CN108414301A - It is a kind of to detect marrow or the pre-treating method of peripheral blood sample for flow cyctometry - Google Patents
It is a kind of to detect marrow or the pre-treating method of peripheral blood sample for flow cyctometry Download PDFInfo
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- CN108414301A CN108414301A CN201711049032.0A CN201711049032A CN108414301A CN 108414301 A CN108414301 A CN 108414301A CN 201711049032 A CN201711049032 A CN 201711049032A CN 108414301 A CN108414301 A CN 108414301A
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- Prior art keywords
- peripheral blood
- blood sample
- treating method
- marrow
- flow
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Classifications
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume, or surface-area of porous materials
- G01N15/10—Investigating individual particles
- G01N15/14—Electro-optical investigation, e.g. flow cytometers
Abstract
The present invention provides a kind of marrow or the pre-treating method of peripheral blood sample are detected for flow cyctometry, technical solution improvement part step on the basis of traditional manipulative steps, fragment generation can be greatly reduced, analysis result can be made more accurate, ensure the accurate of diagnostic result.From the point of view of specific, the present invention maintains 37 DEG C using incubator, and haemolysis, can be optimal effect with this condition；In addition, the present invention is centrifugal force using centrifugal speed unit, ensure different size, the centrifuge of size generates identical centrifugation dynamics, and uses specific 190g centrifugal force, can utmostly remove the fragment that haemolysis generates at 37 DEG C.To significantly reduce the shive content in product to be measured, lay a good foundation for the accuracy of Flow cytometry.The present invention realizes good technique effect with innovative technological improvement, while cost is relatively low, is easily achieved, therefore has promotion prospect outstanding.
Description
Technical field
The present invention relates to biological experiment technical fields, are improved further to the operating method of flow cytometry, specifically
It is related to a kind of pre-treating method being used for flow cyctometry detection marrow or peripheral blood sample.
Background technology
Flow cytometry neoplastic hematologic disorder has been widely used, and pre-processing belongs to critically important in quality control
Link, early period, operation processing was improper, and fragment is excessive when can cause data analysis, false positive occurs, causes mistaken diagnosis.Conventional process side
For method due to methodology disadvantage, generation amount of debris is larger, and in certain samples being not in good state, amount of debris is even larger than detection
Cell itself seriously affects data analysis.The analysis of sample process quality flow cytometric art is most important, and haemolysis, splits at room temperature
It is insufficient to solve red blood cell, generates the inhomogenous fragment of a large amount of sizes, between 25 to 50 degrees Celsius, hemolyzing effect difference is very big,
Temperature is too low, and haemolysis is insufficient, and later stage fragment is difficult to effectively remove, and temperature is excessively high, can crack karyocyte, seriously affects experiment
As a result.
Invention content
The present invention is directed to the technological deficiencies for the prior art, provide a kind of for flow cyctometry detection marrow or periphery
The pre-treating method of blood sample, to solve in the prior art when using Flow cytometry hematological system tumor, to handle bone
The technical issues of marrow and peripheral blood sample are also easy to produce a large amount of fragments and impact analysis.
To realize that the above technical purpose, the present invention use following technical scheme：
It is a kind of to detect marrow or the pre-treating method of peripheral blood sample for flow cyctometry, include the following steps：
1) it by sample and antibody mixing, is protected from light is incubated 15min at room temperature；
2) 1mL hemolysins are added into mixture obtained by step 1), mixing is protected from light under the conditions of 37 DEG C and is incubated 15min；
3) 3mL PBS buffer solution is added into mixture obtained by step 2), is separated by solid-liquid separation after mixing and takes precipitation；
4) 3mL PBS buffer solution is added into solid content obtained by step 3), is separated by solid-liquid separation after mixing and takes precipitation；
5) solid content obtained by step 4) is resuspended to get to product to be measured with 300 μ L PBS buffer solution.
Preferably, it is further comprising the steps of 6)：Utilize the product to be measured obtained by flow cytomery step 5).
Preferably, mixing described in step 1) and step 2), is completed using turbula shaker.
Preferably, the step 1) room temperature is 25 DEG C.
Preferably, the step 2) hemolysin is the commercially available hemolysin produced by coulter companies.
Preferably, step 3) and the separation of solid and liquid described in step 4), are centrifugation.
