CN108950002A - Application and kit based on Caveolin-1 detection chemosensitivity of breast cancer - Google Patents
Application and kit based on Caveolin-1 detection chemosensitivity of breast cancer Download PDFInfo
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Abstract
The invention discloses Caveolin-1 or its correlated series in the application of following either sides: in terms of i) developing, screening the product for being suitable for detecting chemosensitivity of breast cancer；Ii) preparation is suitable for detecting the product aspect of chemosensitivity of breast cancer.The invention also discloses the kits based on Caveolin-1 as molecular marker.The present invention can be used for predicting the sensibility of mammary cancer chemotherapy, and then instruct prediction patient to the sensibility and prognosis of chemotherapy, it can be used for the sensibility that auxiliary judgment patient includes the chemotherapeutics such as alkylating agent, antimetabolite, antitumor antibiotics, plant antineoplastic such as taxol to common chemotherapeutics, provide reference to chemotherapy medication decision.
Description
Technical field
The present invention relates to chemosensitivity of breast cancer detection fields, and in particular to detects mammary gland cancerization based on Caveolin-1
Treat application and the kit of sensibility.
Background technique
Breast cancer is to threaten the most common malignant tumour of women life and health, and disease incidence occupies first of female malignant.
There are about 1,400,000 breast cancer newly to send out patient every year in the whole world at present.China's breast cancer incidence also occupies women and is susceptible to suffer from tumour first place, and
It is incremented by with annual 2~3% rate of rise, while age of onset tends to rejuvenation.Breast cancer has become the great of current social
Public health problem.Clinically, chemotherapy is still before breast cancer operation and postoperative key agents treatment means.Particularly with HER2 sun
Property and triple negative breast cancer invasion high in this way, the pathological of transfer easy to recur, chemotherapy prognosis be the pass for influencing survival
Key.Therefore, the biomarker of prediction chemosensitivity of breast cancer, the inspection of exploitation detection chemotherapy for carcinoma of breast sensibility are found
Survey method improves the therapeutic effect of patient with breast cancer, with important clinical meaning for optimizing the therapeutic scheme of patient with breast cancer
Justice.
Tumor stem cell (Cancer stem cells, CSCs) refers in tumour that content is rare, has self-renewing and more
To the special cells subgroup of differentiation potential.Tumor stem cell theory thinks that tumor stem cell is the initiating cell of tumor development
And driving source.Successfully separation obtains breast cancer tumour stem cell to Michael Clarke in 2003 from breast cancer tissue
(breast carcinoma stem cell).Compared to non-breast cancer stem cell, breast carcinoma stem cell has powerful internal oncogenicity and transfer latent
Energy.Importantly, breast carcinoma stem cell has the drug resistance of height to chemotherapy.Conventional chemotherapy can kill a large amount of point in a short time
Change type breast cancer cell but lacks the specific aim killing to breast carcinoma stem cell, cannot be fully erased by its.Once body inner ring
Border is suitble to, and the breast carcinoma stem cell of a small amount of remaining " can stage a comeback " and then lead to the recurrence of breast cancer.And it recurs exactly
The main reason of the failure of breast cancer clinical treatment and high mortality.To sum up, breast carcinoma stem cell is mammary gland carcinogenesis hair
The root of exhibition is the key factor for determining chemosensitivity of breast cancer, breast cancer caused by breast carcinoma stem cell chemotherapy tolerance
Recurrence has become the bottleneck for restricting breast cancer clinical therapeutic efficacy.
