CN107034128A - A kind of mononuclearcell separating pipe and its application method - Google Patents
A kind of mononuclearcell separating pipe and its application method Download PDFInfo
- Publication number
- CN107034128A CN107034128A CN201710456928.4A CN201710456928A CN107034128A CN 107034128 A CN107034128 A CN 107034128A CN 201710456928 A CN201710456928 A CN 201710456928A CN 107034128 A CN107034128 A CN 107034128A
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- socket bore
- centrifuge tube
- pipe
- cone
- inclined side
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- 210000005087 mononuclear cell Anatomy 0.000 title claims abstract description 25
- 238000000034 method Methods 0.000 title claims abstract description 14
- 239000007788 liquid Substances 0.000 claims abstract description 25
- 238000000926 separation method Methods 0.000 claims abstract description 23
- 210000004027 cell Anatomy 0.000 claims abstract description 21
- 238000005119 centrifugation Methods 0.000 claims abstract description 21
- 230000010100 anticoagulation Effects 0.000 claims abstract description 13
- 210000004369 blood Anatomy 0.000 claims abstract description 11
- 239000008280 blood Substances 0.000 claims abstract description 11
- 238000005192 partition Methods 0.000 claims description 17
- 230000015572 biosynthetic process Effects 0.000 claims description 4
- 238000005406 washing Methods 0.000 claims description 4
- 239000003855 balanced salt solution Substances 0.000 claims description 3
- 210000000601 blood cell Anatomy 0.000 claims description 3
- 239000012930 cell culture fluid Substances 0.000 claims description 3
- 238000004140 cleaning Methods 0.000 claims description 3
- 238000007710 freezing Methods 0.000 claims description 3
- 238000010030 laminating Methods 0.000 claims description 2
- 238000005138 cryopreservation Methods 0.000 claims 1
- 238000010790 dilution Methods 0.000 claims 1
- 239000012895 dilution Substances 0.000 claims 1
- 230000003340 mental effect Effects 0.000 claims 1
- 210000003819 peripheral blood mononuclear cell Anatomy 0.000 abstract description 16
- 230000000694 effects Effects 0.000 abstract description 5
- 210000003743 erythrocyte Anatomy 0.000 abstract description 4
- 230000001194 anti-hemostatic effect Effects 0.000 abstract description 3
- 238000000432 density-gradient centrifugation Methods 0.000 abstract description 3
- 239000000203 mixture Substances 0.000 abstract description 3
- 238000001556 precipitation Methods 0.000 abstract description 3
- 239000010410 layer Substances 0.000 description 9
- 230000001413 cellular effect Effects 0.000 description 5
- 210000002381 plasma Anatomy 0.000 description 4
- 210000005259 peripheral blood Anatomy 0.000 description 3
- 206010028980 Neoplasm Diseases 0.000 description 2
- 230000023555 blood coagulation Effects 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 210000001616 monocyte Anatomy 0.000 description 2
- 239000011886 peripheral blood Substances 0.000 description 2
- 238000012545 processing Methods 0.000 description 2
- 239000004743 Polypropylene Substances 0.000 description 1
- 241000973497 Siphonognathus argyrophanes Species 0.000 description 1
- 238000003491 array Methods 0.000 description 1
- 230000001363 autoimmune Effects 0.000 description 1
- 210000001772 blood platelet Anatomy 0.000 description 1
- 230000010261 cell growth Effects 0.000 description 1
- 210000003855 cell nucleus Anatomy 0.000 description 1
- 238000002512 chemotherapy Methods 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 238000013461 design Methods 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 238000010494 dissociation reaction Methods 0.000 description 1
- 230000005593 dissociations Effects 0.000 description 1
- 238000011124 ex vivo culture Methods 0.000 description 1
- 210000003714 granulocyte Anatomy 0.000 description 1
- 238000009169 immunotherapy Methods 0.000 description 1
- 239000007791 liquid phase Substances 0.000 description 1
- 210000002751 lymph Anatomy 0.000 description 1
- 210000004698 lymphocyte Anatomy 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 239000012071 phase Substances 0.000 description 1
- -1 polypropylene Polymers 0.000 description 1
- 229920001155 polypropylene Polymers 0.000 description 1
- 239000011148 porous material Substances 0.000 description 1
- 230000005855 radiation Effects 0.000 description 1
- 239000002356 single layer Substances 0.000 description 1
- 230000001225 therapeutic effect Effects 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/06—Animal cells or tissues; Human cells or tissues
- C12N5/0602—Vertebrate cells
- C12N5/0634—Cells from the blood or the immune system
Abstract
