CN103308669A - Streaming blood cell analyzer channel for automatic immunofluorescence detection and use method thereof - Google Patents
Streaming blood cell analyzer channel for automatic immunofluorescence detection and use method thereof Download PDFInfo
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- CN103308669A CN103308669A CN2013101673485A CN201310167348A CN103308669A CN 103308669 A CN103308669 A CN 103308669A CN 2013101673485 A CN2013101673485 A CN 2013101673485A CN 201310167348 A CN201310167348 A CN 201310167348A CN 103308669 A CN103308669 A CN 103308669A
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- 238000001514 detection method Methods 0.000 title claims abstract description 33
- 238000000034 method Methods 0.000 title claims abstract description 18
- 238000010166 immunofluorescence Methods 0.000 title abstract description 5
- 210000000601 blood cell Anatomy 0.000 title abstract 5
- 238000005406 washing Methods 0.000 claims abstract description 39
- 206010018910 Haemolysis Diseases 0.000 claims abstract description 25
- 230000008588 hemolysis Effects 0.000 claims abstract description 25
- 238000010790 dilution Methods 0.000 claims abstract description 23
- 239000012895 dilution Substances 0.000 claims abstract description 23
- 210000004369 blood Anatomy 0.000 claims abstract description 14
- 239000008280 blood Substances 0.000 claims abstract description 14
- 210000005259 peripheral blood Anatomy 0.000 claims abstract description 12
- 239000011886 peripheral blood Substances 0.000 claims abstract description 9
- 210000004027 cell Anatomy 0.000 claims abstract description 7
- 238000011534 incubation Methods 0.000 claims abstract description 7
- 239000000523 sample Substances 0.000 claims description 71
- 239000006228 supernatant Substances 0.000 claims description 23
- 239000007788 liquid Substances 0.000 claims description 20
- 230000036039 immunity Effects 0.000 claims description 17
- 238000002156 mixing Methods 0.000 claims description 14
- 238000007710 freezing Methods 0.000 claims description 12
- 206010059484 Haemodilution Diseases 0.000 claims description 9
- 239000000945 filler Substances 0.000 claims description 9
- 238000000926 separation method Methods 0.000 claims description 9
- 238000007619 statistical method Methods 0.000 claims description 9
- 238000003805 vibration mixing Methods 0.000 claims description 9
- 230000033001 locomotion Effects 0.000 claims description 7
- GNBHRKFJIUUOQI-UHFFFAOYSA-N fluorescein Chemical compound O1C(=O)C2=CC=CC=C2C21C1=CC=C(O)C=C1OC1=CC(O)=CC=C21 GNBHRKFJIUUOQI-UHFFFAOYSA-N 0.000 claims description 6
- 238000009534 blood test Methods 0.000 claims description 5
- 230000010355 oscillation Effects 0.000 claims description 4
- 241000931526 Acer campestre Species 0.000 claims description 3
- 239000003146 anticoagulant agent Substances 0.000 claims description 3
- 229940127219 anticoagulant drug Drugs 0.000 claims description 3
- 239000003153 chemical reaction reagent Substances 0.000 claims description 3
- 238000004140 cleaning Methods 0.000 claims description 3
- 230000000694 effects Effects 0.000 claims description 3
- 230000003628 erosive effect Effects 0.000 claims description 3
- 210000003743 erythrocyte Anatomy 0.000 claims description 3
- MHMNJMPURVTYEJ-UHFFFAOYSA-N fluorescein-5-isothiocyanate Chemical compound O1C(=O)C2=CC(N=C=S)=CC=C2C21C1=CC=C(O)C=C1OC1=CC(O)=CC=C21 MHMNJMPURVTYEJ-UHFFFAOYSA-N 0.000 claims description 3
- 239000007850 fluorescent dye Substances 0.000 claims description 3
- 239000003219 hemolytic agent Substances 0.000 claims description 3
- 238000003018 immunoassay Methods 0.000 claims description 3
- 230000003287 optical effect Effects 0.000 claims description 3
- 238000001556 precipitation Methods 0.000 claims description 3
- 238000000197 pyrolysis Methods 0.000 claims description 3
- 230000012447 hatching Effects 0.000 claims description 2
- 210000003958 hematopoietic stem cell Anatomy 0.000 abstract description 3
- 210000000130 stem cell Anatomy 0.000 abstract description 2
- 238000001917 fluorescence detection Methods 0.000 description 4
- 230000007812 deficiency Effects 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 238000000684 flow cytometry Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 239000002244 precipitate Substances 0.000 description 1
- 238000004904 shortening Methods 0.000 description 1
Images
Abstract
The invention provides a streaming blood cell analyzer channel for automatic immunofluorescence detection. The streaming blood cell analyzer channel comprises a valve a, a blood dilution module, a valve b, a hemolysis module, a valve c, a centrifugal washing module, a valve e, an automatic incubation module, a valve d, a valve f and a streaming detection module. The invention further provides a corresponding use method. By using the streaming blood cell analyzer channel and the use method thereof, automatic joint detection of blood routine and specific cells such as peripheral blood stem cells and the like can be realized, the cost for detecting hematopoietic stem cells is reduced, the real-time performance of detection is improved, and the detection function of a blood cell analyzer is extended.
