CN110354142A - Treatment method of people's external amplification natural killer cell to ovarian Cancer of Nude Mice solid tumor - Google Patents
Treatment method of people's external amplification natural killer cell to ovarian Cancer of Nude Mice solid tumor Download PDFInfo
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- A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
- A61K35/14—Blood; Artificial blood
- A61K35/17—Lymphocytes; B-cells; T-cells; Natural killer cells; Interferon-activated or cytokine-activated lymphocytes
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Abstract
The present invention relates to a kind of people's external amplification natural killer cells to the treatment method of ovarian Cancer of Nude Mice solid tumor, successively include people's external amplification natural killer cell, i.e., the culture experiment of NK cell, prepare ovarian cancer cell SK-OV-3+RFP+LUC experiment, NK cell to ovarian cancer cell SK-OV-3+RFP+LUC killing effect in vitro experiment and NK cell lethal effect in ovarian cancer cell SK-OV-3+RFP+LUC body is tested.The present invention solves conventional reagents box and expands NK cell phenotype instability problem, reach 35%-70% with the NK cell purity of this method culture, and stablize, NK cell has apparent killing effect to SK-OV-3 cell, NK cell can be such that nude mice by subcutaneous ovarian cancer cell SK-OV-3 solid tumor is obviously reduced, adjuvant treatment can be provided for clinical treatment SK-OV-3 Ovarian carcinoma, than general chemicotherapy Small side effects, effect is also obvious.
Description
Technical field
The invention belongs to field of biotechnology, are related to the treatment technology of Ovarian carcinoma, and especially a kind of human body extends out
Increase natural killer cells to the treatment method of ovarian Cancer of Nude Mice solid tumor.
Background technique
Malignant tumor of ovary is common one of the malignant tumour of female sex organ, and disease incidence is only second to cervix cancer and son
Carcinoma of corpus uteri and arrange and occupy third position.Since ovary embryonic development, anatomic tissue and endocrine function are more complex, early symptom not allusion quotation
Type, the organization type of preoperative identification of ovarian tumour and good pernicious extremely difficult, so ovarian epithelium mortality of carcinoma but accounts for all kinds of woman
Also there is a great difficulty in the first place of section's tumour in malignant tumor of ovary treatment at present, after best treatment method is operative treatment,
Radiation and chemotherapy.But cancer operation is big to human body wound, makes the reduction of patient's immunity, declines to the resilience of disease, and
And a series of postoperative complications are also easy to produce after performing the operation.Operation at present can only be only applicable to early carcinoma as being local treatment means
Swollen range limitation, postoperative patient also not only will have killing left and right, human body to cancer cell during chemicotherapy through a series of chemicotherapy
Normal cell can also be killed, and human body side effect is big during most chemotherapy, it may appear that Nausea and vomiting, leg fiber crops, body are weary
Power, apathetic etc..
Tumour can be treated by also having many documents to tell about immunocyte at present, but the immunocyte phenotype of people's amplification in vitro is logical
Chang Bugao, that is, purity are bad, and therapeutic effect will be affected, and with immune cell therapy oophoroma (SK-OV-3)
Solid tumor yet seldom has been reported that.
Summary of the invention
It is an object of the invention to provide a kind of NK cell culture processes used in place of overcome the deficiencies in the prior art,
Simple and convenient, oneself configuration factor, it is economical and practical the problems such as not having to cultivate reagent box, avoid the factor unstable, and to nude mice
The apparent people's external amplification natural killer cell of therapeutic effect of oophoroma (SK-OV-3) solid tumor is to ovarian Cancer of Nude Mice solid tumor
Treatment method.
The present invention solves its technical problem and adopts the following technical solutions to achieve:
A kind for the treatment of method of people's external amplification natural killer cell to ovarian Cancer of Nude Mice solid tumor, it is characterised in that: side
Steps are as follows for method: successively progress people's external amplification natural killer cell, the i.e. culture experiment of NK cell prepare ovarian cancer cell
SK-OV-3+RFP+LUC experiment, NK cell test ovarian cancer cell SK-OV-3+RFP+LUC killing effect in vitro
And NK cell tests lethal effect in ovarian cancer cell SK-OV-3+RFP+LUC body.