Preferably, the centrifugation, is the centrifugal force with 190g, centrifugal treating 5min.
The present invention provides a kind of marrow or the pre-treating method of peripheral blood sample, the technology are detected for flow cyctometry
Scheme improvement part step on the basis of traditional manipulative steps can greatly reduce fragment generation, can make analysis result more
Accurately, ensure the accurate of diagnostic result.From the point of view of specific, the present invention maintains 37 DEG C using incubator, with this condition haemolysis, Ke Yida
To optimal effectiveness；In addition, the present invention is centrifugal force using centrifugal speed unit, ensure that different size, the centrifuge of size generate
Identical centrifugation dynamics, and specific 190g centrifugal force is used, it can utmostly remove the fragment that haemolysis generates at 37 DEG C.
To significantly reduce the shive content in product to be measured, lay a good foundation for the accuracy of Flow cytometry.The present invention with
Innovative technological improvement realizes good technique effect, while cost is relatively low, is easily achieved, therefore has popularization outstanding
Foreground.
Specific implementation mode
The specific implementation mode of the present invention will be described in detail below.In order to avoid excessive unnecessary details,
It will not be described in detail in following embodiment to belonging to well known structure or function.Approximation used in following embodiment
Language can be used for quantitative expression, show to allow quantity to have certain variation in the case where not changing basic function.It is fixed except having
Adopted outer, technical and scientific term used in following embodiment has the phase being commonly understood by with those skilled in the art of the invention
Same meaning.
Embodiment 1
It is a kind of to detect marrow or the pre-treating method of peripheral blood sample for flow cyctometry, include the following steps：
1) it by sample and antibody mixing, is protected from light is incubated 15min at room temperature；
2) 1mL hemolysins are added into mixture obtained by step 1), mixing is protected from light under the conditions of 37 DEG C and is incubated 15min；
3) 3mL PBS buffer solution is added into mixture obtained by step 2), is separated by solid-liquid separation after mixing and takes precipitation；
4) 3mL PBS buffer solution is added into solid content obtained by step 3), is separated by solid-liquid separation after mixing and takes precipitation；
5) solid content obtained by step 4) is resuspended to get to product to be measured with 300 μ L PBS buffer solution.
On the basis of above technical scheme, meet the following conditions：
It is further comprising the steps of 6)：Utilize the product to be measured obtained by flow cytomery step 5).
Mixing described in step 1) and step 2), is completed using turbula shaker.
Step 1) the room temperature is 25 DEG C.
Step 2) the hemolysin is the commercially available hemolysin produced by coulter companies.
Separation of solid and liquid described in step 3) and step 4) is centrifugation.
The centrifugation is the centrifugal force with 190g, centrifugal treating 5min.
Embodiment 2
It is a kind of to detect marrow or the pre-treating method of peripheral blood sample for flow cyctometry, include the following steps：
1) it by sample and antibody mixing, is protected from light is incubated 15min at room temperature；
2) 1mL hemolysins are added into mixture obtained by step 1), mixing is protected from light under the conditions of 37 DEG C and is incubated 15min；
3) 3mL PBS buffer solution is added into mixture obtained by step 2), is separated by solid-liquid separation after mixing and takes precipitation；
4) 3mL PBS buffer solution is added into solid content obtained by step 3), is separated by solid-liquid separation after mixing and takes precipitation；
5) solid content obtained by step 4) is resuspended to get to product to be measured with 300 μ L PBS buffer solution.
The embodiments of the present invention have been described in detail above, but content is only the preferred embodiment of the present invention,
It is not intended to limit the invention.All all any modification, equivalent and improvement etc. done in the application range of the present invention, should all
It is included within protection scope of the present invention.
Claims (7)
1. a kind of detecting marrow or the pre-treating method of peripheral blood sample for flow cyctometry, it is characterised in that including following step
Suddenly：
1) it by sample and antibody mixing, is protected from light is incubated 15min at room temperature；
2) 1mL hemolysins are added into mixture obtained by step 1), mixing is protected from light under the conditions of 37 DEG C and is incubated 15min；
3) 3mL PBS buffer solution is added into mixture obtained by step 2), is separated by solid-liquid separation after mixing and takes precipitation；
4) 3mL PBS buffer solution is added into solid content obtained by step 3), is separated by solid-liquid separation after mixing and takes precipitation；
5) solid content obtained by step 4) is resuspended to get to product to be measured with 300 μ L PBS buffer solution.