Description
Technical field
The present invention relates to SABC, optics, mechano-electronic, statistical mathematics, specifically a kind of active immunity fluorescence detection channel and using method thereof for the streaming Hematometer.
Background technology
In the market five the classification Hematometers, except adopting traditional electrical impedance method and radio frequency method, progressively use Flow Cytometry such as Sysmex XE-5000, and adopted immunofluorescence technique, progressively realized the detection to some specific cell, such as peripheral blood hematopoietic stem cells.But present immunofluorescence detects and unrealized robotization detects, but manually dilutes, haemolysis, hatches and the repeatedly work such as centrifuge washing, and process is time-consuming and operate heavy.Therefore, realize that the robotization of immunofluorescence cell detects, to shortening detection time, promote clinical practice, significant.
Summary of the invention
The present invention is directed to the demand and the deficiencies in the prior art, a kind of active immunity fluorescence detection channel and using method thereof of streaming Hematometer is provided.
According to an aspect of the present invention, a kind of streaming Hematometer passage for the active immunity fluoroscopic examination is provided, comprise the valve a, hemodilution module, valve b, haemolysis module, valve c, the centrifuge washing module that connect successively, also comprise valve e, automatic raising module, valve d, valve f, flow cytometer detection module, wherein:
The centrifuge washing module comprises centrifugal oscillating table, resuspended liquid filling syringe needle, supernatant draws syringe needle, the supernatant drawing device, resuspended liquid charging point, sample draws syringe needle, the supernatant drawing device draws syringe needle by supernatant and connects centrifugal oscillating table, resuspended liquid charging point connects centrifugal oscillating table by resuspended liquid filling syringe needle, centrifugal oscillating table connects the end that sample draws syringe needle, sample draws the other end of syringe needle successively by valve e, automatic raising module, valve d connects back centrifugal oscillating table, and the other end that sample draws syringe needle is connected to the flow cytometer detection module by valve f;
The hemodilution module is used for blood sample is detected front anti-freezing and dilution;
The haemolysis module has the erythrocyte cracked liquid filler, and includes the vibration evenly mixing device;
The centrifuge washing module is used for sample is washed purifying;
Automatically raising module has the antibody filler, adopts the hanging drop mode antibody of annotating, and includes vibration mixing and attemperating unit;
The flow cytometer detection module has the color filter group, for the switching that filters for different fluorescent reagents, so that final sample is detected.
Preferably, also comprise control module and statistical analysis module, wherein, control module is used for valve a, hemodilution module, valve b, haemolysis module, valve c, centrifuge washing module, valve e, the running of raising module, valve d, valve f, flow cytometer detection module is controlled and detected automatically, statistical analysis module is processed for the photosignal that the convection type detection module collects, and obtains density, the volumetric parameter of fluorescence immunoassay cell.
Preferably, the centrifuge washing module can be turned round centrifugal motion and the motion of micro oscillation mixing.
Preferably, centrifugal oscillating table comprises the eccentric vibrating wheel and separates resuspended pipe, wherein, separates resuspended pipe and is fixed on the eccentric vibrating wheel, produces vibration by the rotation of eccentric vibrating wheel, guarantees resuspended effect.
Preferably, the bottom of the resuspended pipe of separation of centrifugal oscillating table is provided with groove, to take sample precipitation in, guarantees that supernatant draws and does not lose sample in the process.
