CN103756968B - Spinal highly-metastatic human lung adenocarcinoma cell strain and construction process thereof and application - Google Patents
Spinal highly-metastatic human lung adenocarcinoma cell strain and construction process thereof and application Download PDFInfo
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Abstract
The invention discloses a kind of Spinal highly-metastatic human lung adenocarcinoma cell strain A549L6, this cell strain is preserved in China typical culture collection center (being called for short CCTCC) on April 25th, 2013, does is preserving number CCTCC? No:C201346.The invention also discloses preparation method and the application of described Spinal highly-metastatic human lung adenocarcinoma cell strain.The backbone rate of transform of Spinal highly-metastatic human lung adenocarcinoma cell strain of the present invention is high, can be widely used in adenocarcinoma of lung Bone tumour research model and in study of lung gland cancer metastasis and screening and the application prepared in anti-metastasis drug.
Description
Technical field
The present invention relates to oncobiology technical field and microorganism animal cell line technical field, be specifically related to a kind of Spinal highly-metastatic human lung adenocarcinoma cell strain and construction process thereof and application.
Background technology
Cancer is the number one killer of current threat human health, and according to World Health Organization's incomplete statistics, the patient that cancer is died from the whole world every year reaches more than 700 ten thousand people.Lung cancer is one of cancer that current whole world M & M is the highest.The U.S. every year nearly 170000 people dies from lung cancer, accounts for 30% [1] of whole cancer mortality sum.Along with rapid growth economic in recent years, the sickness rate of China's lung cancer starts to be multiplied, and about has 400,000 people to be made a definite diagnosis every year and suffers from lung cancer, and sickness rate, up to 61.4/10 ten thousand, becomes the country that patients with lung cancer is in the world maximum.Expect 2025, the number dying from lung cancer is often only by China will close to 1,000,000 people.
The same with other malignant tumours, the transfer of lung cancer is the first cause [2] causing Endodontic failure and death.Because the not obvious and disease progression speed of lung cancer early symptom is very fast, the patient of 40% is about had to occur distant metastasis [3] to time medical.Bone is one of the most common metastasis site of lung cancer, and Bone tumour [3] appears in the progressive stage lung cancer of about 30-40%.
Backbone is one of the most common Bone tumour position of lung cancer, accounts for 50% [4] of Bone tumour.The complication such as the transfer of lung cancer backbone often causes that ostalgia, hypercalcemia, pathologic fracture, spinal compression are even paralysed, have a strong impact on life quality and the lifetime of patient, accelerate the dead process of patient, bring heavy burden [5,6] to patient and family thereof and society.Although the complex therapy such as surgical excision, chemicotherapy, Diphosphonate, immunotherapy of code requirement clinically, the median survival interval of lung cancer backbone transfer was still hovered in about half a year, became a great problem that medical circle is urgently to be resolved hurrily.
In recent years, biomedical sector achieves the development of advancing by leaps and bounds, but still understands very few to lung cancer backbone transfer generation, development mechanism, lacks correlative study.Its major cause is to lack reliable lung cancer backbone transfer animal model.Therefore, be necessary to set up a kind of meet Clinical symptoms, reproducible, can real-time tracing monitoring, reliable and stable lung cancer backbone transfer animal model.To cancer backbone transfer molecular mechanism, clinical diagnosis and treatment will have great promoter action, thering is provided research tool for finding new drug targets further and developing corresponding medicine, bringing more wide prospect by giving the treatment of lung cancer and Bone tumour.
A549 cell strain (ATCC CCL-185), within 1972, the cancerous lung tissue of the white male of 58 years old is adopted to set up through vitro culture by Giard etc., be one of the most widely used lung adenocarcinoma cell line [7], the A549 cell strain that the present invention adopts ATCC to originate builds lung cancer backbone metastasis model as source cell.
The background technology relevant with the present invention comprises below with reference to document.
[1]DelaCruzCS，TanoueLT，MatthayRA.Lungcancer：epidemiology,etiology,andprevention.ClinChestMed.2011Dec；32(4)：605-44.
[2]PantelK，Alix-PanabièresC.Circulatingtumourcellsincancerpatients：challengesandperspectives.TrendsMolMed.2010Sep；16(9)：398-406.Epub2010Jul29。
[3]McErleanA，GinsbergMS.Epidemiologyoflungcancer.SeminRoentgenol[J].2011Jul；46(3)：l73-7.
