CN107741497A - For assessing the immunofluorescence detection agent box of liver cancer patient therapeutic effect - Google Patents
For assessing the immunofluorescence detection agent box of liver cancer patient therapeutic effect Download PDFInfo
- Publication number
- CN107741497A CN107741497A CN201710333555.1A CN201710333555A CN107741497A CN 107741497 A CN107741497 A CN 107741497A CN 201710333555 A CN201710333555 A CN 201710333555A CN 107741497 A CN107741497 A CN 107741497A
- Authority
- CN
- China
- Prior art keywords
- liver cancer
- antibody
- therapeutic effect
- cancer patient
- agent box
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Classifications
-
- 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
- G01N33/57438—Specifically defined cancers of liver, pancreas or kidney
Abstract
The invention belongs to biological technical field, is related to a kind of immunofluorescence detection agent box for being used to assess liver cancer patient therapeutic effect, and in particular to a kind of double antibody detection kit for being used to assess the effect that patients with hepatocellular carcinoma receives Sorafenib treatment.The immunofluorescence detection agent box of the present invention for being used to assess liver cancer patient therapeutic effect, including first antibody and secondary antibody, the first antibody are the combinations for being selected from the antibody for following protein molecular label：CD34 (cell differentiation antigen 34) and CXCR4 (C X C Chemokine receptor4s).The kit uses the judge index that double antibody combines, have the characteristics that it is objective and accurate, easy, directly perceived, be easy to repeat, can effectively assess liver cancer patient take Sorafenib therapeutic effect can be used for screening be applicable the liver cancer patient that Sorafenib is treated, so as to preferably help the suitable treatment crowd of doctor's selection, help improves therapeutic effect and extends the survival of patients time.
Description
Technical field
The invention belongs to biological technical field, is related to a kind of Immunofluorescence test for being used to assess liver cancer patient therapeutic effect
A kind of kit, and in particular to double antibody detection examination for being used to assess the effect that patients with hepatocellular carcinoma receives Sorafenib treatment
Agent box.
Background technology
Hepatocellular carcinoma liver cancer (abbreviation liver cancer) is one of most common malignant tumour, and death toll occupies China's malignant tumour
Three.Prognosis in hcc extreme difference, most of patient have lost operative chance in diagnosis, and reason is that liver cancer easily occurs in liver or liver
Outer transfer.Liver cancer angiogenesis enriches, and hematogenous metastasis is the major reason for causing liver cancer to be easy to shift and recur, therefore suppresses blood
Pipe generation is considered as to treat the Critical policies of liver cancer.Sorafenib is the first anti-angiogenic life being approved for treating liver cancer in the whole world
Into medicine, and it is currently the only confirm to extend the molecular targeted agents of liver cancer patient existence through research, but Sorafenib
The disease control rate for treating liver cancer is only 42-62%, and major reason is to lack clear and definite index at present to screen the drug therapy
Target group, patient be only capable of blindly receive Sorafenib treatment, for refractory patient not only delay treatment develop, also result in
The waste of huge economic loss and medical resource.
Clinic, which there is no, at present can be used for judging that liver cancer patient receives molecular marker or the inspection of Sorafenib therapeutic effect
Test agent box.
The content of the invention
It is used to assess the immunofluorescence detection agent box of liver cancer patient therapeutic effect it is an object of the invention to provide a kind of,
The kit use double antibody combine judge index, have the characteristics that it is objective and accurate, easy, directly perceived, be easy to repeatedly, Ke Yiyou
Effect assesses liver cancer patient and takes the therapeutic effect of Sorafenib, and the liver cancer trouble that Sorafenib treated is applicable available for screening
Person, so as to preferably help the suitable treatment crowd of doctor's selection, help improves therapeutic effect and extends the survival of patients time.
The immunofluorescence detection agent box of the present invention for being used to assess liver cancer patient therapeutic effect, including first antibody
And secondary antibody, the first antibody are the combinations for being selected from the antibody for following protein molecular label：CD34 (cell differentiations
Antigen 34) and CXCR4 (C-X-C Chemokine receptor4s).
Preferably, the secondary antibody is the secondary antibody that there is luciferase to mark.
By immunofluorescence detection agent box of the present invention, Immunofluorescence test is carried out to liver cancer tissue sample, it is comprehensive
The expression of the protein molecular label of analysis above two first antibody institute specific marker in the sample is closed, can be reached
Assess liver cancer patient and take the therapeutic effect after Sorafenib.
Immunofluorescence detection agent box of the present invention has advantages below：
(1) test method is highly developed, and detection process is easy, intuitively, is easy to repeat, can be with complete by member of ordinary skill
Into；
(2) the semi-automatic image quantitative analysis of human assistance can be used, it is as a result objective and accurate, better than artificial sxemiquantitative point
Analysis.
