CN106939298B - A kind of full tumour antigen preparation method of glioma and device - Google Patents
A kind of full tumour antigen preparation method of glioma and device Download PDFInfo
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Abstract
The present invention relates to a kind of full tumour antigen preparation method of glioma and device, include the following steps: the Multiplying culture carried out using serum-free medium to the glioma stem cell and glioma non-stem cell；The glioma stem cell of logarithmic growth phase and glioma non-stem cell are pre-processed；Pretreated glioma stem cell is mixed to be placed in lysis process chamber one with glioma non-stem cell and carries out pulse cracking and synchronous centrifugal treating；Supernatant is abandoned, above treated the glioma stem cell and glioma non-stem cell mixed liquor are placed in lysis process chamber two and carry out the cracking of pulse again, is i.e. the harvest full tumour antigen of glioma.The present invention realizes glioma stem cell and glioma non-stem cell treatment by stages and cracking using low frequency and high frequency steep-sided pulse, easy to operate, and cracking more sufficiently, quickly, can get more complete glioma tumour antigen.
Description
Technical field
The invention mainly relates to glioma field more particularly to a kind of full tumour antigen preparation methods of glioma
And device.
Background technique
Glioma is derived from neurepithelial tumour, is the most common malignant tumour of encephalic, accounts for about whole encephalics
The 40-50% of tumour, it is high with disease incidence height, high recurrence rate, lethality, become the problem of oncotherapy.Traditional therapy master
The survival probability of patient is improved if performing the operation with chemicotherapy.But the glioma for being located at critical function area is difficult to accomplish to cut entirely
It removes.Radiotherapy is almost the conventional therapy of various glioma, but therapeutic evaluation is different, except medulloblastoma is quick to radiotherapy height
Feel, outside ependymoma medium sensitivity, other types are insensitive to radiotherapy.Chemotherapeutics is limited to blood-brain barrier and the poison of drug is secondary
Effect, curative effect are not affirmed still.In recent years, immunization therapy becomes effective treatment means of the glioma after operation, chemicotherapy.With
Other modes use in conjunction specifically has very strong complementation, can play recovery to the immune system that patient is damaged and rebuild
Unique curative effect, to further prevent the recurrence and transfer of tumour.
Glioma antigen is obtained by cracking after carrying out Multiplying culture to glioma stem cell in the prior art,
Since tradition cracking is mainly using methods such as multigelation or radiation, cause cracking that can not exclusively obtain holoantigen information, and
And there are certain microbial contamination and animal sources immunogenicities in culture solution, therefore cause the glioma obtained dry
The Antigen Stability of cell preparation is poor, and specificity is weaker, to pass through the therapeutic epidemic disease of antigen glioma obtained
Seedling have stability it is poor, specificity it is weaker, kill the defects of ratio of outflow is not high and immunogenicity is weak.
Cell membrane can preferably hinder the transmission of ion and hydrophilic molecules under normal physiological function situation.But certain
The electric pulse effect of dosage is lower will to occur electroporation phenomenon, and the bilayer lipid membrane of cell, which is temporarily rearranged, forms some quilts
The referred to as hydrophilic pathway of micropore, enables large hydrophilic molecular to pass through.Conductivity changes when membrane perforation, such as right
The conductivity of sodium ion and potassium ion is usually less than 1mS/cm2；When transmembrane voltage be more than film dielectric strength when, conductivity compared with
It will increase sharply in short time to 1S/cm2, after causing cell membrane that particle penetrating power is hindered to reduce electric field cancellation, in most cases
Micropore can be closed without having any impact to cell, and temporary micropore occurs in this cell membrane under the effect of electric pulse in short-term
Physical process is known as electroporation i.e. invertibity electrical breakdown (REB).Electroporation greatly enhances cell membrane permeability, thus proposes
Electroporation therapy.Increasing electric pulse dosage, then the expendable rupture of cell membrane appearance punctures cell, this is irreversibility electricity
Puncture (IREB), and has been used in tumor cell lysis research
Summary of the invention
It is an object of the present invention to cracking is insufficient in order to solve in current glioma tumour antigen preparation process,
Seriously polluted the problems such as can not obtaining holoantigen information, the present invention is based on pulse breakdown principles, provide a kind of glioma
Full tumour antigen preparation method and device.