The invention discloses mononuclearcell separating pipe, including centrifuge tube, lower half cross-section in inclined side leakage bucket dividing plate, centrifuge tube is provided with the mouth of pipe of centrifuge tube and is provided with cone socket bore plate；The half side tube wall of inclined side leakage bucket dividing plate and centrifuge tube forms uniform intervals on inclined side leakage bucket structure, cone socket bore plate and some positive cone socket bore and back taper socket bore is distributed with.The present invention directly can pour into anticoagulation through inclined side leakage bucket, it is ensured that anticoagulation flows down along centrifugation tube wall；Using cone nest AND DEWATERING FOR ORIFICE STRUCTURE, it is allowed to be quickly added to blood sample on the liquid level of density gradient centrifugation liquid, and the mixing of the anti-Hemostatic Oral Liquid layer of energy and centrifugate, the saving time, separation is simple, and due to boring the separation cushioning effect of socket bore plate, red blood cell below will not be poured out therewith；It is PBMC by the mixture the poured out cell precipitation that conventional centrifugal is obtained again；Only the shorter time is needed to be easily accomplished PBMC separation, and whole process uniformity is good, it is time saving and energy saving, make PBMC separation easy, fast, unanimously.
Description
Technical field
The present invention relates to technical field of cell separation, and in particular to a kind of mononuclearcell separating pipe and its application method.
Background technology
The adoptive cellular immunotherapy of tumour is the 4th kind of tumor therapeuticing method after operation, chemotherapy and radiation.Body
, it is necessary to collection patient's 50~100ml peripheral bloods, and separate have in peripheral blood single in advance during outer culture autoimmune cell
Nucleus, i.e. PMNC (Peripheral blood mononuclear cell, PBMC), including lymph are thin
Born of the same parents and monocyte.Separation method main at present is to carry out density gradient centrifugation using sample rate separating liquid, because blood
In the proportion of each visible component have differences, therefore separated.Red blood cell and granulocyte density are deposited on pipe more than separating liquid
Bottom.Blood platelet is then suspended in blood plasma because density is small, has the mononuclearcell suitable with separating liquid density only intensive in blood plasma
In the interface of layer and separating liquid, in tunica albuginea shape, draw the confluent monolayer cells and be resuspended by washing centrifugation, you can for exempting from for therapeutical uses
Epidemic disease cell expansion ex vivo culture.
At present, when application sample rate separating liquid carries out PBMC separation, the centrifuge tube maximum specification of use is generally 15ml
Or 50ml, the peripheral blood sample amount of often pipe separation is 10~15ml every time, and operation requires high during sample-adding, accidentally can cause
Anticoagulation separates liquid phase with sample rate and mixed, and influences the separating effect of mononuclearcell.In the market has a kind of Greiner bio-
One lymph/Monocyte dissociation tube, includes the porous sieve plate of a high-quality weight northylen, and the version is firmly embedded in polypropylene
In the centrifuge tube of material, by strictly controlling the pore size of sieve plate to separate cell, the processing time of sample is substantially reduced.
This kind of separating pipe mainly has two kinds of specifications：12ml and 50ml, its 50ml separating pipes sample size per treatment is 15~30ml blood samples,
Sample throughput is added compared with the common centrifuge tube that conventional method is used, processing time is shortened, reduces operation difficulty, is improved
Operating efficiency, but the product use cost is higher, it is necessary to further develop cheap and good-quality PMNC
Separation consumptive material.
The content of the invention
It is an object of the invention to provide a kind of cell Separating tube and its application method, one in above-mentioned prior art problem is solved
It is individual or multiple.
According to a kind of mononuclearcell separating pipe of the present invention, including centrifuge tube, it is provided with inclined at the mouth of pipe of the centrifuge tube
Lower half cross-section is provided with cone socket bore plate in side leakage bucket dividing plate, the centrifuge tube；The inclined side leakage bucket dividing plate and the centrifugation
Some positive cone socket bore and back taper is distributed with uniform intervals on the half side tube wall one inclined side leakage bucket structure of formation of pipe, the cone socket bore plate
Socket bore.