Caveolin-1 is the main constitutive protein of cell membrane surface caveolae Caveolae, extensive biological study report
Road Caveolin-1 is adjusting mitochondrial function, cell endocytic, cholesterol transport, cell membrane assembling, signal transduction and tumour life
At etc. play an important role in biological processes.Caveolin-1 albumen is encoded by CAV1 gene, and CAV1 gene is located at people
The region Chromosome 7q31 .1, the region are a brittle zone, are easily lacked in tumor patient, prompt Caveolin-1 in cream
The effect of tumor suppressor gene may be shown as in gland cancer generating process.Existing research reports that Caveolin-1 is including breast cancer, pancreas
Several Kinds of Malignancy including gland cancer, colon cancer, gastric cancer, prostate cancer, leukaemia, melanoma, osteosarcoma, rhabdomyosarcoma
The effect of cancer suppressor protein is shown as in generating process.As preclinical study find galactophore epithelial cell vicious transformation during it is adjoint
Caveolin-1 expression reduction.Clinical research discovery mammary gland stroma cell Caveolin-1 expression deletion can be used as breast cancer
The biomarker of early stage recurrence, lymphatic metastasis and low survival rate.It is interesting that the expression of Caveolin-1 and function are swollen
Can fluctuate during tumor occurrence and development, in tumour stage of development, Caveolin-1 as stress protein, normal level
Expression can protect normal cell from the influence of external source sexual stimulus pressure and then prevent the vicious transformation of normal cell.In tumour evening
Phase, Caveolin-1 high under the induction of external source sexual stimulus can be expressed, and play the work for promoting metastases, mediate drug resistance generation
With.
Currently, chemotherapy is still the key agents treatment means that clinically patient with breast cancer is preoperative and postoperative, however at present
Influence of the Caveolin-1 to breast cancer cell drug resistance, and to breast carcinoma stem cell this decision mammary cancer chemotherapy final result
The influence of key cells subgroup drug resistance is still not clear.So far whether Caveolin-1 it is sensitive can be used as detection mammary cancer chemotherapy
Property effective molecular marker, Caveolin-1 as molecular marker detection chemosensitivity of breast cancer field application all
There is not been reported.
Summary of the invention
In order to overcome deficiencies of the prior art, inventor further studies and specifies Caveolin-1 and mammary gland
Relationship between cancer cell and breast carcinoma stem cell, and propose and apply Caveolin-1 or its correlated series in detection mammary gland
In terms of cancer chemotherapy sensibility.The invention is realized by the following technical scheme: Caveolin-1 or its correlated series are following any
The application of aspect:
I) in terms of developing, screening the product for being suitable for detecting chemosensitivity of breast cancer；
Ii) preparation is suitable for detecting the product aspect of chemosensitivity of breast cancer.
Preferably, the product includes kit.
Preferably, the correlated series of the Caveolin-1 include the mRNA of the Caveolin-1.
Preferably, the sample source of the kit detection includes tumor tissues or blood.
Preferably, the kit is used to detect the mRNA of Caveolin-1 in the tumor tissues, or for detecting
The Caveolin-1 of the blood.
Based on the studies above, inventor also proposed the kit for detecting chemosensitivity of breast cancer, wherein a kind of
Based on PCR technology detects the mRNA expression of patients with breast cancer tissue sample Caveolin-1, and then prejudges breast cancer
Chemotherapy in Patients sensibility can be achieved through the following technical solutions:
A kind of kit detecting chemosensitivity of breast cancer, the sample source of detection is in breast tumor tissue, packet
Primer, the first reference substance and the second reference substance are included, the primer detects the mRNA table of Caveolin-1 in the sample for PCR
Up to level, first reference substance includes the standard cDNA that the breast tumor tissue of chemotherapy medicament sensitive extracts, and described second
Reference substance includes the standard cDNA that the breast tumor tissue of chemotherapy resistance extracts.
Another kind is based on double-antibody sandwich elisa technology, passes through the table of Caveolin-1 in detection blood serum of patients with human breast carcinoma
It up to level, and then predicts chemotherapy for carcinoma of breast sensibility, can be achieved through the following technical solutions:
A kind of kit detecting chemosensitivity of breast cancer, the sample source of detection is in blood, including source of people
Caveolin-1 antibody and Caveolin-1 protein standard substance, the source of people Caveolin-1 antibody and the Caveolin-1 egg
White standard items quantitative determine the expression of Caveolin-1 in the sample for ELISA.