The technical scheme to solve the above technical problems is that a kind of full tumour antigen preparation method of glioma
And device, include the following steps:
A kind of full tumour antigen preparation method of glioma, includes the following steps:
Fresh Human Brain Gliomas is cleaned into blood stains with buffer, cuts into lmm3The samples of human glioma block of size is standby
With；
It is unicellular and carry out Multiplying culture glioma will to be obtained in glioma tumor tissue；
Glioma after proliferation is unicellular to isolate glioma stem cell and glioma non-stem cell；
The proliferation that the glioma stem cell and glioma non-stem cell are carried out using serum-free medium
Culture, and obtain glioma stem cell and the glioma non-stem cell of logarithmic growth phase；
The glioma stem cell of logarithmic growth phase and glioma non-stem cell are placed in pretreatment chamber one, pre-
It is pre-processed in process chamber two；
By number than for 1:2-1:5 glioma stem cell and glioma non-stem cell be respectively placed in pre- place
Cracking pretreatment 5-10min is carried out in reason room one and pretreatment chamber two；
With CO in the pretreatment chamber one2Atmosphere is filled up completely, and temperature remains 37 DEG C -45 DEG C, electric pulse parameter setting
Are as follows: peak value 1-2kV/cm, 100 μ s of pulsewidth, frequency 1-4Hz, pulse number 4-10；
With CO in the pretreatment chamber two2Atmosphere is filled up completely, and temperature remains 40 DEG C -45 DEG C, electric pulse parameter setting
Are as follows: peak value 2-5kV/cm, 50 μ s of pulsewidth, frequency 4-8Hz, pulse number 6-10.
The pretreatment chamber one, pretreatment chamber two use CO2Atmosphere is filled up completely, outside air when preventing pretreatment
Into causing to pollute；Pretreatment chamber one, pretreatment chamber two are using low-frequency pulse to glioma stem cell and glioma
Non-stem cell carries out invertibity breakdown, to promote the transmission of glioma stem cell and glioma non-stem cell cell membrane
Property, achieve the purpose that promote glioma stem cell and the expansion of glioma non-stem cell.Pretreatment chamber one, pre- place simultaneously
The temperature control in room two is managed, glioma stem cell and glioma non-stem cell cell membrane can be promoted to change, added
Fast glioma stem cell and glioma non-stem cell expansion process.
Since glioma stem cell is more much larger than glioma non-stem cell volume, itself expansion space is limited,
Therefore pulse frequency will be lower than pulse frequency in pretreatment chamber two in pretreatment chamber one of the present invention.
Pretreated glioma stem cell and glioma non-stem cell are mixed and are placed in lysis process chamber
Pulse cracking and synchronous centrifugal treating are carried out in one；
Supernatant is abandoned, pretreated glioma stem cell and glioma non-stem cell are mixed to be placed in and split
It solves and carries out pulse cracking and synchronous centrifugal treating 4.5-10min in process chamber one；
With CO in the lysis process chamber one2Atmosphere is filled up completely, temperature setting are as follows: rises 1 DEG C to 45 from 37 DEG C of every 30s
℃；
Electric pulse parameter setting are as follows: 20-250V, pulse width 1-20ps, the finger of repetition rate 10Hz-1kHz Independent adjustable
Number decaying waveform, pulse steepness, that is, rise time 90-180ns；
The parameter of noncentricity are as follows: 1000r/min rises to 5000r/min, completes in 5min.
Similarly, with CO in lysis process chamber one2Atmosphere is filled up completely, for avoiding in cracking process by outside contamination；
And using stepped temperature hoisting way can be stable promotion expansion after glioma stem cell, glioma it is non-dry
The metamorphosis of cell film.
Lysis process chamber one using steep-sided pulse to after expansion glioma stem cell and glioma non-stem cell
Irreversible electrical breakdown is carried out, the quick released antigen substance of glioma stem cell can be promoted, while irreversible pulse can
To inactivate polluted bacteria, pollution sources are thoroughly eliminated.
When carrying out steep-sided pulse cracking, centrifugal treating is synchronous to be carried out, this operation can with accelerans glioma stem cells and
The cracking of glioma non-stem cell so that antigenic substance is sufficiently discharged and is merged, and be centrifuged synchronous carry out can be to prevent
Only when pulse breakdown bubble accumulation, occur gas breakdown cause device to damage.