[4]TsuyaA，KurataT，TamuraK，FukuokaM.Skeletalmetastasesinnon-smallcelllungcancer：aretrospectivestudy.LungCancer.2007Aug；57(2)：229-32
[5]ColemanRE.Skeletalcomplicationsofmalignancy.Cancer1997；80：1588-94.
[6] Xiao Jianru. backbone metastatic carcinoma Surgical Strategy is familiar with [J] again. JISUI ZAZHI AUTHOR AND, 2011,21 (7): 531-532.
[7]GiardDJ，AaronsonSA，TodaroGJ，etal：Invitrocultivationofhumantumors：establishmentofcelllinesderivedfromaseriesofsolidtumors.JNatlCancerInst51：1417-1423，1973
[8]KangY，SiegelPM，ShuW，etal.Amultigenicprogrammediatingbreastcancermetastasistobone.CancerCell.2003Jun；3(6)：537-49.
Summary of the invention
The invention discloses a kind of Spinal highly-metastatic human lung adenocarcinoma cell strain and construction process thereof and application.
An object of the present invention is to provide a kind of Spinal highly-metastatic human lung adenocarcinoma cell strain, this cell strain is deposited in the China typical culture collection center (being called for short CCTCC) being positioned at Wuhan on April 25th, 2013, preserving number is CCTCCNo:C201346.
The present invention, using human A459 lung cancer cell line cell strain (ATCC CCL-185) as source cell, by the method for nude mice Vivo Studies on Screening, builds the human lung adenocarcinoma cell line that a strain has the organ such as backbone, brain height transfer ability, called after A549L6.
The detection method that detection backbone high-transfer cell strain A549L6 biological characteristics adopts is with following step B.Nude mice left ventricle injection A549L6 cell, and use IVIS In vivo detection system and the image detection equipment such as x-ray, micro-CT, MRI to monitor its transfer performance, and use HE pathological section to confirm, adopt the experiment in vitro such as woundhealing, transwell to detect its migrate attribute, adopt cell proliferation experiment to detect its multiplication characteristic in vitro.
Compared with source cell strain A549L0, Spinal highly-metastatic human lung adenocarcinoma cell strain A549L6 cell strain of the present invention has following biological characteristics:
(1) detect A549L6 cell strain through woundhealing and transwell, observe 36 hours, the ability of migrating of A549L6 cell is 11.5 times and 22.4 times of source cell A549L0 respectively; Experimentation on animals shows its backbone rate of transform up to 80% (8/10), significantly improves compared with the backbone rate of transform (7.7%, 1/13) of source cell A549L0.
(2) the distant metastasis of human rate after the injection of A549L6 cell strain nude mice left ventricle is 100%, and 70% shifts for multiple organ simultaneously.Bone tumour rate is 90% (9/10), and the backbone rate of transform is 80% (8/10), and the four limbs rate of transform is 30% (3/10); Brain metastes rate is 60% (6/10), and wherein, the median time of backbone transfer is 50 days (40-106).
(3) Spinal highly-metastatic human lung adenocarcinoma cell strain A549L6 cell strain of the present invention marks through luciferase, by living imaging instrument the equipment such as growth in vivo of IVIS live body Real-Time Monitoring tumour, transfer case, and within 6.00 ± 0.63 days, backbone transfer signal detected early than x-ray test set.
(4) Micro-CT, MRI all successfully can detect A549L6 backbone metastatic lesion.Backbone metastasis can be detected through x-ray, micro-CT, MRI after described cell strain transfer, backbone and brain metastes stove can be detected through HE pathological section.
(5), after A549L6 is transferred to backbone, has after shifting to human cancer backbone and the similar symptom of spinal compression occurs.It can be divided into successively five obvious observable nodes according to the development of the course of disease: tail mops floor, instep lands gait, walking step state of sweeping the floor, paralysis, death.It betides 1.43 ± 1.72 days, 3.71 ± 2.06 days, 8.29 ± 2.06 days, 11.57 ± 3.69 days, 14.29 ± 3.59 days after living imaging system discovery backbone metastasis respectively.
(6) after A549L6 is transferred to backbone, its body weight significantly declines, and when injecting A549L6 cell with left ventricle, tail mops floor, instep lands gait, walking step state of sweeping the floor, paralysis, dead corresponding body weight are respectively 17.84 ± 0.83g, 26.20 ± 1.85g, 24.96 ± 1.52g, 22.78 ± 1.44g, 21.02 ± 1.72g, 18.76 ± 1.73g.