Moreover, people's external amplification natural killer cell, i.e. the culture experiment method and step of NK cell is as follows:
(1) healthy human peripheral blood 50mL being acquired with 10mL heparin sodium heparin tube, is centrifuged 800g, 8 liter of 4 drop draws plasma layer,
Upper plasma layer is put into 56 DEG C of water-baths of constant temperature, is inactivated 30min, is centrifuged after the completion of inactivation, revolving speed 2000g, time 10min, 8
8 drops are risen, take supernatant after the completion of centrifugation, are freezed in -20 DEG C, are melted, then be centrifuged 2000g, 8 liter of 8 drop takes supernatant after the completion of centrifugation；
(2) lower layer's red blood cell supplies original volume with physiological saline, i.e., containing blood plasma when volume, then with physiological saline 1:
1 dilution, is uniformly mixed；
(3) red blood cell and ficoll diluted is mixed with the ratio of 4:3, is centrifuged 600g, time 40min, and 1 liter 0 is dropped, from
Tunica albuginea layer is drawn after the heart, is whitened film layer with physiological saline, is repeated twice；
(4) mononuclearcell concentration is adjusted with GT-T551-H3 culture medium, with final concentration of cells for 2 × 106A/mL connects
Kind, cell factor IL-2,2810U/mL, IL-15,50U/mL, CD16,50ng/mL are added in cell | and 5% autoserum；
(5) second days additions CD3,2ng/mL；
Cell was centrifuged in (6) the 5th days, discards culture medium, adjusting concentration with new GT-T551-H3 culture medium is 1 × 106
A/mL adds 5% autoserum and IL-2,2810U/mL；
(7) cultivating the 7th day GT-T551-H3 culture medium and adjusting concentration is 1 × 106A/mL, addition 1% autoserum and
IL-2,1405U/mL, until culture was by the 14th day；
(8) the 14th days, supernatant is removed after centrifugation, cell is resuspended in physiological saline, it is centrifuged, 300g, 5min, 9 liters of 9 drops of revolving speed, from
Supernatant is removed after the heart, is repeated 4 times, and cell is resuspended with physiological saline, 80 mesh cell sieve filtration cells count.
Moreover, described prepares ovarian cancer cell SK-OV-3+RFP+LUC experimental method step are as follows: select lucky Ma slow virus
Ovarian cancer cell SK-OV-3 is transfected, MOI value is 50, and the cell to fluorescence reaches 100%, and can stablize passage.
Moreover, the NK cell is to ovarian cancer cell SK-OV-3+RFP+LUC killing effect in vitro experimental method step
It is as follows:
(1) ovarian cancer cell SK-OV-3+RFP+LUC spreads 96 orifice plates, every 8 × 103/ hole of hole density；
(2) NK cell and ovarian cancer cell SK-OV-3+RFP+LUC, i.e. effector cell: target cell 2.5:1 exist respectively
It is shown with CC-8 kit detection NK cell to the killing rate of sk-ov-3 cell, and in fluorescence within 24 hours, 48 hours, 72 hours
Micro- microscopic observation cell quantity.
Moreover, the NK cell includes ovary to lethal effect experiment in ovarian cancer cell SK-OV-3+RFP+LUC body
The foundation of cancer cell SK-OV-3+RFP+LUC solid tumor models, steps are as follows for specific method:
(1) 5 week old Female nude mices are selected, subcutaneous injection SK-OV-3+RFP+LUC total number of cells are 2 × 106, occur within 7 days
The protrusion of 0.2cm × 0.2cm or so, experiment were randomly divided into four groups at the 7th day, were model control group, NK high dose group, NK respectively
Middle dose group, NK low dose group, every group of 3 nude mices；
(2) start tail vein injection NK cell after a week, point high, normal, basic three dosage, high dose 2 × 106/ only, middle dosage
1×106/ only, low dosage 0.5 × 106/ only, it once a week, is administered 4 times altogether, through flow cytometer detection, administration CD3-, CD5+ is thin for the first time
Born of the same parents, NK cell account for lymph group 70.1%, and second of administration CD3-, CD5+ cell, NK cell accounts for lymph group 53.1%, third time
CD3-, CD5+ cell is administered, NK cell accounts for lymph group 70.6%, the 4th administration CD3-, CD5+ cell, and NK cell accounts for lymph
Group 35.0%；
(3) respectively 10 days after injecting NK cell, 14 days, 21 days, 28 days, 35 days, 47 days, 49 days, 56 days measurement nude mices
Tumor volume；
(4) respectively 10 days after injecting NK cell, 14 days, 21 days, 28 days, 35 days, 47 days, 49 days, 55 days detection knurls
Fluorescence intensity；
(5) nude mice is supported to dissection in 56 days and draws materials, and weighs tumor weight and takes out nude mice spleen progress cytokines measurement, point
Not Jian Ce cell factor IL-2, IL-2 be Porcine HGF in immune system, mainly generated by T cell and T cell system,
At present clinically IL-2 be mainly used for it is antitumor, so this experiment take treatment after nude mice spleen ELISA kit measurement IL-
2 content.