Centrifugal rotational speed rises to 5000r/min by 1000r/min simultaneously, completes in 5min, this is arranged so that revolving speed is steady
It is promoted, is conducive to keep the uniformity in cracking process.
Supernatant is abandoned, above treated the glioma stem cell and glioma non-stem cell mixed liquor are placed
The cracking of pulse again, i.e. the harvest full tumour antigen of glioma are carried out in lysis process chamber two.
Based on the above technical solution, the present invention can also be improved as follows.
Further, supernatant is abandoned, above treated the glioma stem cell and glioma non-stem cell are mixed
Conjunction liquid, which is placed in lysis process chamber two, carries out the cracking 20-30min of pulse again, until pH value fluctuating range is less than in mixed liquor
0.05；
Wherein, with CO in the lysis process chamber two2Atmosphere is filled up completely, and temperature remains 37 DEG C；
Electric pulse parameter setting are as follows: peak value 1-2kV/cm, 100 μ s of pulsewidth, frequency 1-4Hz, pulse number 4-10.
Again low is carried out to high frequency steep-sided pulse treated mixed liquor using low-frequency pulse in the lysis process chamber two
Frequency handle, it is therefore an objective to so that in mixed liquor antigenic information further release.Since cell cracking h substance can cause to mix
Liquid pH numerical value changes, therefore the present invention is to determine that mixed liquor cracking is referring to foundation with pH value fluctuating range in mixed liquor
It is no cmpletely.
The present invention solves the above technical problem and also provides a kind of full tumour antigen preparation facilities of glioma, comprising: micro-
Type controller and the power module being connect with the microcontroller, control module, preprocessing module, detection feedback module,
Processing module is cracked, the preprocessing module is connected with pretreatment chamber one, pretreatment chamber two, and the detection feedback module is connected with
Temperature detection sensor group, pH detection sensor, total reflection air-foam detector, the cracking processing module are connected with cracking processing
Room one, lysis process chamber two, centrifuge, temperature controller, vacuumize process device, the lysis process chamber one be fixedly installed on from
On heart device.
The microcontroller is as present apparatus control centre, for the coordinated control of whole device, the power module
For the power supply of the present apparatus, operation of the control module for whole device is controlled, and the preprocessing module is for mind
Expansion process and sterilization processing through glioma stem cells and glioma non-stem cell, the cracking processing module is for mind
Cracking and antigen preparation through glioma stem cells and glioma non-stem cell.
The lysis process chamber one is fixedly installed on centrifuge, synchronous with centrifugation for realizing the cracking of high frequency steep-sided pulse
Processing is conducive to the acceleration cracking and the generation of bubble breakdown of glioma stem cell and glioma non-stem cell.
Further, the pretreatment chamber one, pretreatment chamber two, lysis process chamber one, in lysis process chamber two on set respectively
It is equipped with impulse generator, the impulse generator is provided with porous nano electrode, and the porous nano electrode is set to pretreatment
Room one, pretreatment chamber two, lysis process chamber one, inside lysis process chamber.
Generation of the impulse generator for period low frequency or high frequency steep-sided pulse, the porous nano electrode is due to porous
Body structure surface product is larger, and the electric field radiation range generated is wider, is conducive to the promotion of efficiency for pre-processing and cracking processing.
Further, the pretreatment chamber one, pretreatment chamber two, lysis process chamber one, be respectively arranged in lysis process chamber
Temperature sensor.The temperature sensor is used for pretreatment chamber one, pretreatment chamber two, lysis process chamber one, lysis process chamber two
The acquisition of middle temperature data, and will acquisition data be sent to detection feedback module amplify, filter, the processing such as numerical value conversion,
It is finally sent to microcontroller to be analyzed, and temperature real-time control is carried out by temperature controller.
Further, the total reflection air-foam detector is set to inside lysis process chamber one, and the pH detection sensor is set
It is placed in inside lysis process chamber two, due to being easy to generate ionized gas, the total reflection bubble detection during high frequency breakdown
Device utilizes total reflection principle to acquire for the data to bubble yield in lysis process chamber one, and acquisition data transmission is extremely examined
Survey feedback module amplify, filter, the processing such as numerical value conversion, be finally sent to microcontroller and analyzed, if bubble stream
Amount reaches preset value, and microcontroller starting vacuumize process device carries out pumping pressurized treatments, so that gas in cracking mixed liquor
It is excluded.Acquisition of the pH detection sensor for pH numerical value change in lysis process chamber two, and data transmission will be acquired extremely
Detection feedback module amplifies, filters, the processing such as numerical value conversion, is finally sent to microcontroller and is analyzed, if cracking
Mixed liquor pH value fluctuating range is less than 0.05pH in process chamber two, and microcontroller determines that entire cracking is completed.