The nutrient solution of cell strain of the present invention, go down to posterity, frozen, recovery condition is all identical with A549, in-vitro multiplication ability is similar to A549; There is notable difference in cellular form and the A549L0 of A549L6, A549L0 and A549 plesiomorphism, major part is the cell clone of light transmission difference, and only containing the cell clone that partial light permeability is strong, and A549L6 (under Figure 13) is almost the strong cell clone of light transmission.
Another aspect of the present invention is to provide the construction process of described Spinal highly-metastatic human lung adenocarcinoma cell strain A549L6, comprises the steps:
(A) human A459 lung cancer cell line cell is inoculated in culture plate, carries out PGL4 plasmid transfection, luciferase assays is carried out to the cell clone obtained through G418 screening after transfection, obtains the cell clone A549L0 of the luciferase expression positive;
(B) A549 expressing luciferase positive cell clone A549L0 suspension is injected in nude mice left ventricle;
(C) screen backbone transfer cell strain, the destruction of bone of selection backbone metastasis is the strongest and fluorescence is the strongest metastatic tumour cell carries out amplification in vitro cultivation; Be injected in nude mice left ventricle by cultivating the cell obtained by step (B) method, then carry out amplification in vitro cultivation by step (C); Repetition three-wheel screens;
(D) the A549 cell of fluorescent mark positive step (C) obtained carries out mono-clonal and selects, and amplification in vitro is cultivated, and obtains backbone high-transfer human lung adenocarcinoma cell cell line A549 L6 of the present invention.
Concrete steps comprise as follows:
Steps A, luciferase (Luciferase) mark human A459 lung cancer cell line
By human A459 lung cancer cell line (ATCC CCL-185) cell according to 1.0 × 10
5/ hole is inoculated in 24 well culture plates, carries out PGL4 plasmid (PromegaBiotechCo.Ltd) transfection next day; Transfection procedure and method are carried out according to Lipofectamin2000 reagent specification sheets; Change liquid after transfection 24h, add 600ug/mlG418 (purchased from Merck company) and screen three weeks, obtain positive cell clone; And luciferase (Luciferase) fluorescence activity mensuration is carried out to the clone obtained.
Step B, left ventricle injecting method set up human lung adenocarcinoma Metastatic nude model
Build the method [8] of bone metastases of lung cancer model foundation with reference to Kang etc., by the A549 cell (called after A549L0) of luciferase (Luciferase) the fluorescent mark positive, be adjusted to 1.0 × 10
5the cell suspension of/100ul, gets 100ul and is injected in nude mice left ventricle, and uses the monitoring adenocarcinoma of lung backbone transfer case such as IVIS living imaging system and x-ray imaging system.
Step C, screening backbone transfer cell strain
Get the strongest and metastatic tumour cell that fluorescence is the strongest of backbone metastasis destruction of bone through vitro culture, amplification, culture condition is that DMEM adds 10% (V/V) foetal calf serum, 37 DEG C, 5%CO
2/ 95% air, saturated humidity is cultivated; The cell of cultivation gained is inoculated in the recycle system of nude mice according to step B, according to step C amplification in vitro, repeats three-wheel screening with method.
Step D, mono-clonal are selected
The A549 cell of the fluorescent mark positive of being taken out by step C carries out mono-clonal and selects, and amplification in vitro, sets up cell strain, called after A549L6, changes nutrient solution every other day, within every three days, goes down to posterity once, conventional cryopreservation, recovery, obtain Spinal highly-metastatic human lung adenocarcinoma cell strain A549L6.
Another aspect of the present invention is to provide described Spinal highly-metastatic human lung adenocarcinoma cell strain A549L6 shifts animal model application at the backbone preparing human lung adenocarcinoma.The preparation process of aforementioned backbone high-transfer human Adenocarcinoma of lung cell line A549 L6 is the application of the present invention in the backbone transfer animal model of human lung adenocarcinoma builds.Present invention also offers the application of described Spinal highly-metastatic human lung adenocarcinoma cell strain A549L6 in bone metastases of lung cancer Mechanism Study and its application at the anti-human adenocarcinoma of lung medicine of preparation and its in screening and the application assessed in anti-human adenocarcinoma of lung medicine.
Spinal highly-metastatic human lung adenocarcinoma cell strain A549L6 of the present invention has the high feature of the backbone rate of transform, be one more preferably adenocarcinoma of lung Bone tumour research model, it will have wide prospect in the application of the metastasis and anti-metastasis drug screening that disclose adenocarcinoma of lung.
Accompanying drawing explanation
Backbone high-transfer human lung adenocarcinoma cell line A549 L6 of the present invention, be preserved in China typical culture collection center (being called for short CCTCC, China, Wuhan, Wuhan University) on April 25th, 2013, preserving number is CCTCCNo:C201346.