The advantages and positive effects of the present invention are:
The NK cell culture processes that the present invention uses solve conventional reagents box and expand NK cell phenotype instability problem, use
The NK cell purity of this method culture reaches 35%-70%, and stablizes, and NK cell has apparent killing to SK-OV-3 cell
Effect, it can be clinical treatment SK-OV-3 that NK cell, which can be such that nude mice by subcutaneous ovarian cancer cell SK-OV-3 solid tumor is obviously reduced,
Ovarian carcinoma provides adjuvant treatment, and than general chemicotherapy Small side effects, effect is also obvious.And it is simple to operate, from
The problems such as oneself configures the factor, does not have to cultivate reagent box, avoids the factor unstable, it is economical and practical, by the way that experimental results demonstrate to naked
The therapeutic effect of mouse oophoroma (SK-OV-3) solid tumor is obvious, has great directive significance to clinical treatment.
Detailed description of the invention
Fig. 1 is NK cell microscopic (20X)；
Fig. 2 is to prepare ovarian cancer cell SK-OV-3+RFP+LUC micrograph, and wherein a is 10X normal visual field, and b is that 10X is glimmering
The light visual field；
Fig. 3 is killing-efficiency analysis chart of the NK cell to SK-OV-3 cell；
Fig. 4 is lethal effect fluorogram of the NK cell to SK-OV-3 cell；
Fig. 5 is tumor formation figure under ovarian Cancer of Nude Mice SK-OV-3+RFP+LUC cell skin；
Fig. 6 is that first time tail vein injection CD3-, CD56+ account for 70.1% analysis chart of lymph group；
Fig. 7 is that second tail vein injection CD3-, CD56+ account for 53.1% analysis chart of lymph group；
Fig. 8 is that third time tail vein injection CD3-, CD56+ account for 70.6% analysis chart of lymph group；
Fig. 9 is that the 4th tail vein injection CD3-, CD56+ account for 35.0% analysis chart of lymph group；
Figure 10 is SK-OV-3+RFP+LUC cell nude mice by subcutaneous tumor growth curve figure；
Figure 11 is tumorous size comparison diagram after the 56th day nude mice dissection materials；
Figure 12 is SK-OV-3+RFP+LUC cell nude mice by subcutaneous tumor fluorescence intensity growth curve；
Figure 13 is the column diagram of each group IL-2 concentration in spleen suspension；
Figure 14 is the column diagram of each group TNF-a concentration in spleen suspension.
Specific embodiment
The invention will be further described with reference to the accompanying drawing and by specific embodiment, and following embodiment is descriptive
, it is not restrictive, this does not limit the scope of protection of the present invention.
A kind for the treatment of method of people's external amplification natural killer cell to ovarian Cancer of Nude Mice solid tumor, it is characterised in that: side
Steps are as follows for method: successively progress people's external amplification natural killer cell, the i.e. culture experiment of NK cell prepare ovarian cancer cell
SK-OV-3+RFP+LUC experiment, NK cell test ovarian cancer cell SK-OV-3+RFP+LUC killing effect in vitro
And NK cell tests lethal effect in ovarian cancer cell SK-OV-3+RFP+LUC body.