Further, the vacuumize process device is connected to by tracheae with lysis process chamber one, for realizing lysis process chamber
Pumping process in one.
The beneficial effects of the present invention are:
The present invention realizes glioma stem cell and glioma non-stem cell point using low frequency and high frequency steep-sided pulse
Phase process and cracking, easy to operate, cracking more sufficiently, quickly, can get more complete glioma tumour antigen.
Detailed description of the invention
A kind of full tumour antigen preparation facilities structure chart of glioma of Fig. 1 present invention.
Specific embodiment
Principles and features of the present invention are described below in conjunction with example, the given examples are served only to explain the present invention, and
It is non-to be used to limit the scope of the invention.
The preparation of the full tumour antigen of glioma
Embodiment 1
Fresh Human Brain Gliomas is cleaned into blood stains with buffer, cuts into lmm3The samples of human glioma block of size is standby
With；It is unicellular and carry out Multiplying culture glioma will to be obtained in glioma tumor tissue；Neuroglia after proliferation
Matter tumor is unicellular to isolate glioma stem cell and glioma non-stem cell；Using serum-free medium to the mind
The Multiplying culture carried out through glioma stem cells and glioma non-stem cell, and obtain the glioma of logarithmic growth phase
Stem cell and glioma non-stem cell；By number than for the glioma stem cell of 1:2 and glioma it is non-dry thin
Born of the same parents, which are respectively placed in pretreatment chamber one and pretreatment chamber two, carries out cracking pretreatment 5-10min；In the pretreatment chamber one with
CO2Atmosphere is filled up completely, and temperature remains 37 DEG C, electric pulse parameter setting are as follows: peak value 1kV/cm, 100 μ s of pulsewidth, frequency 1Hz,
Pulse number 10；With CO in the pretreatment chamber two2Atmosphere is filled up completely, and temperature remains 40 DEG C, electric pulse parameter setting are as follows:
Peak value 2kV/cm, 50 μ s of pulsewidth, frequency 8Hz, pulse number 10.Supernatant is abandoned, by pretreated glioma stem cell
It is placed in lysis process chamber one with the mixing of glioma non-stem cell and carries out pulse cracking and synchronous centrifugal treating 4.5-
10min；With CO in the lysis process chamber one2Atmosphere is filled up completely, temperature setting are as follows: rises 1 DEG C to 45 from 37 DEG C of every 30s
℃；Electric pulse parameter setting are as follows: 250V, pulse width 20ps, the exponential damping waveform of repetition rate 1kHz Independent adjustable, arteries and veins
Rush steepness i.e. rise time 180ns；The parameter of noncentricity are as follows: 1000r/min rises to 5000r/min, completes in 5min.
Embodiment 2
Tumor experiment is killed in experiment one in vitro
Target cell (neuroglial cytoma) is inoculated in 96 orifice plates (three wells), uses silk after cultivating 100h in incubator
Rimocidin C processing.The effector cell after co-culturing is taken to be added in 96 orifice plates by effect target ratio for 1:1,5:1,10:1,20:1,40:l,
It is incubated so that lml is added after KRP buffered containing 37 DEG C of-thymidine of 0.5 μ C/ml [methyl -3H] after cultivating 20h in incubator
It educates 12-18 hours, 10 stopped reactions is got express developed with the PBS of the glucose containing 10mmol/L of pre-cooling.Cell pyrolysis liquid is taken,
Gamma-5500 liquid flashing counting measures umber of pulse (cpm) per minute.
According to formula killing rate (%)=[target control group cpm mono- (effect mono- effect control group cpm of target group cpm)/target control group
Cpm] × 100% calculating killing rate.
As a result: experimental group has stronger killing ability compared with control group.When imitating target ratio 60:1, experimental group vaccine can be killed
86.91% scholar, 6.79% glioma non-stem cell like cell and 80.82% scholar, 6.36% stem cell-like cell, and control group
It is only capable of killing 25.7% scholar, 7.25% glioma non-stem cell like cell and 50.15% scholar, 4.69% stem cell-like cell.