Fig. 1 represents the luciferase expression of A549L0 and A549L6.Through IVIS living imaging systems axiol-ogy, the equal stably express fluorescence of A549L0 and A549L6, after multi-turns screen, the luciferase expression value of A549L6 is significantly higher than A549L0.
Fig. 2 represents the In vivo detection result that lung cancer backbone shifts.In vivo detection result shows 100% mouse generation distant metastasis of human, and mostly is many places organ and shifts simultaneously, and the mouse generation backbone transfer of 80%, 60% brain metastes occurs.
Fig. 3 represents that x-ray imaging system detects the X-ray film image that backbone transfer occurs A549L6.
Fig. 4 represents that micro-CT detects CT imaging performance and the three-dimensional reconstruction image that backbone transfer occurs A549L6.
Fig. 5 represents that the MRI image of backbone transfer occurs A549L6.
Fig. 6 represents A549L6 backbone metastasis histology profile section HE staining pathologic section figure.
Fig. 7 represents A549L6 backbone metastasis histological cross sections section HE staining pathologic section figure.
Fig. 8 represents that spinal compression symptom schematic diagram occurs the transfer of A549L6 backbone, along with the Node Events that PD occurs that tail mops floor, instep lands gait successively, walking step state of sweeping the floor, paralysis, death etc. can be easy to observation.
Fig. 9 represents that spinal compression symptom schematic diagram occurs the transfer of A549L6 backbone, along with PD occur that tail mops floor, instep lands gait successively, walking step state of sweeping the floor, paralysis, death etc. can be easy to the Node Events observed, along with PD, also there is noticeable change in its body weight thereupon.
Figure 10 represents that the cell migration ability of A549L6 and A549L0 is compared in woundhealing experiment.A left side is schematic diagram, and the right side is statistical graph.
Figure 11 represents that the cell migration ability of A549L6 and A549L0 is compared in Transwell experiment.A left side is schematic diagram, and the right side is statistical graph.
Figure 12 represents that A549L6 and A549L0 and A549 multiplication capacity is in vitro similar.
The cellular form that Figure 13 represents A549L6 and A549L0, A549 has notable difference, and A549L6 is almost the strong cell clone of light transmission, and A549L0, A549 are all only containing the cell clone that partial light permeability is strong.
Embodiment
In conjunction with following specific embodiments and the drawings, the present invention is described in further detail, and protection content of the present invention is not limited to following examples.Under the spirit and scope not deviating from inventive concept, the change that those skilled in the art can expect and advantage are all included in the present invention, and with appending claims and equivalent thereof for protection domain.Implement process of the present invention, condition, reagent, experimental technique etc., mention specially except explanation except following, be universal knowledege and the common practise of this area, the present invention is not particularly limited content.Describe the present invention below in conjunction with embodiment and accompanying drawing.But the following example should not regard limitation of the scope of the invention as.
The structure of embodiment 1: Spinal highly-metastatic human lung adenocarcinoma cell strain A549L6
By human A459 lung cancer cell line cell strain (ATCC CCL-185) cell according to 1.0 × 10
5/ hole is inoculated in 24 well culture plates, carries out PGL4 plasmid (PromegaBiotechCo.Ltd) transfection next day; Transfection procedure and method are carried out according to Lipofectamin2000 reagent specification sheets; Change liquid after transfection 24h, add 600ug/mlG418 (purchased from German merck company) and screen three weeks, obtain positive cell clone; And luciferase (Luciferase) fluorescence activity mensuration is carried out to the clone obtained.
By human A459 lung cancer cell line cell suspension 0.1ml (containing 1x10
5individual cell) be injected in nude mice left ventricle, and use IVIS living imaging system and x-ray imaging system monitoring adenocarcinoma of lung backbone transfer case.Nude mice is purchased from (Shanghai western pul-Bi Kai laboratory animal company limited), and product are nu/nu nude mice, male, 4-6 week age, raise in SPF level environment.
After cell is inoculated in nude mice blood circulation by the method that left ventricle is injected, anatomy experiment animal, get the strongest and metastatic tumour cell that fluorescence is the strongest of backbone metastasis destruction of bone through vitro culture, amplification, nutrient solution is that DMEM (gibico) adds 10% (v/v) foetal calf serum (Hyclone, Utah, USA), 37 DEG C, 5%CO
2/ 95% air, saturated humidity is cultivated.