In some embodiments, the inclined side leakage bucket dividing plate includes partition board portion and is connected to inclined below the partition board portion
Side leakage pan, the partition board portion is distributed in the centrifuge tube vertically, the partition board portion both side edges and the centrifugation inside pipe wall phase
The centrifuge tube mouth of pipe is divided into inclined side leakage bucket semicircular opening and centrifuge tube semicircular opening two parts, the inclined side leakage by laminating, the partition board portion
Pan is drawn in side tube wall, is collectively forming inclined side leakage bucket with side centrifugation tube wall, the bottom of the inclined side leakage pan is provided with
It is close to the semi-circular opening of tube wall, the A/F is 0.1mm~3mm.
In some embodiments, the cone socket bore plate is the circular slab being adapted with the centrifuge tube cross section, described
Aperture is small in positive cone socket bore and the distribution of back taper socket bore uniform intervals, the positive cone socket bore, and lower aperture is big, hole in the back taper socket bore
Mouthful big, lower aperture is small, the positive cone socket bore and the back taper socket bore be conical, positive six side taper, square taper, rhombus taper,
One or more combinations in rectangle taper, triangular pyramidal, sine curve taper, the cone socket bore plate periphery and the centrifugation
Inside pipe wall fits.
In some embodiments, the thickness of the cone socket bore plate is 2mm~8mm, the positive cone socket bore and the back taper
The wall thickness of cone nest wall between socket bore is 0.1mm~3mm, the cone nest wall and positive cone socket bore and the folder of back taper socket bore axis
Angle is 20 °~80 °.
A kind of mononuclearcell separating pipe as claimed in claim 1 carries out the application method of cell separation, including step：
1) the sample rate separating liquid that volume is equal below the cone socket bore plate with separating pipe is drawn to arrive through centrifuge tube semicircular opening
In separating pipe；
2) separating pipe is centrifuged 25-35 seconds under 1000g centrifugal force using centrifuge, enters sample rate separating liquid
And full of cone socket bore plate underlying space；
3) keep separating pipe upright, pour into anti-freezing blood sample in the inclined side leakage bucket semicircular opening of separating pipe, anticoagulation is from inclined
Side funnel bottom semi-circular opening outflow, edge centrifuges tube wall downstream to cone socket bore plate top；
4) separating pipe is centrifuged 8-12 minutes under 1000g centrifugal force using centrifuge, or using centrifuge in 800g
Centrifugal force under to separating pipe centrifuge 13-20 minutes；After centrifugation in separating pipe sample from top to bottom successively be layered it is as follows：Blood plasma-
Mononuclearcell-separating liquid-cone socket bore plate-separating liquid-red blood cell；
5) mononuclearcell is drawn from centrifuge tube semicircular opening using pipette, or directly quickly inclined through centrifuge tube semicircular opening
Pour into another conventional centrifugal pipe, PBS is added in new conventional centrifugal pipe, cleaning cell is resuspended with pipette, centrifugation is used
Machine is centrifuged 3-7 minutes under 250g centrifugal force；
6) repeated washing cell 2 times, are resuspended with PBS or cell culture fluid and obtain mononuclearcell, directly deposited in and freeze
Pipe or progress next stage use.
In some embodiments, in step 1, when being preinstalled with sample rate separating liquid below cone socket bore plate, then not
Need step 1.
In some embodiments, the centrifugally operated in step 2,4,5 is carried out at room temperature.
In some embodiments, anticoagulation sample need to be such as diluted, can be diluted outside separating pipe in step 3,
Directly balanced salt solution can be poured into separating pipe to be diluted.