Embodiment 3
Tumor experiment is killed in experiment two in vivo
Experimental procedure: tumor-bearing mice is randomly divided into three groups: experimental group, control group and blank group, blank group root of the tail portion skin
Lower injecting normal saline；Existing glioma therapeutic vaccine (10 is subcutaneously injected in control group root of the tail portion5A/mL, 0.3mL)；
The glioma therapeutic vaccine (10 in the inventive embodiments 1 is subcutaneously injected in experimental group root of the tail portion5A/mL, 0.3mL) epidemic disease
Seedling；Observe the life cycle of each group mouse in 50d.
As a result: find through analysis life cycle: the median survival interval of each experimental group rat is respectively as follows: control group 18d days, real
Testing group is 33d blank control group 6d.
Experimental animal peripheral blood IFN-γ testing result: each group rat peripheral blood IFN-γ concentration is respectively as follows: existing vaccine
120.12 scholar 2.13pg/ml of group, 181.76 scholar 6.95pg/ml of experimental group, blank group is 78.98 scholar 5.74pg/ml.
The foregoing is merely presently preferred embodiments of the present invention, is not intended to limit the invention, it is all in spirit of the invention and
Within principle, any modification, equivalent replacement, improvement and so on be should all be included in the protection scope of the present invention.
Claims (7)
1. a kind of full tumour antigen preparation method of glioma, which comprises the steps of:
Fresh Human Brain Gliomas is cleaned into blood stains with buffer, cuts into lmm3The samples of human glioma block of size is spare；
It is unicellular and carry out Multiplying culture glioma will to be obtained in glioma tumor tissue；
Glioma after proliferation is unicellular to isolate glioma stem cell and glioma non-stem cell；
The Multiplying culture that the glioma stem cell and glioma non-stem cell are carried out using serum-free medium,
And obtain glioma stem cell and the glioma non-stem cell of logarithmic growth phase；
The glioma stem cell of logarithmic growth phase and glioma non-stem cell are placed in pretreatment chamber one, pretreatment
It is pre-processed in room two；
By number than for 1:2-1:5 glioma stem cell and glioma non-stem cell be respectively placed in pretreatment chamber
One and pretreatment chamber two in carry out cracking pretreatment 5-10min；
With CO in the pretreatment chamber one2Atmosphere is filled up completely, and temperature remains 37 DEG C -45 DEG C, electric pulse parameter setting are as follows: peak
Value 1-2kV/cm, 100 μ s of pulsewidth, frequency 1-4Hz, pulse number 4-10；
With CO in the pretreatment chamber two2Atmosphere is filled up completely, and temperature remains 40 DEG C -45 DEG C, electric pulse parameter setting are as follows: peak
Value 2-5kV/cm, 50 μ s of pulsewidth, frequency 4-8Hz, pulse number 6-10；
Pretreated glioma stem cell and glioma non-stem cell are mixed and are placed in lysis process chamber one
Carry out pulse cracking and synchronous centrifugal treating；
Supernatant is abandoned, pretreated glioma stem cell and glioma non-stem cell are mixed and are placed at cracking
It manages and carries out pulse cracking and synchronous centrifugal treating 4.5-10min in room one；
With CO in the lysis process chamber one2Atmosphere is filled up completely, temperature setting are as follows: rises 1 DEG C to 45 DEG C from 37 DEG C of every 30s；
Electric pulse parameter setting are as follows: the index of 20-250V, pulse width 1-20ps, repetition rate 10Hz-1kHz Independent adjustable decline
Cut waveform, pulse steepness, that is, rise time 90-180ns；
The parameter of noncentricity are as follows: 1000r/min rises to 5000r/min, completes in 5min；
Supernatant is abandoned, above treated the glioma stem cell and glioma non-stem cell mixed liquor are placed in and are split
The cracking of pulse again, i.e. the harvest full tumour antigen of glioma are carried out in solution process chamber two.