It preferably, further include ELISA Plate, the Caveolin-1 protein standard substance dilution, detection solution, tmb substrate
And reaction terminating liquid, the source of people Caveolin-1 antibody are coated with the ELISA Plate.
The present invention is based on Caveolin-1 as molecular marker, can be used for predicting the sensibility of mammary cancer chemotherapy, in turn
Guidance prediction patient to the sensibility and prognosis of chemotherapy, can be used for auxiliary judgment patient to common chemotherapeutics include alkylating agent,
The sensibility of the chemotherapeutics such as antimetabolite, antitumor antibiotics, plant antineoplastic such as taxol, to chemotherapy medication decision
Reference is provided.
Detailed description of the invention
Invention is further described in detail in the following with reference to the drawings and specific embodiments.
Fig. 1: breast carcinoma resistance cell and breast carcinoma stem cell height express Caveolin-1；
Fig. 2: Caveolin-1 expression up-regulation can mediate the generation of breast cancer cell drug resistance；
Fig. 3: Caveolin-1 expression up-regulation can mediate the generation of breast carcinoma stem cell drug resistance；
Fig. 4: breast cancer cell and breast tumor tissues Caveolin-1 can express up-regulation, and companion under chemotherapy stimulation induction
With the up-regulation of mammary cancer chemotherapy drug resistance index of correlation；
Fig. 5: chemotherapy stimulates the table of the expression of lower Caveolin-1 and ABCG2 the and β-catenin of breast cancer tissue
Up to horizontal significant positive correlation；
Fig. 6: Caveolin-1 strikes the low tumor of breast for increasing breast carcinoma stem cell formation to the chemotherapy of epirubicin
Sensibility.
Specific embodiment
The present invention is further described below:
One: a variety of inside and outside experiments confirm correlation of the Caveolin-1 with chemosensitivity of breast cancer
(1) breast carcinoma resistance cell and breast carcinoma stem cell height express Caveolin-1
Experimental result as shown in Figure 1 confirms that the Caveolin-1 expression of breast carcinoma resistance cell MCF-7/ADR is significant
Higher than mammary cancer chemotherapy sensitive cells MCF-7, the process is along with breast carcinoma resistance key regulatory albumin A BCG2 and breast cancer
The expression of stem cell key regulatory albumen β-catenin is raised.Stem cell-like cell and tumor stem cell are to influence chemotherapy of tumors
The key factor that drug resistance generates.Breast carcinoma resistance cell MCF-7/ADR is dry compared to mammary cancer chemotherapy sensitive cells MCF-7 thin
Born of the same parents' like cell ratio increases, and epirubicin accumulation reduces (Figure 1A-C).In addition, in mammary stem cells like cell subgroup and tumour
In stem cell subgroup, expression of the expression of Caveoin-1 respectively compared to non-stem cell like cell, non-tumor stem cell significantly increases
Add, (Fig. 1 D-E) is raised in the expression of above-mentioned change simultaneous ABCG2 and β-catenin.In summary, breast carcinoma resistance is thin
Born of the same parents and the high expression Caveolin-1 of breast carcinoma stem cell, Caveolin-1 can be used as breast carcinoma resistance cell and breast cancer is dry thin
The molecular marker of born of the same parents.
(2) the expression up-regulation of Caveolin-1 can mediate the generation of breast cancer cell and breast carcinoma stem cell drug resistance
Experimental result as shown in Figure 2 further confirms breast cancer MCF-7 and breast cancer MDA-MB-231 cell Caveolin-
1 overexpression can reduce above-mentioned cell to the chemosensitivity of breast cancer front-line chemotherapeutic agents Taxol.And strike low MCF-7 and
The expression of MDA-MB-231 cell Caveolin-1 can increase above-mentioned cell to the chemosensitivity of chemotherapeutics Taxol.
