1 hour at 37 DEG C；
（6）Sample-adding
1. 100 μ l sample is added to each hole of microtiter plate by, it is incubated 60 minutes at 37 DEG C;Obtain standard
True quantitative result, it is common practice that compare the signal of unknown sample and standard curve.Each ELISA Plate must bioassay standard
Product（Double measure or triplicate）And blank sample, to ensure accuracy；
2. discards sample, and washs microtiter plate three times, 200 μ l PBST are added in micropore every time（Phosphate is told
Warm buffer solution）；
3. the detection antibody that 100 μ l concentration are 0.5 μ g/ml is added to each hole by；
4. covers microtiter plate and is incubated 1 hour at 37 DEG C；
5. washs microtiter plate four times with PBST；
(7) detect
1. is by TMB（3,3', 5,5'- tetramethyl benzidine）Solution is added to each hole, is incubated 15-30 minutes, addition etc.
The terminate liquid of volume, optical density is then read at 450nm.
2. the data that is obtained by serial dilutions draw standard curve, concentration is marked on X-axis（Logarithmic scale）On, and extinction
Scale is in Y-axis（Lineal scale）On.Sample concentration is drawn on this standard curve by interpolation method.
The present invention, by a variety of permutation and combination, filters out 3 tumor markers from numerous tumor markers
（CBP-35, human aspartyl's B-hydroxylase (HAAH), cytokeratin fragment antigen 21-1 (CYFRA21-
1) lung cancer quick diagnosis reagent kit is formed.Wherein, human aspartyl's B-hydroxylase (Human Aspartyl/
Asparaginyl β-hydroxylase, HAAH) it is to be present in a kind of intracellular enzyme embryonic period, embryonic phase, belong to and rely on α -one penta 2
Sour dioxygenase family, the aspartic acid or asparagus fern in specific protein Concentrations of Epidermal Growth Factor (EGF) acceptor spline structure domain can be catalyzed
Beta carbon hydroxylating in amide residues.Research shows height expression of the HAAH in tumor cell surface, is a kind of new pernicious
The molecular marked compound of tumour high specific.Nearest research also confirms that HAAH is the ideal mark thing of lung cancer.Galactose agglutinin-
3 (Galectin-3) are a kind of galactose-binding proteins, are the members uniquely in Glectin families with embedded structure.Gala
Sugared galectin-3 (Galectin-3) is distributed widely in tumor tissues, the expression of hL-31 with its tumour invasion and attack and turn
Move closely related.Galectin-3 can be combined with the CD98 of cell surface glycosylation promotes integrin to be gathered into cell surface
Cluster, strengthen the affinity of integrin and its part indirectly;Also can be directly in conjunction with Integrin alphα1 β 1 and CD11b/18, positivity or negativity
The activity of integrin is adjusted, influences the combination of integrin and extracellular ligand.Galectin-3 can also increase integrin thin
The expression of cellular surface, and promote secretion of the collagen in cell compartment.Because CBP-35 has affinity to polysaccharide, it
Can also be directly in conjunction with glycosylated cells epimatrix composition, mediated cell and matrix are sticked.Research shows patients with lung cancer blood
Galectin-3 levels are significantly more than healthy population in clear.Cytokeratin fragment antigen 21-1 (CYFRA21-1) is cell angle
Protein 19（CYK-19 soluble fragments), CYK-19 are widely distributed in normal structure surface, in stratiform or scaly epithelium.
In malignant epithelial cell, the protease of activation accelerates the degraded of cell so that a large amount of cytokeratin fragment CYFRA21-1 are released
Blood is put into, in pernicious cancerous lung tissue, CYFRA21-1 rich contents, especially there is high expression in lung squamous cancer.
Compared with prior art, beneficial effects of the present invention are as follows：With high sensitivity accuracy it is strong the characteristics of, its is sensitive
Degree and accuracy reach more than 90%, can effectively reduce testing cost in addition, reduce operating procedure and reduce detection error.