A kind of cell Separating tube provided in the present invention, the mouth of pipe is provided with inclined side leakage bucket dividing plate, is total to side centrifugation tube wall
With inclined side leakage bucket structure is formed, in the case where keeping separating pipe upright, anticoagulation directly is poured into through inclined side leakage bucket structure, can be with
Ensure that anticoagulation flows down along centrifugation tube wall；Using cone nest AND DEWATERING FOR ORIFICE STRUCTURE, it is allowed to blood sample quickly is added into density gradient centrifugation
On the liquid level of liquid, and the mixing of the anti-Hemostatic Oral Liquid layer of energy and centrifugate, at the end of centrifugation, slow down without excessively gentle very long.Centrifugation
Afterwards, both PBMC cellular layers can be drawn with suction pipe, also can be directly by plasma layer, PBMC cellular layers and the part sample above cone socket bore plate
This Density Separation liquid is easily quickly poured into new conventional centrifugal pipe, due to boring the separation cushioning effect of socket bore plate, below
Red blood cell will not be poured out therewith；It is PBMC by the mixture the poured out cell precipitation that conventional centrifugal is obtained again；Only need shorter
Time can be easily accomplished PBMC separation, and whole process uniformity is good, time saving and energy saving, makes PBMC separation easy, fast,
Unanimously.
Brief description of the drawings
Fig. 1 is a kind of structural representation of mononuclearcell separating pipe of one embodiment of the present invention；
Fig. 2 removes the structural representation of lid for a kind of mononuclearcell separating pipe of one embodiment of the present invention；
Fig. 3 shows for a kind of plan structure of the cone socket bore plate of mononuclearcell separating pipe of one embodiment of the present invention
It is intended to；
Fig. 4 shows for a kind of side view structure of the cone socket bore plate of mononuclearcell separating pipe of one embodiment of the present invention
It is intended to.
Embodiment
With reference to Figure of description, the present invention is described in more detail.
As illustrated in fig. 1 or fig. 2, inclined side is provided with a kind of cell Separating tube, including centrifuge tube 1, the mouth of pipe of centrifuge tube 1
The mouth of pipe of centrifuge tube 1 is divided into two parts by funnel dividing plate 2, inclined side leakage bucket dividing plate 2, and the effect of inclined side leakage bucket dividing plate 2 is to pour into anti-
Blood coagulation sample, anticoagulation flows out from the inclined semi-circular opening 23 of side funnel bottom 2, along the tube wall of centrifuge tube 1 downstream to cone socket bore plate 3 on
Side, the inside lower half cross-section of centrifuge tube 1 is provided with cone socket bore plate 3, and overall cone socket bore plate 3 is in circular plate shape；Inclined side leakage bucket every
Plate 2 and the one inclined side leakage of formation of the half side tube wall of centrifuge tube 1 are bucket-shaped, and inclined side leakage bucket dividing plate 2 includes partition board portion 21 and is integrally formed or welds
The mode of connecing is fixedly connected on the inclined side leakage pan 22 of the bottom of partition board portion 21, and partition board portion 21 is distributed in centrifuge tube 1 vertically, partition board portion
21 both side edges are fitted with the inwall of centrifuge tube 1, and the mouth of pipe of centrifuge tube 1 is divided into inclined side leakage bucket semicircular opening 11 and centrifugation by partition board portion 21
In the two parts of pipe semicircular opening 12, the present embodiment, the mouth of pipe of separating pipe 1 is divided into inclined side leakage bucket semicircular opening 11 and centrifugation by partition board portion 21
Pipe semicircular opening 12, inclined side leakage bucket semicircular opening 11 and centrifuge tube semicircular opening 12 are in semicircle shape, and partition board portion 21 is straight down along separation
The tube wall of pipe 1 extends and is connected with inclined side leakage pan 22, and the inclined edge of side leakage pan 22 is affixed to merge with centrifuge tube 1 makes inclined side leakage bucket
Portion 22 is drawn at edge to side, and one inclined side leakage of formation is bucket-shaped between inclined side leakage pan 22 and separating pipe 1, the inclined bottom of side leakage pan 22
Provided with the semi-circular opening 23 for being close to tube wall, the width of semi-circular opening 23 is 0.1mm~3mm, can be according to actual use demand
Limit A/F size.