Moreover, people's external amplification natural killer cell, i.e. the culture experiment method and step of NK cell is as follows:
(1) healthy human peripheral blood 50mL being acquired with 10mL heparin sodium heparin tube, is centrifuged 800g, 8 liter of 4 drop draws plasma layer,
Upper plasma layer is put into 56 DEG C of water-baths of constant temperature, is inactivated 30min, is centrifuged after the completion of inactivation, revolving speed 2000g, time 10min, 8
8 drops are risen, take supernatant after the completion of centrifugation, are freezed in -20 DEG C, are melted, then be centrifuged 2000g, 8 liter of 8 drop takes supernatant after the completion of centrifugation；
(2) lower layer's red blood cell supplies original volume with physiological saline, i.e., containing blood plasma when volume, then with physiological saline 1:
1 dilution, is uniformly mixed；
(3) red blood cell and ficoll diluted is mixed with the ratio of 4:3, is centrifuged 600g, time 40min, and 1 liter 0 is dropped, from
Tunica albuginea layer is drawn after the heart, is whitened film layer with physiological saline, is repeated twice；
(4) mononuclearcell concentration is adjusted with GT-T551-H3 culture medium, with final concentration of cells for 2 × 106A/mL connects
Kind, cell factor IL-2,2810U/mL, IL-15,50U/mL, CD16,50ng/mL are added in cell | and 5% autoserum；
(5) second days additions CD3,2ng/mL；
Cell was centrifuged in (6) the 5th days, discards culture medium, adjusting concentration with new GT-T551-H3 culture medium is 1 × 106
A/mL adds 5% autoserum and IL-2,2810U/mL；
(7) cultivating the 7th day GT-T551-H3 culture medium and adjusting concentration is 1 × 106A/mL, addition 1% autoserum and
IL-2,1405U/mL, until culture was by the 14th day；
(8) the 14th days, supernatant is removed after centrifugation, cell is resuspended in physiological saline, it is centrifuged, 300g, 5min, 9 liters of 9 drops of revolving speed, from
Supernatant is removed after the heart, is repeated 4 times, and cell is resuspended with physiological saline, 80 mesh cell sieve filtration cells count, as a result as shown in Figure 1.
Moreover, described prepares ovarian cancer cell SK-OV-3+RFP+LUC experimental method step are as follows: select lucky Ma slow virus
Ovarian cancer cell SK-OV-3 is transfected, MOI value is 50, and the cell to fluorescence reaches 100%, and can stablize passage, as a result such as
Shown in Fig. 2, Fig. 2 a is normal visual field 10X, and Fig. 2 a is fluorescence visual field 10X, and experiment needs the stable cell lines with fluorescence, cell note
Enter to be determined with fluorescence intensity in nude mouse internal SK-OV-3 cell number, general fluorescence intensity is strong, cell just it is more.
It can also be observed simultaneously with cell fluorescence in vitro to the lethal effect of SK-OV-3 cell.
Moreover, the NK cell is to ovarian cancer cell SK-OV-3+RFP+LUC killing effect in vitro experimental method step
It is as follows:
(1) ovarian cancer cell SK-OV-3+RFP+LUC spreads 96 orifice plates, every 8 × 103/ hole of hole density；
(2) NK cell and ovarian cancer cell SK-OV-3+RFP+LUC, i.e. effector cell: target cell 2.5:1 exist respectively
It is shown with CC-8 kit detection NK cell to the killing rate of sk-ov-3 cell, and in fluorescence within 24 hours, 48 hours, 72 hours
Micro- microscopic observation cell quantity finds out that experiment needs the stable cell lines with fluorescence, cell injection as shown in Figure 3, Figure 4, in Fig. 3
Can be determined with fluorescence intensity in nude mouse internal SK-OV-3 cell number, general fluorescence intensity is strong, cell just it is more.Together
When can also be observed with cell fluorescence in vitro to the lethal effect of SK-OV-3 cell.As can be seen from Figure 4 NK groups of cells
SK-OV-3 cell fluorescence number is significantly lower than control group, at 48 hours, the substantially all death of cell.
Moreover, the NK cell includes ovary to lethal effect experiment in ovarian cancer cell SK-OV-3+RFP+LUC body
The foundation of cancer cell SK-OV-3+RFP+LUC solid tumor models, steps are as follows for specific method:
(1) 5 week old Female nude mices are selected, subcutaneous injection SK-OV-3+RFP+LUC total number of cells are 2 × 106, occur within 7 days
The protrusion of 0.2cm × 0.2cm or so, experiment were randomly divided into four groups at the 7th day, were model control group, NK high dose group, NK respectively
Middle dose group, NK low dose group, every group of 3 nude mices, as shown in figure 5, tumor formation rate 100% under SK-OV-3+RFP+LUC cell skin；
(2) start tail vein injection NK cell after a week, point high, normal, basic three dosage, high dose 2 × 106/ only, middle dosage
1×106/ only, low dosage 0.5 × 106/ only, it once a week, is administered 4 times altogether, through flow cytometer detection, administration CD3-, CD5+ is thin for the first time
Born of the same parents, NK cell account for lymph group 70.1%, as shown in fig. 6, second of administration CD3-, CD5+ cell, NK cell account for lymph group
53.1%, as shown in fig. 7, third time administration CD3-, CD5+ cell, NK cell accounts for lymph group 70.6%, as shown in figure 8, the 4th
Secondary administration CD3-, CD5+ cell, NK cell account for lymph group 35.0%, as shown in Figure 9；
(3) respectively 10 days after injecting NK cell, 14 days, 21 days, 28 days, 35 days, 47 days, 49 days, 56 days measurement nude mices
Tumor volume, as shown in Figure 10, within injection NK cell 42 days, each group knurl average external volume difference is little, opens from the 49th day
Begin, the average tumor volume of tumor model group is higher by NK group and is higher by 2 times, and the 56th day, the average tumor volume of tumor model group was high
NK group is higher by 3 times out, and such as Figure 11, the tumorous size of model control group is apparently higher than the tumorous size of NK each group；
(4) respectively 10 days after injecting NK cell, 14 days, 21 days, 28 days, 35 days, 47 days, 49 days, 55 days detection knurls
Fluorescence intensity, as shown in figure 12, injection NK cell 21 days after, the fluorescence intensity of tumor model group is always above NK each group.