Brief description of the drawings
Fig. 1 is the result figure of CXCR4 positive vessels in immunofluorescence method detection liver cancer tissue.In figure, green fluorescence shows
Show CD34 molecules, red fluorescence shows CXCR4 molecules, and yellow shows the distribution of CXCR4 positive vessels.
Fig. 2 is survivorship curve analysis chart, before display liver cancer patient takes Sorafenib treatment, the CXCR4 in tumor tissues
The relation of overall survival situation after the density of positive vessels and medication.
Embodiment
Embodiment one：The preparation of immunofluorescence detection agent box of the present invention
The present invention is from CD34 antigens as the overall vascular endothelial cell mark in tumor tissues, CXCR4 antigen conducts
Blood vessel endothelium apical cell marks, and marks the double positive blood vessels of CXCR4 and CD34 antigens using the double dyeing techniques of immunofluorescence
Endothelial cell.
Table 1：The source of antibody and specific experiment condition
| Index | Antibody sources | Restorative procedure | Antibody thinner ratio |
| CD34 | Zhong Shan Golden Bridge, article No.：ZM-0046 | Microwave 20min, 10mM CB (pH6.0) | 1:200 |
| CXCR4 | Abcam, article No. ab124824 | Microwave 20min, 10mM CB (pH6.0) | 1:100 |
CB：Citrate buffer solution.
Immunofluorescence detection agent box of the present invention is determined by following experiment：
First, two kinds of molecular marked compounds are detected in liver cancer patient paraffin section tumor region using immunofluorescence method
Expression.
Then, the picture 5 for cancer nests region being obtained under the high power field (200 times) of fluorescence microscope is opened.Schemed using specialty
As double positive signals of software analysis CXCR4 and CD34 molecule account for the ratio of single positive signal of CD34 molecules.Final result is 5
The average value of individual high power field.
Finally, the packet of " more " and " few " are carried out to gained ratio using median method.
The immunofluorescence detection agent box phase of the preparation method of immunofluorescence detection agent box of the present invention and routine
Together, difference is that the protein molecular that the first antibody that the kit uses is detected is respectively：CD34 and CXCR4.Both
One antibody can detect clearly positive signal in liver cancer tissue paraffin section.
Embodiment two：The test experience of immunofluorescence detection agent box of the present invention
For immunofluorescence detection agent box of the present invention, its validity is detected by following steps.
(1) Samples detection
1. choosing the liver cancer paraffin section (before taking Sorafenib treatment) comprising tumor region, and ensure without large stretch of bad
Extremely；
2. obtain 4 μm of paraffin section 8, in 60 DEG C of roasting pieces 2 hours, take out, it is slightly cold；
3. dewaxed 2 times, every time 10 minutes with dimethylbenzene at room temperature；
4. wash away dimethylbenzene in 100% ethanol, then successively by 95% alcohol, 80% alcohol, 70% alcohol, every time 5
Minute；
5. washed in distilled water 5 minutes；
6. use 0.3%H2O2Close endogenous peroxidase activity, room temperature 10 minutes；
7. washed in distilled water 4 times, every time 5 minutes；
8. antigen retrieval：10mM citrate buffer solutions (pH6.0), microwave thermal repair 20min；
9. room temperature natural cooling 30 minutes；
Washed in 10.PBS buffer solutions 4 times, every time 3 minutes；
It is 11. first antibody CD34 (being purchased from company of Zhong Shan Golden Bridge, article No. ZM-0046) and CXCR4 is (public purchased from Abcam
Department, article No. ab124824) it is added dropwise on tissue sections, 4 DEG C are incubated 12 hours；
12.PBS buffer solutions Xian 4 times, 5 minutes every time；
13. the secondary antibody of luciferase mark is added dropwise, 37 DEG C are incubated 30 minutes；It is donkey respectively including 2 kinds of secondary antibodies
Anti- mouse IgG green fluorescent label antibody (being purchased from ThermoFisher companies, article No. A-21202) and donkey anti-rabbit IgG's is red
Color fluorescein labelled antibody (is purchased from ThermoFisher companies, article No. A-31572).
14.PBS is washed 4 times, every time 5 minutes；
15. nucleus dyestuff is added dropwise；
16. anti-quencher mounting.
(2) graphical analysis
1. the picture 5 that cancer nests region is obtained under the high power field (200 times) of fluorescence microscope is opened；
2. identify CD34 (green) and CXCR4 (red) single positive signal and both double sun respectively using analysis software
Property (yellow) signal (such as Fig. 1)；
3. analyze the double positive signal areas of CXCR4 and CD34, and the mono- positive signal areas of CD34, calculate CXCR4 and
The double positive signal areas of CD34 account for the ratios of the mono- positive signal areas of CD34, and (i.e. CXCR4 positive vessels account for unit vessel area
Ratio, abbreviation CXCR4 positive vessels ratio)；
4. final result is the average value of 5 high power fields.