Preferably, the resuspended pipe of the separation of centrifugal oscillating table and resuspended liquid filler, supernatant draw syringe needle, sample and draw syringe needle and carry out station according to demand and align, and guarantee separation, washing, resuspended carrying out.
According to another aspect of the present invention, also provide a kind of using method of described streaming Hematometer passage for the active immunity fluoroscopic examination, comprise the steps:
Step 4, sample draws syringe needle and draws sample, sends into automatic raising module by valve e, and the haemolysis sample of washing purifying carries out the filling of sample fluorescein and automatically hatches;
Step 6, sample are drawn and are hatched sample after syringe needle draws washing, send into the flow cytometer detection module by valve f, and the flow cytometer detection module detects final sample;
Step 7, statistical analysis module is integrated recognition result and the peripheral blood routine blood test result that PMT obtains.
Step 8 cleans backward erosion some times to the liquid road in the whole sense channel.
Preferably, in step 1, the peripheral blood extension rate is 10-100 times.
Preferably, in step 4, incubation temperature is 30-40 degree centigrade, and incubation time is 10-15 minute, and fluorescein is FITC.
Preferably, in step 6, the optical maser wavelength that the flow cytometer detection module adopts is 488nm, and the fluorescent probe that the flow cytometer detection module comprises is PMT.
Compared with prior art, advantage of the present invention is, can realize routine blood test and specific cell, automatic combined detection such as peripheral blood hematopoietic stem cells etc., reduce the expense that candidate stem cell detects, increased the real-time that detects, expanded the measuring ability of Hematometer.
Description of drawings
By reading the detailed description of non-limiting example being done with reference to the following drawings, it is more obvious that other features, objects and advantages of the present invention will become:
Fig. 1 is structure principle chart of the present invention;
Fig. 2 is that the sample that separates resuspended pipe is taken groove design in.
Embodiment
The present invention is described in detail below in conjunction with specific embodiment.Following examples will help those skilled in the art further to understand the present invention, but not limit in any form the present invention.Should be pointed out that to those skilled in the art, without departing from the inventive concept of the premise, can also make some distortion and improvement.These all belong to protection scope of the present invention.
Referring to Fig. 1 and Fig. 2,
The present invention comprises following technical scheme: a kind of active immunity fluorescence detection channel of streaming Hematometer comprises anti-freezing dilution module, haemolysis module, automatic raising module, centrifuge washing module, flow cytometer detection module, control module, statistical analysis module.The routine blood test dilution system of described anti-freezing dilution module and streaming Hematometer shares, and by a valve a UNICOM, described active immunity fluorescence detection channel is carried out switch control, and its major function is anti-freezing and the dilution before sample is detected.Described haemolysis module has the erythrocyte cracked liquid filler, and the vibration evenly mixing device is arranged.Described automatic raising module has the antibody filler, adopts the hanging drop mode antibody of annotating, and vibration mixing and attemperating unit are arranged.Described centrifuge washing module has the vibration centrifugal chamber and separates resuspended pipe, and is furnished with that locating device, resuspended liquid filling syringe needle, supernatant draw syringe needle, sample draws syringe needle, can wash purifying to sample.Described flow cytometer detection module has the color filter group, and switching can filter for different fluorescent reagents.The mode of operation of described control module convection type Hematometer is selected, and to each channel switch of Hematometer, the running of submodule is controlled and detected.The photosignal that described statistical analysis module convection type detection module collects is processed at host computer, the parameters such as the density of acquisition fluorescence immunoassay cell, volume.
Further, described haemolysis module, described automatic raising module by valve b and valve c and described centrifugal module UNICOM, can as required, enter respectively the centrifuge washing module and wash purifying with assurance haemolysis sample and fluorescence calibration sample respectively.
Further, described centrifuge washing module can realize two kinds of motions, namely turns round the motion of centrifugal motion and micro oscillation mixing.
Further, described centrifuge washing module makes by locating device separates resuspended pipe and resuspended liquid filler, supernatant and draws syringe needle, sample and draw syringe needle and carry out station according to demand and align, assurance separation, washing, resuspended carrying out.
Further, the resuspended pipe of described separation has unique groove design, takes sample precipitation in, guarantees that supernatant draws not lose sample in the process.