2. a kind of pre-treating method being used for flow cyctometry detection marrow or peripheral blood sample according to claim 1,
Characterized by further comprising following steps 6)：Utilize the product to be measured obtained by flow cytomery step 5).
3. a kind of pre-treating method being used for flow cyctometry detection marrow or peripheral blood sample according to claim 1,
It is characterized in that mixing described in step 1) and step 2), is completed using turbula shaker.
4. a kind of pre-treating method being used for flow cyctometry detection marrow or peripheral blood sample according to claim 1,
It is characterized in that the step 1) room temperature is 25 DEG C.
5. a kind of pre-treating method being used for flow cyctometry detection marrow or peripheral blood sample according to claim 1,
It is characterized in that the step 2) hemolysin is the commercially available hemolysin produced by coulter companies.
6. a kind of pre-treating method being used for flow cyctometry detection marrow or peripheral blood sample according to claim 1,
It is characterized in that step 3) and the separation of solid and liquid described in step 4), are centrifugation.
7. a kind of pre-treating method being used for flow cyctometry detection marrow or peripheral blood sample according to claim 6,
It is characterized in that the centrifugation, is the centrifugal force with 190g, centrifugal treating 5min.
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|---|---|---|---|
| CN201711049032.0A CN108414301A (en) | 2017-10-31 | 2017-10-31 | It is a kind of to detect marrow or the pre-treating method of peripheral blood sample for flow cyctometry |
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| CN201711049032.0A CN108414301A (en) | 2017-10-31 | 2017-10-31 | It is a kind of to detect marrow or the pre-treating method of peripheral blood sample for flow cyctometry |
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Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2004201574A (en) * | 2002-12-25 | 2004-07-22 | Dai Ichi Seiyaku Co Ltd | Hematopoietic stem cell activation marker |
| CN101327324A (en) * | 2007-06-21 | 2008-12-24 | 中国科学院上海生命科学研究院 | Method and composition for treating autoimmunity disease |
| CN101620220A (en) * | 2008-06-30 | 2010-01-06 | 张晖 | Cytoperm capable of being used as hemolytic agent and using method thereof |
| CN103217374A (en) * | 2013-02-20 | 2013-07-24 | 北京大学人民医院 | Acute B lymphocytic leukemia initiating cell characteristic determination system and method |
| CN105223361A (en) * | 2015-08-28 | 2016-01-06 | 北京大学人民医院 | A kind of detect Pancytopenia Naive T cells kit, application and method |
| CN105242046A (en) * | 2015-09-17 | 2016-01-13 | 中国人民解放军第三军医大学第三附属医院 | Detection method for tuberculosis infection Treg/Th17 cell |
| CN107233296A (en) * | 2017-05-27 | 2017-10-10 | 中山大学 | Thymopeptide-5 solubility micropin and preparation method thereof |
-
2017
- 2017-10-31 CN CN201711049032.0A patent/CN108414301A/en active Pending
Patent Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2004201574A (en) * | 2002-12-25 | 2004-07-22 | Dai Ichi Seiyaku Co Ltd | Hematopoietic stem cell activation marker |
| CN101327324A (en) * | 2007-06-21 | 2008-12-24 | 中国科学院上海生命科学研究院 | Method and composition for treating autoimmunity disease |
| CN101620220A (en) * | 2008-06-30 | 2010-01-06 | 张晖 | Cytoperm capable of being used as hemolytic agent and using method thereof |
| CN103217374A (en) * | 2013-02-20 | 2013-07-24 | 北京大学人民医院 | Acute B lymphocytic leukemia initiating cell characteristic determination system and method |
| CN105223361A (en) * | 2015-08-28 | 2016-01-06 | 北京大学人民医院 | A kind of detect Pancytopenia Naive T cells kit, application and method |
| CN105242046A (en) * | 2015-09-17 | 2016-01-13 | 中国人民解放军第三军医大学第三附属医院 | Detection method for tuberculosis infection Treg/Th17 cell |
| CN107233296A (en) * | 2017-05-27 | 2017-10-10 | 中山大学 | Thymopeptide-5 solubility micropin and preparation method thereof |
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