Experimental result as shown in Figure 3 further confirms the overexpression of Caveolin-1 and knocks out not influence breast carcinoma stem cell
Cell viability, but the overexpression of Caveolin-1 can mediate breast carcinoma stem cell to increase the drug resistance of epirubicin, mammary gland
The expression of cancer drug resistance key regulatory albumin A BCG2 and breast carcinoma stem cell key regulatory albumen β-catenin increase, breast cancer
Epirubicin accumulation is reduced in stem cell.On the contrary, breast carcinoma stem cell can be increased to epirubicin by striking low Caveolin-1
Chemosensitivity lowers the expression of breast carcinoma stem cell ABCG2 and β-catenin, increases epirubicin in breast carcinoma stem cell
Accumulation.
In summary, the expression up-regulation of Caveolin-1 can mediate the production of breast cancer cell and breast carcinoma stem cell drug resistance
It is raw.
(3) chemotherapy stimulation can further inducing mammary cancer cell and breast cancer tissue Caveolin-1 expression up-regulation, increase
Add the ratio of breast carcinoma stem cell
Epirubicin, taxol, 5-Fluorouracil are the First-line chemotherapy schemes of clinically patient with breast cancer, as shown in figure 4,
Through experiment in vitro confirm said medicine can concentration dependent up-regulation breast cancer MCF-7 and MDA-MB-231 cell
The expression of Caveolin-1, simultaneous ABCG2 (breast carcinoma resistance key regulatory albumen) and β-catenin (breast cancer
Stem cell key regulatory albumen) expression raise (Fig. 4 A-B).Further experiment in vivo confirms that epirubicin is inhibiting nude mice cream
While gland cancer growth of transplanted human, the expression of Caveolin-1, ABCG2 and β-catenin in breast tumor tissues can be raised, is increased
Add the ratio (Fig. 4 C-E) of breast carcinoma stem cell in breast cancer tissue.Above-mentioned discovery shows breast cancer cell and breast tumor tissues
Caveolin-1 can express up-regulation under chemotherapy stimulation induction, and along with the up-regulation of mammary cancer chemotherapy drug resistance index of correlation.
As shown in figure 5, comparing 10 chemotherapy for carcinoma of breast front and back Caveolin-1's by extracting blood total serum IgE
As a result the change of mRNA expression confirms that after the chemotherapy in 6 periods, the Caveolin-1mRNA level of patient with breast cancer is aobvious
It writes and increases (Fig. 5 A).The above results show that chemotherapy stimulation can further inducing mammary cancer cell and breast cancer tissue Caveolin-1
Expression up-regulation.It is found in addition, carrying out analysis to the breast tumor tissues of 46 patient with breast cancers, Caveolin-1 is in three feminine genders
Relatively high expression in patient with breast cancer, this clinical manifestation that is poor with triple negative breast cancer Chemotherapy in Patients effect, easily shifting are consistent (figure
5A).At the same time, immunohistochemical analysis discovery is carried out to the clinical tumor tissue of 46 patient with breast cancers, Caveolin-1's
The expression of expression and ABCG2 the and β-catenin of breast cancer tissue are significantly positively correlated (Fig. 5 B).
The above results further demonstrate the relationship between Caveolin-1 expression and chemosensitivity of breast cancer.
(4) tumour that the breast carcinoma stem cell of Caveolin-1 low expression is formed increases the chemosensitivity of epirubicin
Divide from the breast cancer MDA-MB-231 cell of different Caveolin-1 expressions by using flow cytometry
Breast carcinoma stem cell is selected, is then inoculated with the breast carcinoma stem cell of different Caveolin-1 expressions to the cream of NOD/SCID mouse
Gland fat pad constructs breast cancer transplantable tumor model.As shown in fig. 6, confirmed on above-mentioned model Caveolin-1 strike it is low can
Increase chemosensitivity of the tumor of breast to epirubicin of breast carcinoma stem cell formation, which provides Caveolin-1 work
For the positive evidence for predicting tumor of breast chemosensitivity molecular marker.