Further, the design of the vibration evenly mixing device of described centrifuge washing module is that the resuspended pipe of described separation is fixed on the eccentric vibrating wheel, produces vibration by the eccentric wheel rotation, guarantees resuspended effect.
Further, described anti-freezing dilution module, the haemolysis module, automatic raising module, centrifuge washing module, flow cytometer detection module have independent Washing device, can carry out independent washing away.
Correspondingly, method provided by the invention comprises the steps:
Step 4, sample are drawn syringe needle 36 and are drawn sample, send into automatic raising module 4 by valve e, and the haemolysis sample of washing purifying carries out the filling of sample fluorescein and automatically hatches.Be primarily characterized in that incubation temperature is 30-40 degree centigrade, incubation time is 10-15 minute, and fluorescein is FITC in the present embodiment.
Step 6, sample are drawn and are hatched sample after syringe needle 36 draws washing, send into flow cytometer detection module 5 by valve f.5 pairs of final samples of flow cytometer detection module detect, and optical maser wavelength is 488nm in the present embodiment, and fluorescent probe is PMT.
Step 7, statistical analysis module is integrated recognition result and the peripheral blood routine blood test result that PMT obtains.
Step 8 cleans backward erosion some times to the liquid road in the whole sense channel.
Above specific embodiments of the invention are described.It will be appreciated that, the present invention is not limited to above-mentioned specific implementations, and those skilled in the art can make various distortion or modification within the scope of the claims, and this does not affect flesh and blood of the present invention.
Claims (10)
1. streaming Hematometer passage that is used for the active immunity fluoroscopic examination, it is characterized in that, comprise the valve a, hemodilution module, valve b, haemolysis module, valve c, the centrifuge washing module that connect successively, also comprise valve e, automatic raising module, valve d, valve f, flow cytometer detection module, wherein:
The centrifuge washing module comprises centrifugal oscillating table, resuspended liquid filling syringe needle, supernatant draws syringe needle, the supernatant drawing device, resuspended liquid charging point, sample draws syringe needle, the supernatant drawing device draws syringe needle by supernatant and connects centrifugal oscillating table, resuspended liquid charging point connects centrifugal oscillating table by resuspended liquid filling syringe needle, centrifugal oscillating table connects the end that sample draws syringe needle, sample draws the other end of syringe needle successively by valve e, automatic raising module, valve d connects back centrifugal oscillating table, and the other end that sample draws syringe needle is connected to the flow cytometer detection module by valve f;
The hemodilution module is used for blood sample is detected front anti-freezing and dilution;
The haemolysis module has the erythrocyte cracked liquid filler, and includes the vibration evenly mixing device;
The centrifuge washing module is used for sample is washed purifying;
Automatically raising module has the antibody filler, adopts the hanging drop mode antibody of annotating, and includes vibration mixing and attemperating unit;
The flow cytometer detection module has the color filter group, for the switching that filters for different fluorescent reagents, so that final sample is detected.
2. the streaming Hematometer passage for the active immunity fluoroscopic examination according to claim 1, it is characterized in that, also comprise control module and statistical analysis module, wherein, control module is used for valve a, hemodilution module, valve b, haemolysis module, valve c, centrifuge washing module, valve e, the running of raising module, valve d, valve f, flow cytometer detection module is controlled and detected automatically, statistical analysis module is processed for the photosignal that the convection type detection module collects, and obtains density, the volumetric parameter of fluorescence immunoassay cell.
3. the streaming Hematometer passage for the active immunity fluoroscopic examination according to claim 1 is characterized in that, the centrifuge washing module can be turned round centrifugal motion and the motion of micro oscillation mixing.
4. the streaming Hematometer passage for the active immunity fluoroscopic examination according to claim 1, it is characterized in that, centrifugal oscillating table comprises the eccentric vibrating wheel and separates resuspended pipe, wherein, separating resuspended pipe is fixed on the eccentric vibrating wheel, produce vibration by the rotation of eccentric vibrating wheel, guarantee resuspended effect.
5. the streaming Hematometer passage for the active immunity fluoroscopic examination according to claim 4 is characterized in that, the bottom of the resuspended pipe of separation of centrifugal oscillating table is provided with groove, to take sample precipitation in, guarantees that supernatant draws and does not lose sample in the process.