(3) interpretation of result
1. by CXCR4 positive vessels ratio according to median be divided at high proportion with low two kinds of ratio.When CXCR4 positive vessels
When ratio is higher than 10%, group (being designated as " CXCR4 is high ") at high proportion is divided into；When CXCR4 positive vessels ratio is less than 10%, division
(it is designated as " CXCR4 is low ") for low ratio group.
2. judge patient outcomes：When patient's CXCR4 positive vessels ratio is CXCR4 high, it is believed that patient receives rope
The effect of after La Feini treatments, is preferable, it is proposed that receives Sorafenib treatment；When patient's CXCR4 positive vessels ratio is that CXCR4 is low
When, it is believed that the effect of patient receives after Sorafenib is treated is poor, it is not recommended that receives Sorafenib treatment.
3. survival analysis result shows, two groups of patients (CXCR4 positive vessels ratio is high low with CXCR4 positive vessels ratios)
There were significant differences (P=0.028, such as Fig. 2) for overall survival after Sorafenib treatment is taken.
Claims (2)
1. a kind of immunofluorescence detection agent box for being used to assess liver cancer patient therapeutic effect, including first antibody and second resist
Body, it is characterised in that：The first antibody is the combination for being selected from the antibody for following protein molecular label：CD34 (cells
Differentiation antigen 34) and CXCR4 (C-X-C Chemokine receptor4s).
2. immunofluorescence detection agent box according to claim 1, it is characterised in that：The secondary antibody is with fluorescence
The secondary antibody of plain enzyme mark.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710333555.1A CN107741497A (en) | 2017-05-12 | 2017-05-12 | For assessing the immunofluorescence detection agent box of liver cancer patient therapeutic effect |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710333555.1A CN107741497A (en) | 2017-05-12 | 2017-05-12 | For assessing the immunofluorescence detection agent box of liver cancer patient therapeutic effect |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN107741497A true CN107741497A (en) | 2018-02-27 |
Family
ID=61235151
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201710333555.1A Pending CN107741497A (en) | 2017-05-12 | 2017-05-12 | For assessing the immunofluorescence detection agent box of liver cancer patient therapeutic effect |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN107741497A (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109633155A (en) * | 2018-12-12 | 2019-04-16 | 中山大学 | Application of the ang2 as the indication molecule of Sorafenib treatment liver cancer efficacy |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101107011A (en) * | 2004-11-19 | 2008-01-16 | 康乃尔研究基金会有限公司 | Use of vascular endothelial growth factor receptor 1+ cells in treating and monitoring cancer and in screening for chemotherapeutics |
| WO2012047339A2 (en) * | 2010-06-28 | 2012-04-12 | The General Hospital Corporation | Anti-cxcr4 as a sensitizer to cancer therapeutics |
| US20140235495A1 (en) * | 2011-09-23 | 2014-08-21 | Siemens Healthcare Diagnostics Inc. | Cell response assay for cancer and methods of producing and using same |
| US9415011B1 (en) * | 2015-04-08 | 2016-08-16 | National Tsing Hua University | Method for treatment of liver cancer and inhibition of metastasis with CXC-chemokine-receptor 4-targeted nanoparticle |
| WO2016164359A1 (en) * | 2015-04-10 | 2016-10-13 | Tumorgen Mdx Llc | Rare cell isolation device and method of use thereof |
-
2017
- 2017-05-12 CN CN201710333555.1A patent/CN107741497A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101107011A (en) * | 2004-11-19 | 2008-01-16 | 康乃尔研究基金会有限公司 | Use of vascular endothelial growth factor receptor 1+ cells in treating and monitoring cancer and in screening for chemotherapeutics |
| WO2012047339A2 (en) * | 2010-06-28 | 2012-04-12 | The General Hospital Corporation | Anti-cxcr4 as a sensitizer to cancer therapeutics |
| US20140235495A1 (en) * | 2011-09-23 | 2014-08-21 | Siemens Healthcare Diagnostics Inc. | Cell response assay for cancer and methods of producing and using same |
| US9415011B1 (en) * | 2015-04-08 | 2016-08-16 | National Tsing Hua University | Method for treatment of liver cancer and inhibition of metastasis with CXC-chemokine-receptor 4-targeted nanoparticle |
| WO2016164359A1 (en) * | 2015-04-10 | 2016-10-13 | Tumorgen Mdx Llc | Rare cell isolation device and method of use thereof |
