CN103048467A - Method for detecting tissue cell protein by using housekeeping protein - Google Patents
Method for detecting tissue cell protein by using housekeeping protein Download PDFInfo
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- CN103048467A CN103048467A CN2012105201489A CN201210520148A CN103048467A CN 103048467 A CN103048467 A CN 103048467A CN 2012105201489 A CN2012105201489 A CN 2012105201489A CN 201210520148 A CN201210520148 A CN 201210520148A CN 103048467 A CN103048467 A CN 103048467A
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Abstract
The invention relates to a method for detecting tissue cell protein by using housekeeping protein. According to the method, housekeeping protein and a monoclonal antibody or a polyclone antibody of protein to be detected are coupled with colloidal gold or colloidal silver respectively, the colloidal gold or silver antibody compound formed after coupling is dripped on a gold marking pad of a test paper; and another monoclonal antibody or polyclone antibody against corresponding housekeeping protein or protein to be detected is fixed at a detection area of the test paper, and the negative or the positive of the protein detected is judged according to the relative calculation ratio of the color of the housekeeping protein on the test paper to the color of the detected protein. The detection method solves the quantitative problem in colloidal gold immunoassay, and widens the application of colloidal gold immunoassay in protein detection; and in addition, the detection method has the advantages of fastness in speed, high accuracy and the like compared with other protein detection.
Description
Technical field
The present invention relates to a kind of histocyte protein quantification detection method, particularly a kind of method of utilizing house keeping protein that histocyte albumen is quantitatively detected.
Background technology
Oncotherapy in the past, particularly chemotherapy because drug effect is in extensive range, in the kill tumor cell, have also been killed a large amount of normal cells, have caused insufferable toxic and side effect.Along with the rapid progress of medical research, scientists has been developed neoplasm targeted therapy medicine with strong points, that toxic and side effect is little gradually.At present, this targeted therapy is because good, the little development trend that more and more becomes oncotherapy of toxic and side effect of therapeutic action.Yet, how can in clinical, judge fast and accurately the molecule type of patient tumors, and then determine whether this patient is applicable to certain targeted therapy scheme and is still the difficult problem that present medical circle is badly in need of solution.
Take breast cancer as example, the patient with breast cancer of the about 20%-30% of China is human epidermal growth factor acceptor-2 (HER2) positive breast cancer.The patient of the HER2 positive compares with other breast cancer patients, and malignancy is higher, make progress faster, easier recurrence and to DISTANT METASTASES IN, and the common Endocrine treatment of this type of patient is insensitive, prognosis is relatively poor.So several international drugmakers develop the target therapeutic agent for the HER2 positive breast cancer, such as Trastuzumab, have received reasonable result for the treatment of.Yet, this class target medicine is only effective to the patient with breast cancer of the HER2 positive, for the patient of 70%-80%HER2 feminine gender but without any result for the treatment of, if untimely diagnosis is clear and definite before treatment, not only delay treatment, also can bear heavy financial burden (medical expense) to the patient.Therefore, it is significant for its targeted therapy how in the shortest time HER2 expression in the patients with breast cancer tissue (negative or positive) to be made accurate judgement.
At present the cancer cell tumor tissues is detected main employing immunohistochemistry (IHC) and detect the HER2 protein expression level.Although SABC (IHC) has been passed through the FDA approval, the selection of different detection methods, the accuracy of calculating and reagent all can affect the result that HER2 detects.An investigation result of the U.S. shows that nearly 1/4 patient has accepted inappropriate treatment because testing result is inaccurate.IHC detects the error rate average out to 18% of HER2 protein overexpression, and FISH detects the error rate of HER2 gene magnification 13%.Therefore, China virologist reaches common understanding after according to up-to-date both at home and abroad data discussion, works out in October, 2006 and has issued China's " breast cancer HER2 guide detection ".American Society of Clinical Oncology (ASCO) and U.S. pathology medical college (CAP) also unite on Dec 11st, 2006 and have issued " the ASCO/CAP guide common recognition that breast cancer HER2 detects ".During having emphasized to detect, these guides are prone to link, inside and external mass control and the assurance program of error, be intended to make running program that HER2 detects and to the standardization of as a result interpretation, improve repeatability and accuracy that HER2 detects, filter out more accurately the patient with breast cancer who is applicable to the drug therapies such as Herceptin (Trastuzumab), avoid futile treatment, make the patient bear unnecessary financial burden.