For two, the present invention is based on above-mentioned Caveolin-1 research relevant to chemosensitivity of breast cancer, proposing will
In terms of Caveolin-1 or its correlated series are applied to the product of detection chemosensitivity of breast cancer, in terms of kit.
As an example, the present embodiment proposes a kind of mRNA expression water for detecting breast tumor tissues Caveolin-1
Kit that is flat and then prejudging chemosensitivity, details are as follows:
Sample requirement
The fresh breast tumor tissues that breast tumor tissues detected should be operation excision or tissue biopsy procedure obtains,
It can also be the breast tumor tissues deposited in RNAlater liquid.
Reagent constituents
Kit includes following components:
Component 1-4 is the following primer dry powder of sequence, each 2OD, and when use is diluted to 10 μM of working solution with DEPC water.
Component 1:F:5'-gagcgagaagcaagtgtacga-3' is labeled as 1；
Component 2:R:5'-acagacggtgtggacgaagat-3' is labeled as 2；
Component 3:F:5'-gactaaccctgcgctcctg-3' is labeled as 3；
Component 4:R:5'-gcccaatacgaccaaatcag-3' is labeled as 4；
Component 5: reference substance 1, for the standard cDNA each 1 of the patients with breast cancer tissue extraction to chemotherapy medicament sensitive
Branch, is respectively labeled as 5A, 5B, 5C, 5D, 5E, 5F, 5G, 5H, 5I, 5K by totally 10, every 50 μ l, when experiment each 20 μ l
2 μ l reference substances 1 are added in pcr amplification reaction system.
Component 6: reference substance 2, for the standard cDNA of the patients with breast cancer tissue extraction to chemotherapy resistance, totally 10,
Every 50 μ l, are respectively labeled as 6A, 6B, 6C, 6D, 6E, 6F, 6G, 6H, 6I, 6K, when experiment each 20 μ l PCR reaction system
2 μ l reference substances 2 of middle addition.
Said components freeze under the conditions of being placed in -20 DEG C, as early as possible using finishing after dilution use, avoid freezing repeatedly.
Required preparation article except kit
Including but not limited to 8 needed for chloroform, isopropanol, 75% ethyl alcohol, RNAase-free water or DEPC water, PCR reaction
The pipettor and corresponding suction nozzle, Real Time PCR amplification instrument of connecting leg, different ranges.Quotient needed for RNA reverse transcription and cDNA are expanded
Product kit.
Operating procedure is as follows:
1. taking fresh Mammary cancer tumor tissues or tissue biopsy tumor tissues, or it is placed in RNAlater liquid and saves
50~100mg of breast tumor tissues, after liquid nitrogen freezing, mortar grinder are at powder, be added RNA extract reagent extract Total
RNA.It is recommended to use Takara RNAiso Plus (Code No.9108) product, method is referring to its specification.Every 50~100mg
Breast tumor tissue samples add the RNAiso Plus of 1ml to be advisable.Extracted RNA precipitate is dissolved in suitable DEPC water
Afterwards, purity analysis, OD are carried out260/OD280Ratio can carry out next step experiment in 1.7~2.1 ranges.
2. being cDNA by its reverse transcription after pair extracted Total RNA row removal genomic DNA reaction.It is recommended to use
The PrimeScript of TakaraTMRT reagent Kit with gDNA Eraser (Perfect Real Time) product
(Code No.RR047A) carries out reverse transcription reaction.The Total extracted in step 1 is added in the reverse transcription reaction system of every 20 μ l
RNA 500ng is advisable.Reaction condition is as follows: 37 DEG C, 15min；85 DEG C are denaturalized 5 seconds.
3. breast tumor tissues cDNA row amplified reaction obtained in pair step 2.It is recommended to use Takara's
Premix Ex TaqTMII (Tli RNaseH Plus) product (Code No.RR820A) carries out amplification reaction.This part experiment
The middle component 5 provided using kit and component 6 (are the cDNA solution extracted, have been diluted to working concentration, when use
Every 20 μ l reaction system adds 2 μ l) it is used as control sample.CAV1 genetic test hole and house-keeping gene is respectively set in each sample
Detection hole, is separately added into the combination of component 1/2 and component 3/4 combines.