Non-Patent Citations (5)
| Title |
|---|
| LIEPELT,A等: "Stromal cell-derived factor-1 (SDF-1) as a target in liver diseases", 《AMERICAN JOURNAL OF PHYSIOLOGY-GASTROINTESTINAL AND LIVER PHYSIOLOGY》 * |
| MA,X等: "Predictive Immunohistochemical Markers Related to Drug Selection for Patients Treated with Sunitinib or Sorafenib for Metastatic Renal Cell Cancer", 《SCIENTIFIC REPORTS》 * |
| RATAJCZAK,MZ等: "Induction of a tumor-metastasis-receptive microenvironment as an unwanted and underestimated side effect of treatment by chemotherapy or radiotherapy", 《JOURNAL OF OVARIAN RESEARCH》 * |
| 孙甲甲等: "CXCR4在原发性肝细胞癌中的表达及意义", 《中国肿瘤临床》 * |
| 陆萌英等: "肝脏微血管密度在索拉非尼联合冷冻消融治疗进展期肝癌疗效评价中的意义", 《第二军医大学学报》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109633155A (en) * | 2018-12-12 | 2019-04-16 | 中山大学 | Application of the ang2 as the indication molecule of Sorafenib treatment liver cancer efficacy |
| CN109633155B (en) * | 2018-12-12 | 2022-04-01 | 中山大学 | application of ang2 as indicator molecule for liver cancer treatment effect of sorafenib |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Gabr et al. | Liver transplantation following Yttrium‐90 radioembolization: 15‐year experience in 207‐patient cohort | |
| Woodward et al. | Cancer stem cells: markers or biomarkers? | |
| Saunders et al. | Breast cancer | |
| Jeong et al. | Ischemia induces autophagy of endothelial cells and stimulates angiogenic effects in a hindlimb ischemia mouse model | |
| Dehcordi et al. | Stemness marker detection in the periphery of glioblastoma and ability of glioblastoma to generate glioma stem cells: clinical correlations | |
| Lin et al. | Clinicopathological features of salivary and non-salivary adenoid cystic carcinomas | |
| Filby et al. | Cellular origins of endometriosis: towards novel diagnostics and therapeutics | |
| CN107603946A (en) | Astragalus polyose produces the application during bystander cell damages to BMSCs in protection radiation A549 cells | |
| Horn et al. | Outcome of heavily pretreated recurrent oral squamous cell carcinoma after salvage resection: A monocentric retrospective analysis | |
| Zil-E-Ali et al. | Diagnosing necrotizing fasciitis using procalcitonin and a laboratory risk indicator: brief overview | |
| CN108398556A (en) | Double antibody for Post hepatectomy of liver cancer prognosis combines fluorescence detection reagent kit | |
| CN107741497A (en) | For assessing the immunofluorescence detection agent box of liver cancer patient therapeutic effect | |
| CN105560255B (en) | Application and its medicine of the obakulactone in treatment pulmonary fibrosis, chronic pneumonia, chronic respiratory disease medicine is prepared | |
| CN105158468B (en) | CK19 associating OV6 application in preparation hepatocarcinoma molecule parting test kit and hepatocarcinoma individualized treatment | |
| CN109793749A (en) | MiR-145-3p is preparing the application in Apoptosis and autophagy reinforcing agent | |
| CN103243163A (en) | Novel applications of miR-27a (microRNA-27a) | |
| Mavrikakis et al. | The role of thrombosis as a mechanism of exacerbation in venous and combined venous lymphatic vascular malformations of the orbit | |
| Wang et al. | Correlations between claudin-1 and PIGF expressions in retinoblastoma. | |
| CN103417629B (en) | Traditional Chinese medicine for treating hepatitis B liver cirrhosis | |
| Wu et al. | Yang-Gan-Jiang-Mei formula alleviates non-alcoholic steatohepatitis by inhibiting NLRP3 inflammasome through mitophagy | |
| Tewari et al. | Estrogen and progesterone receptor status in breast cancer: effect of oral contraceptive pills and hormone replacement therapy | |
| Zhou et al. | Research hotspot and frontier progress of cancer under the background of precision medicine | |
| Zeng et al. | Expressions of orphan nuclear receptor TR3/Nur77 in chronic hepatopathy and its clinical significance | |
| CN102297971A (en) | Kit for detecting expression of JWA in cancer or malignant tumor tissue | |
| Fležar et al. | Radiosensitivity of squamous cell carcinoma metastases to the neck assessed by immunocytochemical profiling of fine-needle aspiration biopsy cell specimens: a pilot study |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| WD01 | Invention patent application deemed withdrawn after publication | ||
| WD01 | Invention patent application deemed withdrawn after publication |
Application publication date: 20180227 |