In addition, a kind of method one gold medal mark method of clenbuterol hydrochloride etc. being carried out fast detecting is arranged in the market.Gold mark method is a kind of solid phase labelling immunoassays new technology that grows up on the basis of enzyme linked immunosorbent assay.It detects principle, take miillpore filter as carrier, coated known antigens, add sample to be checked after, through the capillarity of filter membrane, the antigen combination that the antigen in the sample is combined with collaurum is combined colour developing by the antigen that is coated with again and is reached testing goal on film.The reagent of gold mark method is the test strips form, is gold label test strip.
But golden mark method not only sensitivity is too low, and the experimental error between different batches is too large, and, can only be by developing the color to judge having and nothing of detected albumen, can not interpretation by the absolute quantitation of Reichl's test or relative quantification.Therefore, existing golden mark method can't detect the albumen in the histocyte at all.So, develop a kind of technology that tumor tissues point of impact on target protein expression level is carried out fast, accurately detects, not only in the colloid immunoassay technology, belong to a kind of breakthrough, significant, and have widely market application foreground.
Summary of the invention
The shortcoming of prior art the object of the present invention is to provide a kind of method of utilizing house keeping protein that histocyte albumen is quantitatively detected in view of the above.
Reach for achieving the above object other relevant purposes, the invention provides a kind of method of utilizing house keeping protein that histocyte albumen is detected, it comprises the steps:
1) with the monoclonal antibody of house keeping protein, detected albumen or how anti-respectively with collaurum or collargol coupling, collaurum or the collargol antibody complex that forms after the coupling dripped on the gold mark pad of test strips;
2) detection zone in test strips resists monoclonal antibody or many anti-being fixed on the test strips of corresponding house keeping protein or detected albumen with another;
3) calculate corresponding ratio according to house keeping protein colour developing on the test strips and the relative quantity that is detected the albumen colour developing, and then judge detected protein expression level to be negative or positive.
Preferably, described house keeping protein is GAPDH or Actin.
Preferably, described ELISA test strip district is provided with several and divides other to detect round dot by array-like.
As mentioned above, the method of utilizing house keeping protein that histocyte albumen is detected of the present invention has following beneficial effect: this detection method adopts house keeping protein is set when detecting, make house keeping protein and collaurum or collargol coupling, calculate corresponding ratio according to house keeping protein colour developing on the test strips and the relative quantity that is detected the albumen colour developing at last, and then judge detected protein expression level to be negative or positive.This detection method has solved a quantitative difficult problem in the colloid gold immune detection method, has improved detection sensitivity, has enlarged its application in Protein Detection; In addition, this detection method has the advantages such as speed is fast, degree of accuracy is high with respect to other protein detection technology.
Description of drawings
Fig. 1 is the front view of test strips of the present invention.
Fig. 2 is the vertical view of test strips of the present invention.
The element numbers explanation
1 base plate
2 polyester films
3 sample pad
4 gold medal mark pads
5 nitrocellulose membranes
6 detection zones
61 detect round dot
7 absorption layers
Embodiment
Below by specific instantiation explanation embodiments of the present invention, those skilled in the art can understand other advantages of the present invention and effect easily by the disclosed content of this instructions.The present invention can also be implemented or be used by other different embodiment, and the every details in this instructions also can be based on different viewpoints and application, carries out various modifications or change under the spirit of the present invention not deviating from.
See also Fig. 1,2.Need to prove, the diagram that provides in the present embodiment only illustrates basic conception of the present invention in a schematic way, so only show in graphic with the present invention in relevant assembly but not component count, shape and size drafting when implementing according to reality, kenel, quantity and the ratio of each assembly can be a kind of random change during its actual enforcement, and its assembly layout kenel also may be more complicated.
The below is further described technical solution of the present invention take breast cancer as example:
The method comprises the steps: at first as shown in Figure 1, 2, one test strips is set, this test strips comprises base plate 1, base plate 1 is provided with polyester film 2, polyester film 2 is provided with nitrocellulose membrane 5, sample pad 3, gold mark pad 4, nitrocellulose membrane 5 is provided with detection zone 6, absorption layer 7, under absorption layer 7 syphonic effects, and the movement that the liquid on the sample pad 3 can be from left to right.Detection zone 6 is provided with a plurality of detection round dots 61 by the array distribution of shapes.
When detecting, utilize collaurum or collargol and corresponding house keeping protein actin, the monoclonal antibody that reaches detected albumen Her2 or the coupling of how anti-difference, and collaurum or the collargol antibody complex that forms after the coupling dripped on gold mark pad shady and cool preservation the after the oven dry under 37 ° of C environment.Monoclonal antibody or resist of another anti-corresponding house keeping protein actin or detected albumen Her2 are fixed on the detection zone in the display mode more; After the detection zone colour developing in 5 minutes, number by house keeping protein colour developing round dot on the magnifier observation detection zone and detected albumen colour developing round dot, calculate corresponding ratio, and then judge that detected protein expression is positive or negative in this tumour cell, and then determine whether to adopt the target therapeutic agent (such as Trastuzumab) take Her2 as target spot to treat.