Brief description of the drawings
Fig. 1 is the schematic diagram that the present invention is used for lung cancer early metaphase quick diagnosis reagent kit；
Fig. 2 is the selection pair for the detection antibody best effort concentration that the present invention is used for lung cancer early metaphase quick diagnosis reagent kit
Than figure；
Fig. 3 is that the present invention detects for lung cancer early metaphase quick diagnosis reagent kit in blood before and after lung cancer therapy
Galectin-3 change comparison diagram；
Fig. 4 is that the present invention detects for lung cancer early metaphase quick diagnosis reagent kit in blood before and after lung cancer therapy
CYFRA21-1 change comparison diagram；
Fig. 5 is that the present invention detects HAAH for lung cancer early metaphase quick diagnosis reagent kit in blood before and after lung cancer therapy
Change comparison diagram.
Embodiment
It is used to lung cancer early metaphase quick diagnosis reagent kit to the present invention below in conjunction with the drawings and specific embodiments carry out in detail
Description explanation.
For lung cancer early metaphase quick diagnosis reagent kit, including antibody is captured, capture antibody Block buffer, standard items,
Mark HRP detection antibody, cleaning solution and nitrite ion etc..The capture antibody includes galactose agglutinin and cytokeratin piece
Monoclonal antibody made from Duan Kangyuan 21-1 (CYFRA21-1) difference；By to galactose agglutinin and cytokeratin fragment
Antigen 21-1 (CYFRA21-1) common quick detection, can effectively improve Detection accuracy and sensitivity, effectively overcome
Only according to deficiency caused by detection CYFRA21-1.
The capture antibody also includes human aspartyl's B-hydroxylase（HAAH）Obtained monoclonal antibody, passes through
Detect galactose agglutinin, cytokeratin fragment antigen 21-1 (CYFRA21-1) and human aspartyl's B-hydroxylase
（HAAH）, its sensitivity and accuracy is reached more than 90%.
The capture antibody is by galactose agglutinin, cytokeratin fragment antigen 21-1 (CYFRA21-1) and the mankind
Aspartyl B-hydroxylase（HAAH）Carrier for expression of eukaryon is cloned into, and the expression of albumen is realized in mammalian cell,
Corresponding antigen is obtained after purification, the corresponding monoclonal antibody that the antigen immune mammal is obtained.
The step of preparation method of the capture antibody includes is as follows：
（1）The preparation of antigen：By the method for molecular cloning Galectin-3, HAAH and CYFRA21-1 gene clonings
To carrier for expression of eukaryon, and the expression of albumen is realized in mammalian cell, obtain antigen after purification, this antigen also can conduct
Standard items are used；The preparation of the antigen can also be prepared using existing conventional method, no longer burdensome herein.
（2）Capture the preparation of antibody：By above-mentioned antigen immune mammal, it is anti-for capture to obtain corresponding monoclonal antibody
Body.The mammal is mouse and rabbit, preferably mouse.The preparation of the capture antibody can also use existing conventional method
Prepare, it is no longer burdensome herein.
The detection antibody includes galactose agglutinin, cytokeratin fragment antigen 21-1 (CYFRA21-1) and the mankind
Aspartyl B-hydroxylase（HAAH）Polyclonal antibody after HRP marks made from respectively.
The detection antibody is by galactose agglutinin, cytokeratin fragment antigen 21-1 (CYFRA21-1) and the mankind
Aspartyl B-hydroxylase（HAAH）Carrier for expression of eukaryon is cloned into, and the expression of albumen is realized in mammalian cell,
Corresponding antigen is obtained after purification, the polyclonal antibody after the progress HRP marks that the antigen immune mammal is obtained.Institute
It is as follows the step of including to state the preparation method of detection antibody：
（1）The preparation of antigen：By the method for molecular cloning galactose agglutinin, cytokeratin fragment antigen 21-1
And the gene cloning of human aspartyl's B-hydroxylase is to carrier for expression of eukaryon, and albumen is realized in mammalian cell
Expression, obtain antigen after purification, this antigen also can be used as standard items use；
（2）Detect the preparation of antibody：By above-mentioned antigen immune mammal, corresponding polyclonal antibody is obtained；
（3）HRP is marked：HRP marks are carried out to the polyclonal antibody.
The mammal is mouse and rabbit, preferably rabbit.HRP marks are carried out to the polyclonal antibody to be used
Conventional existing method at present.
Wherein, the galactose agglutinin preferably galactose galectin-3 or Galectin-8；The cell angle
The preferred serum cytokeratin fragment antigen 21-1 of quality fragment antigen 21-1.