3.1 be CAV1 genetic test hole reaction system as follows:
3.2 be house-keeping gene detection hole reaction system as follows:
3.3 reaction conditions are as follows:
Interpretation of result:
After reaction, according to various kinds sample wells Ct value, 2 are utilized-△△CtMethod analyzes data, it is more detected to
Expression of the sample compared to control sample (6 sample well of component 5 and component).Such as the mRNA of test sample Caveolin-1
Relative expression levels are not higher than 5 each sample of component, show that the patient with breast cancer may be to chemosensitivity.Such as test sample
The mRNA relative expression levels of Caveolin-1 are not less than 6 each sample of component, show that the patient with breast cancer may be resistance to for chemotherapy
Medicine.In view of Caveolin-1 to the universal reactivity of chemotherapeutics, therefore this kit results can be used for auxiliary judgment patient couple
Common chemotherapeutics includes the chemotherapeutics such as alkylating agent, antimetabolite, antitumor antibiotics, plant antineoplastic such as taxol
Sensibility.
The kit of the present embodiment can accurately detect the coding base of the Caveolin-1 of patient with breast cancer's breast tumor tissues
Because of CAV1mRNA expression, by using kit offer clinically to chemosensitivity and drug resistant patients with breast cancer
The cDNA of tissue extraction judges the opposite Caveolin-1 expression of patient to be measured as control, and then instructs prediction
Patient is to chemosensitivity and prognosis.
As one another kind of example, it is horizontal that the present embodiment proposes a kind of Caveolin-1 detected in blood serum of patients with human breast carcinoma
And then the kit of chemosensitivity is prejudged, details are as follows:
Test sample acquisition and processing requirement
With vacuum blood collection blood sampling tube 5~10ml venous blood, 2h or 4 DEG C is placed naturally at room temperature overnight, after waiting for blood to solidify
It is centrifuged 20 minutes in 1000g, absorption upper serum sample is set 4 DEG C of refrigerators and temporarily saved, and completes detection as early as possible, avoids freezing repeatedly
Melt.Haemolysis, which occurs, in test sample will affect last testing result, therefore haemolysis sample should not carry out this detection.Before sample detection
Slowly equilibrium it should not should heat to room temperature and be allowed to melt.
Reagent constituents
Kit includes following components:
1 piece and its 1 piece of overlay film of component 1:96 orifice plate (pre-coated source of people Caveolin-1 antibody)；
Component 2:Caveolin-1 protein standard substance 1 is people Caveolin-1 protein freeze-dried powder, total 2ng；
1 bottle of total 20ml of component 3:Caveolin-1 protein standard substance dilution；
Component 4: detection solution A, 1 bottle of total 20ml；
Component 5: detection solution B, 1 bottle of total 20ml；
1 bottle of total 9ml of component 6:TMB substrate；
Component 7: 1 bottle of total 6ml of reaction terminating liquid；
Component 8: 1 bottle of total 600ml of cleaning solution；
In mentioned reagent box component, component 1, component 4 and component 5 save under the conditions of being placed in -20 DEG C, remaining reagent please be set
It is saved backup in 4 DEG C.
Required preparation article except kit
Microplate reader, multichannel micropipettor and suction nozzle, diluted sample including but not limited to 450 ± 10nm optical filter
The EP pipe of product, distilled water or deionized water and blotting paper etc..
Operating procedure is as follows:
1.1 using preceding slowly balanced to room temperature (18-25 DEG C) by all reagent and sample.