As shown in Fig. 3 or Fig. 4, some positive cone socket bore 31 and some back taper nests are distributed with uniform intervals on cone socket bore plate 3
Hole 32, the generally circular plate-like shape of cone socket bore plate 3, is arranged at aperture in the cross section that separating pipe 1 manages interior bottom, positive cone socket bore 31
Small, lower aperture is big, and aperture is big in back taper socket bore 32, and lower aperture is small, thus on cone socket bore plate 3, positive cone socket bore 31 and back taper nest
The uniform intervals of hole 32 are distributed on cone socket bore plate 3, and positive cone socket bore 31 and back taper socket bore 32 are conical, positive six side taper, side's cone
In one or more combinations in shape, rhombus taper, rectangle taper, triangular pyramidal, sine curve cone taper, the present embodiment,
Using cone, thus, the center of circle of positive cone socket bore 31 and back taper socket bore 32 is arranged in arrays on cone socket bore plate 3, and this measure can be most
Utilize on a large scale and bore socket bore plate 3.
The thickness for boring socket bore plate 3 is 2mm~8mm, and the thickness of cone socket bore plate, positive cone can be selected according to actual use demand
The wall thickness of cone nest wall between socket bore 31 and back taper socket bore 32 is 0.1mm~3mm, cone nest wall and positive cone socket bore 31 and back taper nest
The angle of the axis of hole 32 is 20 °~80 °, is 45 ° in the present embodiment.
A kind of mononuclearcell separating pipe as claimed in claim 1 carries out the application method of cell separation, including step：
1) the sample rate separating liquid that volume is equal below the cone socket bore plate with separating pipe is drawn to arrive through centrifuge tube semicircular opening
In separating pipe；
2) separating pipe is centrifuged 25-35 seconds under 1000g centrifugal force using centrifuge, enters sample rate separating liquid
And full of cone socket bore plate underlying space；
3) keep separating pipe upright, pour into anti-freezing blood sample in the inclined side leakage bucket semicircular opening of separating pipe, anticoagulation is from inclined
Side funnel bottom semi-circular opening outflow, edge centrifuges tube wall downstream to cone socket bore plate top；
4) separating pipe is centrifuged 8-12 minutes under 1000g centrifugal force using centrifuge, or using centrifuge in 800g
Centrifugal force under to separating pipe centrifuge 13-20 minutes；After centrifugation in separating pipe sample from top to bottom successively be layered it is as follows：Blood plasma-
Mononuclearcell-separating liquid-cone socket bore plate-separating liquid-red blood cell；
5) mononuclearcell is drawn from centrifuge tube semicircular opening using pipette, or directly quickly inclined through centrifuge tube semicircular opening
Pour into another conventional centrifugal pipe, PBS is added in new conventional centrifugal pipe, cleaning cell is resuspended with pipette, centrifugation is used
Machine is centrifuged 3-7 minutes under 250g centrifugal force；
6) repeated washing cell 2 times, are resuspended with PBS or cell culture fluid and obtain mononuclearcell, directly deposited in and freeze
Pipe or progress next stage use.
In step 1, when being preinstalled with sample rate separating liquid below cone socket bore plate, then step 1 is not needed.
Centrifugally operated in step 2,4,5 is carried out at room temperature.
Anticoagulation sample need to be such as diluted, can be diluted outside separating pipe in step 3, can also fallen directly in separating pipe
Enter balanced salt solution to be diluted.
A kind of mononuclearcell separating pipe provided in the present invention, the mouth of pipe is provided with inclined side leakage bucket dividing plate, with side from
Heart tube wall is collectively forming inclined side leakage bucket structure, in the case where keeping separating pipe upright, is directly poured into through inclined side leakage bucket structure anti-
Blood coagulation, it is ensured that anticoagulation flows down along centrifugation tube wall；Using cone nest AND DEWATERING FOR ORIFICE STRUCTURE, it is allowed to blood sample quickly is added into density
On the liquid level of gradient centrifugation liquid, and the mixing of the anti-Hemostatic Oral Liquid layer of energy and centrifugate, at the end of centrifugation, slow down and overflow without excessively gentle
It is long.After centrifugation, both PBMC cellular layers can be drawn with suction pipe, also can be directly by the plasma layer above cone socket bore plate, PBMC cellular layers
Easily poured into part sample rate separating liquid in new conventional centrifugal pipe, due to boring the separation cushioning effect of socket bore plate, under it
The red blood cell of side will not be poured out therewith；It is PBMC by the mixture the poured out cell precipitation that conventional centrifugal is obtained again；Only need
The shorter time can be easily accomplished PBMC separation, and whole process uniformity is good, time saving and energy saving, make PBMC separation easy,
Fast, unanimously.