Wherein, in the hole for the microtiter plate that the capture antibody can be coated in PVC material in advance, step can be simplified
Suddenly, detection efficiency is improved.
For the preparation method of lung cancer early metaphase quick diagnosis reagent kit, it comprises the following steps：
（1）The preparation of antigen：By the method for molecular cloning by Galectin-3, HAAH and CYFRA21-1 gene clonings
To carrier for expression of eukaryon, and the expression of albumen is realized in mammalian cell, obtain required antigen after purification, wherein, institute
Stating antigen can also be used as standard items to use；
（2）Capture the preparation of antibody：Above-mentioned antigen immune mammal is obtained into corresponding monoclonal antibody, the Dan Ke
Capture antibody of the grand antibody as this kit；
（3）Detect the preparation of antibody：Above-mentioned antigen immune mammal is obtained into corresponding polyclonal antibody, to described more
Clonal antibody carries out HRP marks；The detection antibody has mark function, can save labelled antibody and addition labelled antibody
Operating procedure, it is effective cost-effective, reduce operating procedure, reduce detection error.
（4）Capture antibody coating：
1. the capture antibody that concentration is 1 μ g/ml is coated in micro drop by with carbonate/bicarbonate buffer solution (pH9.6)
The hole of fixed board；
2. covers microtiter plate with bond plastic product and is incubated overnight at 4 DEG C；
3. discards coating buffer（The capture antibody of carbonate/bicarbonate buffer solution dilution）, and washed with cleaning solution micro
Titer plate adds 200 μ l PBST in micropore twice, every time（Phosphate Tween buffer）, gently whipping is micro- above tank
Measure titer plate, remove cleaning solution, microtiter plate is patted on paper handkerchief, removes remaining drop, dry be put in it is standby in 4 DEG C of environment
With；The cleaning solution is PBS（Phosphate buffer）It is middle to add a certain amount of Tween20, the mass percent of the Tween20
Concentration is 0.05%；
（5）Closing
1. adds 200 μ l Block buffers in every hole of microtiter plate（For containing 1.2%BSA（Bovine serum albumin(BSA)）'s
PBS（Phosphate buffer））, remaining protein binding site in hole is coated with for closing；
2. covers microtiter plate and is incubated 1 hour at 37 DEG C.
For the detection method of lung cancer early metaphase quick diagnosis reagent kit, it comprises the following steps：
（1）The preparation of antigen：By the method for molecular cloning by Galectin-3, HAAH and CYFRA21-1 gene clonings
To carrier for expression of eukaryon, and the expression of albumen is realized in mammalian cell, obtain required antigen after purification, wherein, institute
Stating antigen can also be used as standard items to use；
（2）Capture the preparation of antibody：Above-mentioned antigen immune mammal is obtained into corresponding monoclonal antibody, the Dan Ke
Capture antibody of the grand antibody as this kit；
（3）Detect the preparation of antibody：Above-mentioned antigen immune mammal is obtained into corresponding polyclonal antibody, to described more
Clonal antibody carries out HRP marks；
（4）Capture antibody coating：
1. the capture antibody that concentration is 1 μ g/ml is coated with microtitration by carbonate/bicarbonates buffer solution (pH9.6)
The hole of plate；
2. covers microtiter plate with bond plastic product and is incubated overnight at 4 DEG C；
3. discards coating buffer（The capture antibody of carbonate/bicarbonate buffer solution dilution）, and washed with cleaning solution micro
Titer plate adds 200 μ l PBST in micropore twice, every time（Phosphate Tween buffer, can contain 0.05% Tween-20
PH7.4 phosphate buffer）, the gently whipping microtiter plate above tank, cleaning solution is removed, is patted on paper handkerchief micro
Titer plate, remove remaining drop, dry be put in it is standby in 4 DEG C of environment；The cleaning solution is PBS（Phosphate buffer）In plus
Enter a certain amount of Tween20, the mass percent concentration of the Tween20 is 0.05%；
（5）Closing
1. adds 200 μ l Block buffers in each hole of microtiter plate（1.2%BSA/PBS）, close in coating hole
Remaining protein binding site；
2. covers microtiter plate with bond plastic product and is incubated 1 hour at 37 DEG C；
（6）Sample-adding
1. suitably dilutes 100 μ l（20 times of dilution）Sample be added to each hole of microtiter plate, incubated at 37 DEG C
Educate 60 minutes;Obtain accurate quantitative result, it is common practice that compare the signal of unknown sample and standard curve.Each enzyme mark
Plate all must bioassay standard product（Double measure or triplicate）And blank sample, to ensure accuracy；
2. discards sample, and washs microtiter plate three times, 200 μ l PBST are added in micropore every time（Phosphate is told
Warm buffer solution）；
3. the detection antibody that 100 μ l concentration are 0.5 μ g/ml is added to each hole by；
4. covers microtiter plate with bond plastic product and is incubated 1 hour at 37 DEG C；
5. washs microtiter plate four times with PBST；
(7) detect
1. is by TMB（3,3 ', 5,5 '-tetramethyl benzidine）Solution is added to each hole, is incubated 15-30 minutes, addition etc.