This detection method adopts house keeping protein is set when detecting, make house keeping protein and collaurum or collargol coupling, calculate corresponding ratio according to house keeping protein colour developing on the test strips and the relative quantity that is detected the albumen colour developing at last, and then judge detected protein expression level to be negative or positive.This detection method has solved in the colloid gold immune detection method the difficult problem of protein quantification, has enlarged its application in Protein Detection; In addition, this detection method has the advantages such as speed is fast, good reproducibility, degree of accuracy height with respect to other protein detection technology.In sum, the present invention has effectively overcome various shortcoming of the prior art and the tool high industrial utilization.
Above-described embodiment is illustrative principle of the present invention and effect thereof only, but not is used for restriction the present invention.Any person skilled in the art scholar all can be under spirit of the present invention and category, and above-described embodiment is modified or changed.Therefore, have in the technical field under such as and know that usually the knowledgeable modifies or changes not breaking away from all equivalences of finishing under disclosed spirit and the technological thought, must be contained by claim of the present invention.
Claims (3)
1. one kind is utilized house keeping protein to the method that histocyte albumen detects, and it is characterized in that it comprises the steps:
1) with the monoclonal antibody of house keeping protein, detected albumen or how anti-respectively with collaurum or collargol coupling, collaurum or the collargol antibody complex that forms after the coupling dripped on the gold mark pad of test strips;
2) on test strips with monoclonal antibody or many anti-detection zones that is fixed on test strips of another anti-corresponding house keeping protein or detected albumen;
3) according to house keeping protein colour developing and the relative quantity ratio calculated that is detected the albumen colour developing on the test strips, and then judge detected protein expression level to be negative or positive.
2. the house keeping protein that utilizes according to claim 1 is characterized in that the method that histocyte albumen detects: described house keeping protein is GAPDH or Actin.
3. the house keeping protein that utilizes according to claim 1 is characterized in that the method that histocyte albumen detects: described ELISA test strip district is provided with several detection round dots of arranging by array-like.
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| CN201210520148.9A CN103048467B (en) | 2012-12-06 | 2012-12-06 | Utilize the method that house keeping protein detects histocyte albumen |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103499691A (en) * | 2013-07-11 | 2014-01-08 | 苏州默锐克生物科技有限责任公司 | Detection test paper of HER2 protein expression level |
| WO2020233533A1 (en) * | 2019-05-17 | 2020-11-26 | Mei Jun | Test piece for rapid quantitative detection of tissue cell proteins |
Citations (5)
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| US20030073121A1 (en) * | 2001-09-17 | 2003-04-17 | Ib Mendel-Hartvig | Multi-analyte assay device with multi-spot detection zone |
| CN101344526A (en) * | 2008-08-22 | 2009-01-14 | 无锡纳生生物科技有限公司 | Fast and accurate prostatic cancer detection test paper strip and its preparation and application |
| CN102072956A (en) * | 2010-12-20 | 2011-05-25 | 重庆市科学技术研究院 | Nano influenza virus testing paper |
| CN201917575U (en) * | 2010-12-20 | 2011-08-03 | 重庆市科学技术研究院 | Nano influenza virus detection test paper |
| WO2012099897A1 (en) * | 2011-01-18 | 2012-07-26 | Symbolics, Llc | Lateral flow assays using two dimensional features |
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Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20030073121A1 (en) * | 2001-09-17 | 2003-04-17 | Ib Mendel-Hartvig | Multi-analyte assay device with multi-spot detection zone |
| CN101344526A (en) * | 2008-08-22 | 2009-01-14 | 无锡纳生生物科技有限公司 | Fast and accurate prostatic cancer detection test paper strip and its preparation and application |
| CN102072956A (en) * | 2010-12-20 | 2011-05-25 | 重庆市科学技术研究院 | Nano influenza virus testing paper |
| CN201917575U (en) * | 2010-12-20 | 2011-08-03 | 重庆市科学技术研究院 | Nano influenza virus detection test paper |
| WO2012099897A1 (en) * | 2011-01-18 | 2012-07-26 | Symbolics, Llc | Lateral flow assays using two dimensional features |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103499691A (en) * | 2013-07-11 | 2014-01-08 | 苏州默锐克生物科技有限责任公司 | Detection test paper of HER2 protein expression level |
| WO2020233533A1 (en) * | 2019-05-17 | 2020-11-26 | Mei Jun | Test piece for rapid quantitative detection of tissue cell proteins |
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