It opens within 49th day, the average knurl fluorescence intensity of tumor model group is higher by NK group and is higher by 2 times；
(5) nude mice is supported to dissection in 56 days and draws materials, and weighs tumor weight and takes out nude mice spleen progress cytokines measurement, point
Not Jian Ce cell factor IL-2, IL-2 be Porcine HGF in immune system, mainly generated by T cell and T cell system,
At present clinically IL-2 be mainly used for it is antitumor, so this experiment take treatment after nude mice spleen ELISA kit measurement IL-
2 content.
Such as Figure 13, the IL-2 concentration of each group IL-2 concentration in spleen suspension, blank control is higher than tumor model group, and NK is high, normal, basic
Three groups of IL-2 concentration is higher than tumor model group.
TNF-a be a kind can direct killing tumour cell and the cell factor to normal cell without overt toxicity, be
One of strongest bioactie agent of direct killing function of tumor found so far.Such as Figure 14, each group in spleen suspension
TNF-a concentration, the TNF-a concentration of blank control are higher than tumor model group, and the TNF-a concentration that high, normal, basic three groups of NK is higher than tumour mould
Type group.
Although disclosing the embodiment of the present invention and attached drawing for the purpose of illustration, those skilled in the art can be managed
Solution: do not departing from the present invention and spirit and scope of the appended claims in, various substitutions, changes and modifications be all it is possible,
Therefore, the scope of the present invention is not limited to the embodiment and attached drawing disclosure of that.
Claims (5)
1. a kind of people's external amplification natural killer cell is to the treatment method of ovarian Cancer of Nude Mice solid tumor, it is characterised in that: method
Steps are as follows: successively progress people's external amplification natural killer cell, the i.e. culture experiment of NK cell prepare ovarian cancer cell SK-
OV-3+RFP+LUC experiment, NK cell is tested to ovarian cancer cell SK-OV-3+RFP+LUC killing effect in vitro and NK cell pair
Lethal effect is tested in ovarian cancer cell SK-OV-3+RFP+LUC body.
2. a kind of people's external amplification natural killer cell according to claim 1 is to the treatment side of ovarian Cancer of Nude Mice solid tumor
Method, it is characterised in that: the culture experiment method and step of people's external amplification natural killer cell, i.e. NK cell is as follows:
(1) healthy human peripheral blood 50mL is acquired with 10mL heparin sodium heparin tube, is centrifuged 800g, 8 liter of 4 drop draws plasma layer, upper layer
Plasma layer is put into 56 DEG C of water-baths of constant temperature, is inactivated 30min, is centrifuged after the completion of inactivation, revolving speed 2000g, time 10min, 8 liter 8
Drop takes supernatant after the completion of centrifugation, freezes in -20 DEG C, melts, then be centrifuged 2000g, and 8 liter of 8 drop takes supernatant after the completion of centrifugation；
(2) lower layer's red blood cell supplies original volume with physiological saline, i.e., containing blood plasma when volume, then it is dilute with physiological saline 1:1
It releases, is uniformly mixed；
(3) red blood cell and ficoll diluted is mixed with the ratio of 4:3, is centrifuged 600g, time 40min, 1 liter of 0 drop, after centrifugation
Tunica albuginea layer is drawn, film layer is whitened with physiological saline, is repeated twice；
(4) mononuclearcell concentration is adjusted with GT-T551-H3 culture medium, with final concentration of cells for 2 × 106A/mL inoculation, thin
Cell factor IL-2,2810U/mL, IL-15,50U/mL, CD16,50ng/mL are added in born of the same parents | and 5% autoserum；
(5) second days additions CD3,2ng/mL；
Cell was centrifuged in (6) the 5th days, discards culture medium, adjusting concentration with new GT-T551-H3 culture medium is 1 × 106A/mL,
Add 5% autoserum and IL-2,2810U/mL；
(7) cultivating the 7th day GT-T551-H3 culture medium and adjusting concentration is 1 × 106A/mL adds 1% autoserum and IL-2,
1405U/mL, until culture was by the 14th day；
(8) the 14th days, supernatant is removed after centrifugation, cell, centrifugation, 300g, 5min, 9 liters of 9 drops of revolving speed, after centrifugation are resuspended in physiological saline
Supernatant is removed, is repeated 4 times, cell is resuspended with physiological saline, 80 mesh cell sieve filtration cells count.