Terminate liquid (the 2M H of volume2SO4), optical density is then read at 450nm with enzyme-linked immunosorbent assay instrument.
2. the data that is obtained by serial dilutions draw standard curve, concentration is marked on X-axis（Logarithmic scale）On, and extinction
Scale is in Y-axis（Lineal scale）On.Sample concentration is drawn on this standard curve by interpolation method.
The present invention, which is used for lung cancer early metaphase quick diagnosis reagent kit, includes CBP-35（Galectin-3）, the mankind
Aspartyl B-hydroxylase（HAAH）, two in tri- indexs of CYFRA21-1 raise as diagnosis early metaphase mammary gland simultaneously
The standard of cancer, Cleaning Principle are as shown in Figure 1；CBP-35（Galectin-3）, human aspartyl's B-hydroxylase
（HAAH）, two standards declined simultaneously as lung cancer therapy recruitment evaluation in tri- indexs of CYFRA21-1.It can be used for facing
Dynamic evaluation is carried out to the therapeutic effect of lung cancer by detecting the height of 3 tumor marker levels in blood on bed；May be used also in addition
For the application of the clinically relapse and metastasis to lung cancer and Index for diagnosis.
Test effect explanation
The selection of double-antibody sandwich elisa optimum experimental condition
It is coated with the selection of mouse anti-human Galectin-3, HAAH and CYFRA21-1 monoclonal antibody best effort concentration：According to square formation method
It is determined that when coating concentration is 1ug/mL, the OD values of monoclonal antibody are 1.03, so its optimal coating concentration is 1ug/mL.Such as figure
Shown in 2, the selection of the how anti-best effort concentration of rabbit-anti people Galectin-3, HAAH and CYFRA21-1：With detection antibody dilution
The increase of multiple, cases of lung cancer serum and normal human serum OD values to be measured have the trend successively decreased, when antibody concentration is 1：When 200,
(normal control OD values subtract blank control OD to positive control (positive control OD values subtract blank control OD values) with normal control
Value) the ratio between A450nm (abbreviation P/N values) is higher, therefore selection rabbit-anti human antibody best effort concentration is 1：200.Serum is groped most
Good working concentration is 1：25.It is 1.2%BSA that confining liquid, which gropes best effort solubility,.
Clinical serum Samples detection
400 parts of serum specimens are have detected altogether, using hospital through being diagnosed as lung cancer I-II phases patients serum as positive controls
（110）, other benign patients and normal population serum are negative control group（290（Normal 170, benign 120
Example））, PBST is blank control, carries out qualitative by above-mentioned double crush syndrome method and quantitatively detects clinical serum sample.Such as
Shown in following table：
With P/N values>2 be double crush syndrome Positive judgement standards, and clinical serum sample is entered using pathological diagnosis as standard
Row detection, lung cancer result detection sensitivity（SN）For 95.45%（105/110）, accuracy rate（SP）Reach 95.25%（381/
400）.
The clinical therapeutic effect to lung cancer carries out dynamic evaluation
To determine that can this kit be used to carry out dynamic evaluation to the therapeutic effect of breast cancer, we have collected 102 parts
Serum before and after treatment in patients with lung cancer.The testing result of 102 patients is referring to Fig. 3-5, before and after as a result showing treatment in patients with lung cancer
Serum in Galectin-3, HAAH and CYFRA21-1 horizontal there were significant differences（P<0.05）.Effectively after treatment in serum
Galectin-3, HAAH and CYFRA21-1 level can be greatly reduced, and prompt this kit to be used for the therapeutic effect to lung cancer
Carry out dynamic evaluation.