2. the full tumour antigen preparation method of a kind of glioma according to claim 1, which is characterized in that the step is abandoned
Above treated the glioma stem cell and glioma non-stem cell mixed liquor are placed in cracking processing by supernatant
The cracking of pulse again is carried out in room two, that is, harvests being implemented as follows for the full tumour antigen of glioma:
Supernatant is abandoned, above treated the glioma stem cell and glioma non-stem cell mixed liquor are placed in and are split
Solution process chamber two in carry out pulse again cracking 20-30min, until in mixed liquor pH value fluctuating range less than 0.05；
Wherein, with CO in the lysis process chamber two2Atmosphere is filled up completely, and temperature remains 37 DEG C；
Electric pulse parameter setting are as follows: peak value 1-2kV/cm, 100 μ s of pulsewidth, frequency 1-4Hz, pulse number 4-10.
3. a kind of full tumour antigen preparation facilities of glioma characterized by comprising microcontroller and with it is described micro-
The power module of type controller connection, control module, preprocessing module, detection feedback module, cracking processing module, the pre- place
Reason module is connected with pretreatment chamber one, pretreatment chamber two, and the detection feedback module is connected with temperature detection sensor group, pH inspection
Survey sensor, total reflection air-foam detector, the cracking processing module be connected with lysis process chamber one, lysis process chamber two, from
Heart device, temperature controller, vacuumize process device, the lysis process chamber one are fixedly installed on centrifuge.
4. the full tumour antigen preparation facilities of a kind of glioma according to claim 3, which is characterized in that the pretreatment
Room one, pretreatment chamber two, lysis process chamber one, in lysis process chamber two on be respectively arranged with impulse generator, the pulse hair
Raw device is provided with porous nano electrode, and the porous nano electrode is set to pretreatment chamber one, pretreatment chamber two, lysis process chamber
One, inside lysis process chamber.
5. the full tumour antigen preparation facilities of a kind of glioma according to claim 3, which is characterized in that the pretreatment
Room one, lysis process chamber one, is respectively arranged with temperature sensor in lysis process chamber two at pretreatment chamber two.
6. the full tumour antigen preparation facilities of a kind of glioma according to claim 3, which is characterized in that the total reflection
Air-foam detector is set to inside lysis process chamber one, and the pH detection sensor is set to inside lysis process chamber two.
7. the full tumour antigen preparation facilities of a kind of glioma according to claim 3, which is characterized in that described to vacuumize
Processor is connected to by tracheae with lysis process chamber one.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710166858.9A CN106939298B (en) | 2017-03-20 | 2017-03-20 | A kind of full tumour antigen preparation method of glioma and device |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710166858.9A CN106939298B (en) | 2017-03-20 | 2017-03-20 | A kind of full tumour antigen preparation method of glioma and device |
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| Publication Number | Publication Date |
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| CN106939298A CN106939298A (en) | 2017-07-11 |
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101085391A (en) * | 2007-06-29 | 2007-12-12 | 重庆大学 | Device for inducing tumor cell apoptosis by high-voltage nanosecond pulse |
| CN105055015A (en) * | 2015-08-05 | 2015-11-18 | 天津市鹰泰利安康医疗科技有限责任公司 | Irreversible electroporation system |
| CN105233279A (en) * | 2015-10-10 | 2016-01-13 | 深圳爱生再生医学科技有限公司 | Glioma holoantigen and preparation method and application thereof |
| CN105233280A (en) * | 2015-10-10 | 2016-01-13 | 深圳爱生再生医学科技有限公司 | DC-based glioma holoantigen vaccine and preparation method thereof |
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Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101085391A (en) * | 2007-06-29 | 2007-12-12 | 重庆大学 | Device for inducing tumor cell apoptosis by high-voltage nanosecond pulse |
| CN105055015A (en) * | 2015-08-05 | 2015-11-18 | 天津市鹰泰利安康医疗科技有限责任公司 | Irreversible electroporation system |
| CN105233279A (en) * | 2015-10-10 | 2016-01-13 | 深圳爱生再生医学科技有限公司 | Glioma holoantigen and preparation method and application thereof |
| CN105233280A (en) * | 2015-10-10 | 2016-01-13 | 深圳爱生再生医学科技有限公司 | DC-based glioma holoantigen vaccine and preparation method thereof |
Non-Patent Citations (1)
| Title |
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| 陡脉冲对恶性肿瘤细胞不可逆性电击穿的实验研究;姚陈果 等;《中国生物医学工程学报》;20040229;第23卷(第1期);第92-97页 |
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