3. a kind of people's external amplification natural killer cell according to claim 1 is to the treatment side of ovarian Cancer of Nude Mice solid tumor
Method, it is characterised in that: described prepares ovarian cancer cell SK-OV-3+RFP+LUC experimental method step are as follows: select lucky Ma sick slowly
Poison transfection ovarian cancer cell SK-OV-3, MOI value are 50, and the cell to fluorescence reaches 100%, and can stablize passage.
4. a kind of people's external amplification natural killer cell according to claim 1 is to the treatment side of ovarian Cancer of Nude Mice solid tumor
Method, it is characterised in that: the NK cell is to ovarian cancer cell SK-OV-3+RFP+LUC killing effect in vitro experimental method step
It is as follows:
(1) ovarian cancer cell SK-OV-3+RFP+LUC spreads 96 orifice plates, every 8 × 103/ hole of hole density；
(2) NK cell and ovarian cancer cell SK-OV-3+RFP+LUC, i.e. effector cell: target cell 2.5:1 are small 24 respectively
When, 48 hours, 72 hours with CC-8 kit detection NK cell to the killing rate of sk-ov-3 cell, and in fluorescence microscope
Lower observation cell quantity.
5. a kind of people's external amplification natural killer cell according to claim 1 is to the treatment side of ovarian Cancer of Nude Mice solid tumor
Method, it is characterised in that: the NK cell includes ovary to lethal effect experiment in ovarian cancer cell SK-OV-3+RFP+LUC body
The foundation of cancer cell SK-OV-3+RFP+LUC solid tumor models, steps are as follows for specific method:
(1) 5 week old Female nude mices are selected, subcutaneous injection SK-OV-3+RFP+LUC total number of cells are 2 × 106, there is 0.2cm within 7 days
The protrusion of × 0.2cm or so, experiment at the 7th day were randomly divided into four groups, were model control group, NK high dose group, agent in NK respectively
Amount group, NK low dose group, every group of 3 nude mices；
(2) start tail vein injection NK cell after a week, point high, normal, basic three dosage, high dose 2 × 106/ only, middle dosage 1 ×
106/ only, low dosage 0.5 × 106/ only, it once a week, is administered 4 times altogether, CD3-, CD5+ cell is administered for the first time through flow cytometer detection,
NK cell accounts for lymph group 70.1%, and second of administration CD3-, CD5+ cell, NK cell accounts for lymph group 53.1%, gives for the third time
Medicine CD3-, CD5+ cell, NK cell account for lymph group 70.6%, the 4th administration CD3-, CD5+ cell, and NK cell accounts for lymph group
35.0%；
(3) respectively 10 days after injecting NK cell, 14 days, 21 days, 28 days, 35 days, 47 days, 49 days, 56 days measurement nude mice knurls
Volume；
(4) respectively 10 days after injecting NK cell, 14 days, 21 days, 28 days, 35 days, 47 days, 49 days, 55 days detection knurl it is glimmering
Luminous intensity；
(5) nude mice is supported to dissection in 56 days and draws materials, and weighs tumor weight and takes out nude mice spleen progress cytokines measurement, examines respectively
Cell factor IL-2 is surveyed, IL-2 is the Porcine HGF in immune system, is mainly generated by T cell and T cell system, at present
Clinically IL-2 is mainly used for antitumor, so this experiment takes nude mice spleen ELISA kit after treatment to measure IL-2's
Content.
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| CN112285081A (en) * | 2020-10-28 | 2021-01-29 | 上海睿钰生物科技有限公司 | Method for detecting cell killing efficacy and application thereof |
| WO2022089552A1 (en) * | 2020-10-28 | 2022-05-05 | 上海睿钰生物科技有限公司 | Method and system for detecting cell killing efficacy and/or immune activity, and application thereof |
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