6. the streaming Hematometer passage for the active immunity fluoroscopic examination according to claim 4, it is characterized in that, the resuspended pipe of the separation of centrifugal oscillating table and resuspended liquid filler, supernatant draw syringe needle, sample and draw syringe needle and carry out station according to demand and align, and guarantee separation, washing, resuspended carrying out.
7. the using method of each described streaming Hematometer passage for the active immunity fluoroscopic examination in the claim 1 to 6 is characterized in that, comprises the steps:
Step 1 is opened valve a, and the peripheral blood blood sample enters the hemodilution module, to blood sample filling anti-coagulants, dilution and the mixing that vibrates, carries out anti-freezing and dilution;
Step 2 is opened valve b, and blood sample enters the haemolysis module, and sample filling hemolytic agent after the anti-freezing dilution is carried out haemolysis, automatically keeps some minutes behind the automatic mixing as pyrolysis time;
Step 3 is opened valve c, blood sample enter the centrifuge washing module to haemolysis after sample repeat that centrifuging, supernatant are drawn, resuspended dilution, vibration mixing step some times;
Step 4, sample draws syringe needle and draws sample, sends into automatic raising module by valve e, and the haemolysis sample of washing purifying carries out the filling of sample fluorescein and automatically hatches;
Step 5 is opened valve d, and the centrifuge washing module after finishing separate cleaning and washing away carries out to hatching rear sample that centrifuging, supernatant are drawn, resuspended dilution, vibration mixing step some times;
Step 6, sample are drawn and are hatched sample after syringe needle draws washing, send into the flow cytometer detection module by valve f, and the flow cytometer detection module detects final sample;
Step 7, statistical analysis module is integrated recognition result and the peripheral blood routine blood test result that PMT obtains.
Step 8 cleans backward erosion some times to the liquid road in the whole sense channel.
8. the using method of the streaming Hematometer passage for the active immunity fluoroscopic examination according to claim 7 is characterized in that, in step 1, the peripheral blood extension rate be 10-100 doubly.
9. the using method of the streaming Hematometer passage for the active immunity fluoroscopic examination according to claim 7 is characterized in that, in step 4, incubation temperature is 30-40 degree centigrade, and incubation time is 10-15 minute, and fluorescein is FITC.
10. the using method of the streaming Hematometer passage for the active immunity fluoroscopic examination according to claim 7 is characterized in that, in step 6, the optical maser wavelength that the flow cytometer detection module adopts is 488nm, and the fluorescent probe that the flow cytometer detection module comprises is PMT.
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| CN201310167348.5A CN103308669B (en) | 2013-05-08 | 2013-05-08 | Streaming blood cell analyzer channel for automatic immunofluorescence detection and use method thereof |
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| CN201310167348.5A CN103308669B (en) | 2013-05-08 | 2013-05-08 | Streaming blood cell analyzer channel for automatic immunofluorescence detection and use method thereof |
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| CN103308669A true CN103308669A (en) | 2013-09-18 |
| CN103308669B CN103308669B (en) | 2014-07-09 |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111493891A (en) * | 2020-04-02 | 2020-08-07 | 深圳市儿童医院 | Blood collection vibration detection equipment |
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| WO2002004921A2 (en) * | 2000-07-07 | 2002-01-17 | Coulter International Corp. | Apparatus and method for biological sample preparation and analysis |
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-
2013
- 2013-05-08 CN CN201310167348.5A patent/CN103308669B/en not_active Expired - Fee Related
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS6488251A (en) * | 1987-09-30 | 1989-04-03 | Canon Kk | Particle analyzer |
| WO2002004921A2 (en) * | 2000-07-07 | 2002-01-17 | Coulter International Corp. | Apparatus and method for biological sample preparation and analysis |
| CN101236149A (en) * | 2007-02-02 | 2008-08-06 | 深圳迈瑞生物医疗电子股份有限公司 | Stream type cell detection device and its accomplished stream type cell detection method |
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Non-Patent Citations (4)
| Title |
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| CHARLES GOOLSBY ET AL.: "Flow cytometric detection of herpes simplex virus type 2 infected and transformed cells by immunoenzymatic and by indirect immunofluorescence staining", 《CYTOMETRY》 * |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111493891A (en) * | 2020-04-02 | 2020-08-07 | 深圳市儿童医院 | Blood collection vibration detection equipment |
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