3 (the standard of component of 1ml is added in the preparation of 1.2 standard items that is, into component 2 (Caveolin-1 standard protein) bottle
Diluted protein solution dilution), it is stored at room temperature after covering about 10 minutes, while gently shaking and (avoiding blistering), concentration is
2000pg/ml (mother liquor).Then take mother liquor 0.5ml, with component 3 (standard dilutions) gradually sesquialter be diluted to 1000pg/ml,
Each 0.5ml of the standard sample of 500pg/ml, 250pg/ml, 125pg/ml, 62.5pg/ml, 31.2pg/ml, 15.6pg/ml.Mark
Quasi- product are please prepared in 15 minutes before use.
1.3 sample-addings: gauge orifice, sample to be tested hole, blank well are set respectively.8 hole of gauge orifice is set altogether, sequentially adds 100 μ l not
With concentration standard solution (2000pg/ml, 1000pg/ml, 500pg/ml, 250pg/ml, 125pg/ml, 62.5pg/ml,
31.2pg/ml and 15.6pg/ml), 100 μ l of test serum sample is added in sample to be tested hole, and 3 (standard items of component are added in blank well
Dilution) 100 μ l.
1.4 reactions: ELISA Plate adds overlay film, is placed in 37 DEG C of incubators, incubates 1 hour.
1.5 discard liquid in hole, and after drying 96 orifice plates, every hole adds 100 μ l to detect solution A (thaw rewarming before use), enzyme
Target overlay film is placed in 37 DEG C of incubators, is incubated 1 hour.
1.6 discard liquid in hole, and every hole is added the cleaning solution washing of 300 μ l, impregnates 2 minutes, enzyme is patted on blotting paper
Target removes all liq in hole.Remaining washing buffer is poured out after repeating above-mentioned board-washing process 3 times, ELISA Plate is fallen
It is buckled on blotting paper, the liquid remained in hole is all blotted.
1.7 every holes add 100 μ l to detect solution B, and ELISA Plate overlay film is placed in 37 DEG C of incubators, incubate 30min.
1.8 discard liquid in hole, drying, and board-washing 3 times, method is the same as our department step by step 1.6.
1.9 every holes add 90 μ l of tmb substrate solution, and ELISA Plate overlay film is placed in 37 DEG C of incubators and incubates, when gauge orifice color goes out
Now when apparent gradient blue, it can terminate and (be usually no more than 30min).It is anti-to terminate that subsequent every hole adds 50 μ l of stop bath
It answers.Such as there is irregular colour one, ELISA Plate please be shake gently so that solution is uniformly mixed.
The 1.10 optical density OD value with microplate reader in each hole of 450nm wavelength measurement.
Interpretation of result:
After each standard sample wells and sample aperture OD value deduct blank well OD value, using the concentration of standard items as abscissa, OD value is
Ordinate draws standard curve and calculates calibration curve equation.It brings the OD value of sample well into calibration curve equation, can calculate
The Caveolin-1 expression of blood serum of patients with human breast carcinoma sample out judges patient to be measured according to standard serum caveolin-1 value
Chemosensitivity and clinical prognosis.In view of Caveolin-1 to the universal reactivity of Treated with Chemotherapeutic Drugs object, therefore this kit is examined
Surveying result can be used for assisting prediction patient with breast cancer including alkylating agent, antimetabolite, anti-tumor to other common chemotherapeutics
The sensibility of the chemotherapeutics such as raw element, plant antineoplastic such as taxol.
The kit of the present embodiment is in double antibody sandwich ELISA quantitative determination blood serum of patients with human breast carcinoma
Caveolin-1 is horizontal, and then prediction patient can be instructed to chemosensitivity and clinical prognosis.
It should be understood that for those of ordinary skills, it can be modified or changed according to the above description,
And all these modifications and variations should all belong to the protection domain of appended claims of the present invention.
Illustrative description is carried out to the invention patent above, it is clear that the realization of the invention patent is not by aforesaid way
Limitation, as long as use the invention patent method concept and technical solution carry out various improvement, or it is not improved will this
The conception and technical scheme of patent of invention directly apply to other occasions, are within the scope of the invention.