Described above is only the preferred embodiment of the present invention, it is noted that to those skilled in the art, not
On the premise of departing from the invention design, some similar modification and improvement can also be made, these also should be regarded as the present invention
Protection domain within.
Claims (8)
1. it is provided with a kind of mononuclearcell separating pipe, it is characterised in that including centrifuge tube (1), the mouth of pipe of the centrifuge tube (1)
Inclined side leakage bucket dividing plate (2), the interior lower half cross-section of centrifuge tube (1) is provided with cone socket bore plate (3)；The inclined side leakage bucket dividing plate
(2) with the half side tube wall one inclined side leakage bucket structure of formation of the centrifuge tube (1), uniform intervals distribution on the cone socket bore plate (3)
There are some positive cone socket bore (31) and back taper socket bore (32).
2. a kind of mononuclearcell separating pipe according to claim 1, it is characterised in that the inclined side leakage bucket dividing plate (2)
Including partition board portion (21) and the inclined side leakage pan (22) being connected to below the partition board portion (21), the partition board portion (21) is divided vertically
It is distributed in the centrifuge tube (1), partition board portion (21) both side edges fit with the centrifuge tube (1) inwall, the partition board portion
(21) centrifuge tube (1) mouth of pipe is divided into inclined side leakage bucket semicircular opening (11) and centrifuge tube semicircular opening (12) two parts, the inclined side leakage
Pan (22) is drawn in side tube wall, and inclined side leakage bucket, the inclined side leakage pan (22) are collectively forming with side centrifuge tube (1) tube wall
Bottom be provided with and be close to the semi-circular opening (23) of tube wall, described opening (23) width is 0.1mm~3mm.
3. a kind of mononuclearcell separating pipe according to claim 1, it is characterised in that the cone socket bore plate (3) be with
The adaptable circular slab in centrifuge tube (1) cross section, the positive cone socket bore (31) and the distribution of back taper socket bore (32) uniform intervals,
Aperture is small on the positive cone socket bore (31), and lower aperture is big, and aperture is big on the back taper socket bore (32), and lower aperture is small, the positive cone
Socket bore (31) and the back taper socket bore (32) are conical, positive six side taper, square taper, rhombus taper, rectangle taper, pyrometric cone
One or more combinations in shape, sine curve taper, cone socket bore plate (3) periphery and the centrifuge tube (1) inwall phase
Laminating.
4. a kind of mononuclearcell separating pipe according to claim 3, it is characterised in that the thickness of the cone socket bore plate (3)
Spend for 2mm~8mm, the wall thickness of the cone nest wall between the positive cone socket bore (31) and the back taper socket bore (32) for 0.1mm~
3mm, the cone nest wall and the angle of positive cone socket bore (31) and back taper socket bore (32) axis are 20 °~80 °.
5. a kind of method that mononuclearcell separating pipe as claimed in claim 1 carries out cell separation, it is characterised in that including
Step：
1) the sample rate separating liquid equal with the cone socket bore plate of separating pipe lower section volume is drawn through centrifuge tube semicircular opening to separating
In pipe；
2) separating pipe is centrifuged 25-35 seconds under 1000g centrifugal force using centrifuge, sample rate separating liquid is entered and is filled
Completely bore socket bore plate underlying space；
3) keep separating pipe upright, pour into anti-freezing blood sample in the inclined side leakage bucket semicircular opening of separating pipe, anticoagulation is from inclined side leakage
Bucket bottom semi-circular opening outflow, edge centrifuges tube wall downstream to cone socket bore plate top；
4) using centrifuge under 1000g centrifugal force to separating pipe centrifuge 8-12 minute, or use centrifuge 800g from
Separating pipe is centrifuged 13-20 minutes under mental and physical efforts；After centrifugation in separating pipe sample from top to bottom successively be layered it is as follows：Blood plasma-single
Nucleus-separating liquid-cone socket bore plate-separating liquid-red blood cell；
5) mononuclearcell is drawn from centrifuge tube semicircular opening using pipette, or is directly quickly poured into through centrifuge tube semicircular opening
In another conventional centrifugal pipe, PBS is added in new conventional centrifugal pipe, cleaning cell is resuspended with pipette, is existed using centrifuge
Centrifuged 3-7 minutes under 250g centrifugal force；
6) repeated washing cell 2 times, are resuspended with PBS or cell culture fluid and obtain mononuclearcell, directly deposit in cryopreservation tube or
Progress next stage uses.