Preferred embodiment of the invention described in detail above, it will be appreciated that the ordinary skill of this area is without wound
The property made work can makes many modifications and variations according to the design of the present invention.Therefore, all technician in the art
According to present inventive concept in prior art basis by logic analysis, reasoning or according to the limited available technology of experiment
Scheme, should be among the protection domain determined by the claims.
Claims (1)
1. for the preparation method of lung cancer early metaphase quick diagnosis reagent kit, its feature is in it comprises the following steps：
(1) preparation of antigen：By galactose agglutinin, cytokeratin fragment antigen 21-1 and human aspartyl β-hydroxyl
Change the gene cloning of enzyme to carrier for expression of eukaryon, and the expression of albumen is realized in mammalian cell, obtain after purification required
Antigen；
(2) preparation of antibody is captured：Above-mentioned antigen immune mammal is obtained into corresponding monoclonal antibody；
(3) preparation of antibody is detected：Above-mentioned antigen immune mammal is obtained into corresponding polyclonal antibody, to described polyclonal
Antibody carries out HRP marks；
(4) antibody coating is captured：
1. the capture antibody that concentration is 1 μ g/ml is coated with the hole of microtiter plate by carbonate/bicarbonates buffer solution；②.
Capping microtiter plate is simultaneously incubated overnight at 4 DEG C；3. discards coating buffer, and washs microtiter plate twice with cleaning solution, often
It is secondary that 200 μ l PBST are added in micropore, cleaning solution and remaining drop are removed, dries and is put in 4 DEG C of environment；
(5) close：200 μ l Block buffers are added per hole, closing is coated with remaining protein binding site in hole.
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Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101363051A (en) * | 2008-08-29 | 2009-02-11 | 芮屈生物技术（上海）有限公司 | Kit for GALECTIN-3 gene hybridization in situ, detection method and application thereof |
| CN101377501A (en) * | 2007-08-31 | 2009-03-04 | 北京科美东雅生物技术有限公司 | Cell keratin 19 fragments chemiluminescence immune analysis determination reagent kit and preparing method thereof |
| WO2012115885A1 (en) * | 2011-02-22 | 2012-08-30 | Caris Life Sciences Luxembourg Holdings, S.A.R.L. | Circulating biomarkers |
| CN103033619A (en) * | 2013-01-08 | 2013-04-10 | 河南生生医疗器械有限公司 | Protein chip reagent kit and method for comprehensively detecting lung cancer marker |
| CN103163293A (en) * | 2012-06-19 | 2013-06-19 | 中国医学科学院肿瘤医院 | Test kit of auxiliary diagnosis of non-small cell lung cancer patients |
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Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101377501A (en) * | 2007-08-31 | 2009-03-04 | 北京科美东雅生物技术有限公司 | Cell keratin 19 fragments chemiluminescence immune analysis determination reagent kit and preparing method thereof |
| CN101363051A (en) * | 2008-08-29 | 2009-02-11 | 芮屈生物技术（上海）有限公司 | Kit for GALECTIN-3 gene hybridization in situ, detection method and application thereof |
| WO2012115885A1 (en) * | 2011-02-22 | 2012-08-30 | Caris Life Sciences Luxembourg Holdings, S.A.R.L. | Circulating biomarkers |
| CN103163293A (en) * | 2012-06-19 | 2013-06-19 | 中国医学科学院肿瘤医院 | Test kit of auxiliary diagnosis of non-small cell lung cancer patients |
| CN103033619A (en) * | 2013-01-08 | 2013-04-10 | 河南生生医疗器械有限公司 | Protein chip reagent kit and method for comprehensively detecting lung cancer marker |
Non-Patent Citations (2)
| Title |
|---|
| 半乳糖凝集素3及骨桥蛋白在非小细胞肺癌亚临床转移中的意义;梁勇等;《中华外科杂志》;20090731;第47卷(第14期);1061-1063 * |
| 血清肿瘤标记物CEA和CYFRA21-1在肺癌临床诊断中的意义;张爱霞等;《放射免疫学杂志》;20091231;第22卷(第1期);95-96 * |
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