Claims (8)
- The application of 1.Caveolin-1 or its correlated series in following either sides:I) in terms of developing, screening the product for being suitable for detecting chemosensitivity of breast cancer；Ii) preparation is suitable for detecting the product aspect of chemosensitivity of breast cancer.
- 2. application according to claim 1, which is characterized in that the product includes kit.
- 3. application according to claim 1, which is characterized in that the correlated series of the Caveolin-1 include described The mRNA of Caveolin-1.
- 4. application according to claim 2, which is characterized in that the sample source of the kit detection includes tumor tissues Or blood.
- 5. application according to claim 4, which is characterized in that the kit is for detecting in the tumor tissues The mRNA of Caveolin-1, or the Caveolin-1 for detecting the blood.
- 6. a kind of kit for detecting chemosensitivity of breast cancer, the sample source of detection is special in breast tumor tissue Sign is, including primer, the first reference substance and the second reference substance, and the primer detects Caveolin- in the sample for PCR 1 mRNA expression, first reference substance include the standard cDNA that the breast tumor tissue of chemotherapy medicament sensitive extracts, Second reference substance includes the standard cDNA that the breast tumor tissue of chemotherapy resistance extracts.
- 7. a kind of kit for detecting chemosensitivity of breast cancer, the sample source of detection is in blood, which is characterized in that including Source of people Caveolin-1 antibody and Caveolin-1 protein standard substance, the source of people Caveolin-1 antibody and described Caveolin-1 protein standard substance quantitative determines the expression of Caveolin-1 in the sample for ELISA.
- 8. kit according to claim 7, which is characterized in that further include ELISA Plate, the Caveolin-1 albumen mark Quasi- product dilution, detection solution, tmb substrate and reaction terminating liquid, the source of people Caveolin-1 antibody are coated with the enzyme mark Plate.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201811037980.7A CN108950002A (en) | 2018-09-06 | 2018-09-06 | Application and kit based on Caveolin-1 detection chemosensitivity of breast cancer |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201811037980.7A CN108950002A (en) | 2018-09-06 | 2018-09-06 | Application and kit based on Caveolin-1 detection chemosensitivity of breast cancer |
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| Publication Number | Publication Date |
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| CN108950002A true CN108950002A (en) | 2018-12-07 |
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| Application Number | Title | Priority Date | Filing Date |
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| CN201811037980.7A Pending CN108950002A (en) | 2018-09-06 | 2018-09-06 | Application and kit based on Caveolin-1 detection chemosensitivity of breast cancer |
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| CN (1) | CN108950002A (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN110184351A (en) * | 2019-05-29 | 2019-08-30 | 南通普惠精准医疗科技有限公司 | The application of detection kit |
| CN111939255A (en) * | 2019-05-16 | 2020-11-17 | 中央研究院 | Caveolin 1 antibodies for treating brain inflammation and injury and improving functional recovery |
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| CN103237901A (en) * | 2010-03-01 | 2013-08-07 | 卡里斯生命科学卢森堡控股有限责任公司 | Biomarkers for theranostics |
| CN106383235A (en) * | 2016-08-31 | 2017-02-08 | 中国人民解放军第三军医大学第附属医院 | Application of breast cancer multidrug resistant protein molecule IBP |
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| CN103237901A (en) * | 2010-03-01 | 2013-08-07 | 卡里斯生命科学卢森堡控股有限责任公司 | Biomarkers for theranostics |
| CN106383235A (en) * | 2016-08-31 | 2017-02-08 | 中国人民解放军第三军医大学第附属医院 | Application of breast cancer multidrug resistant protein molecule IBP |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111939255A (en) * | 2019-05-16 | 2020-11-17 | 中央研究院 | Caveolin 1 antibodies for treating brain inflammation and injury and improving functional recovery |
| CN110184351A (en) * | 2019-05-29 | 2019-08-30 | 南通普惠精准医疗科技有限公司 | The application of detection kit |
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Application publication date: 20181207 |