6. the method for cell separation according to claim 5, it is characterised in that in step 1, pre- below cone socket bore plate
During equipped with sample rate separating liquid, then step 1 is not needed.
7. the method for cell separation according to claim 5, it is characterised in that the centrifugally operated in step 2,4,5 is equal
Carry out at room temperature.
8. the method for cell separation according to claim 5, it is characterised in that anticoagulation sample need to such as be entered in step 3
Row dilution, can dilute, also directly can pour into balanced salt solution in separating pipe is diluted outside separating pipe.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710456928.4A CN107034128A (en) | 2017-06-16 | 2017-06-16 | A kind of mononuclearcell separating pipe and its application method |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710456928.4A CN107034128A (en) | 2017-06-16 | 2017-06-16 | A kind of mononuclearcell separating pipe and its application method |
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| Publication Number | Publication Date |
|---|---|
| CN107034128A true CN107034128A (en) | 2017-08-11 |
Family
ID=59541649
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|---|---|---|---|
| CN201710456928.4A Pending CN107034128A (en) | 2017-06-16 | 2017-06-16 | A kind of mononuclearcell separating pipe and its application method |
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| CN (1) | CN107034128A (en) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109794082A (en) * | 2019-03-20 | 2019-05-24 | 林侯汎 | It is a kind of for extracting the device of mononuclearcell in blood |
| CN110591901A (en) * | 2019-10-27 | 2019-12-20 | 李文强 | Cell separation tube |
| CN111117957A (en) * | 2019-12-24 | 2020-05-08 | 泰州市人民医院 | Separation method for separating peripheral blood mononuclear cells |
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| US5474687A (en) * | 1994-08-31 | 1995-12-12 | Activated Cell Therapy, Inc. | Methods for enriching CD34+ human hematopoietic progenitor cells |
| CN103952295A (en) * | 2014-05-04 | 2014-07-30 | 李文强 | Cell separating device |
| CN104031831A (en) * | 2014-07-01 | 2014-09-10 | 北京圣浦博大生物科技有限公司 | Blood cell separation tube and separation method of mononuclear cell |
| CN204022830U (en) * | 2014-08-19 | 2014-12-17 | 任峰 | A kind of cell Separating tube for separating of extracting PBMC |
| CN207079232U (en) * | 2017-06-16 | 2018-03-09 | 何向锋 | A kind of mononuclearcell separating pipe |
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2017
- 2017-06-16 CN CN201710456928.4A patent/CN107034128A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5474687A (en) * | 1994-08-31 | 1995-12-12 | Activated Cell Therapy, Inc. | Methods for enriching CD34+ human hematopoietic progenitor cells |
| CN103952295A (en) * | 2014-05-04 | 2014-07-30 | 李文强 | Cell separating device |
| CN104031831A (en) * | 2014-07-01 | 2014-09-10 | 北京圣浦博大生物科技有限公司 | Blood cell separation tube and separation method of mononuclear cell |
| CN204022830U (en) * | 2014-08-19 | 2014-12-17 | 任峰 | A kind of cell Separating tube for separating of extracting PBMC |
| CN207079232U (en) * | 2017-06-16 | 2018-03-09 | 何向锋 | A kind of mononuclearcell separating pipe |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109794082A (en) * | 2019-03-20 | 2019-05-24 | 林侯汎 | It is a kind of for extracting the device of mononuclearcell in blood |
| CN110591901A (en) * | 2019-10-27 | 2019-12-20 | 李文强 | Cell separation tube |
| CN111117957A (en) * | 2019-12-24 | 2020-05-08 | 泰州市人民医院 | Separation method for separating peripheral blood mononuclear cells |
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Effective date of registration: 20240326 Address after: Room 308, No. 42, Guangzhou Road, Zhongxing Street, Development Zone, Nantong City, Jiangsu Province, 226000 Applicant after: Mingchang Biopharmaceutical Nantong Co.,Ltd. Country or region after: China Address before: 226,000 205, Building 7, Chenyuan, Chongchuan District, Nantong, Jiangsu Applicant before: He Xiangfeng Country or region before: China |