The cell of cultivation gained is inoculated in the recycle system of nude mice according to above-mentioned steps, according to this step amplification in vitro, repeats three-wheel screening with method.
The A549 cell of the fluorescent mark positive of above-mentioned steps being taken out carries out mono-clonal and selects, and amplification in vitro, sets up cell strain, called after A549L6, changes nutrient solution every other day, within every three days, goes down to posterity once, conventional cryopreservation, recovery, obtain Spinal highly-metastatic human lung adenocarcinoma cell strain A549L6.
Embodiment 2: the Characteristics Detection of Spinal highly-metastatic human lung adenocarcinoma cell strain A549L6 of the present invention
Serial experiment builds to embodiment 1 the cell line A549 L6 of the present invention obtained and detects routinely:
1, by after centrifugal for cell routine trysinization, density is adjusted to 1x10
6/ ml, gets 100ul/ hole and adds black 96 orifice plate, carry out doubling dilution successively, and each gradient establishes 2 multiple holes.Every hole adds the luciferase substrate 100ul of 300 μ g/ml, within 10 minutes, be placed on IVIS living imaging system photographs (CaliperLifeSciencesIVISLuminaII, model: DS934N-BV-286), time shutter is 5 minutes, shows the equal stably express luciferase of A549L0 and A549L6 after testing.As shown in Fig. 1 left side, after too much taking turns Vivo Studies on Screening, when cell count is 1x10
6time, A549L6 luciferase expression value is approximately 5.512 times of A549L0; When cell count dilution is 5x10
4time A549L0 almost without macroscopic fluorescence, and A549L6 is until lower than 1.25x10
4the macroscopic fluorescence of Shi Caiwu.Fig. 1 right side is depicted as the luciferase expression value curve of A549L6 under each cell density condition.As seen from Figure 1, through IVIS living imaging systems axiol-ogy, the equal stably express fluorescence of A549L0 and A549L6, after multi-turns screen, the luciferase expression value of A549L6 is significantly higher than A549L0.
2,1x10 is got
5cell (0.1ml) is injected in nude mice left ventricle, within latter 25 days, starts the In vivo detection carrying out the transfer of lung cancer backbone in injection.Method is: abdominal injection injected fluorescein enzyme substrates 200ul/ is (D-luciferin only, potassiumsalt, 15mg/ml, sigma), within 10 minutes, be placed on IVIS living imaging system photographs (CaliperLifeSciencesIVISLuminaII, model: DS934N-BV-286), the time shutter is 5 minutes.As shown in Figure 2, the distant metastasis ability of cell line A549 L6 of the present invention comparatively A549L0 (distant metastasis of human rate is 7.7%) significantly improves, overall Distant metastasis rates is 100%, mostly be bone (Bone tumour rate is 90%) and brain (brain metastes rate is 60%) shifts simultaneously, and mostly be many places organ and shift simultaneously.Wherein, the backbone rate of transform is 80%, and the four limbs rate of transform is 30%, and the meta number of days that backbone transfer occurs is 50 days (40-106 days).
3, through animalcule x-ray test set (KodakDXS4000ProSystem, model: MicroFocusX-RayImagingSource, 35KVP, the parameter that instrument uses is Bringing:4XBringing, f-stop:2.51, Fov:59.4mm, Resolution:220ppi, CCDtemperature:-29 DEG C, time shutter is 1 minute) monitoring, 6.00 ± 0.63 days after IVIS detects cell line A549 L6 backbone metastasis signal of the present invention, x-ray test set can be utilized the metastasis of mouse backbone to be detected.As shown in Figure 3, Fig. 3 is the next corresponding x-ray image of IVIS living imaging figure, Fig. 3.
4, animalcule CT (Skyscan1076,40Kv, 250uA is used, BrukermicroCTNV, Antwerp, Belgium) and 3.0TMRI (MagnetomVerio, Siemens, Germany) effectively can detect the destruction situation of A549L6 backbone metastasis.As shown in Figure 4, Fig. 4 upper left represents that the centrum of mouse is obviously destroyed by tumour, and upper right is normal control; Be corresponding three-dimensional reconstruction figure under Fig. 4.Fig. 5 right side is respectively the coronal-plane of backbone metastasis, sagittal plane, transverse section T2 weighted mri imaging, and left side is respectively corresponding normal control.
5, find that A549L6 backbone transfer characteristic is similar to the clinical picture that lung cancer backbone shifts through HE pathological section, as shown in Figure 6, representing the whole centrum of tumor cell invasion, but do not invade intervertebral disk under Fig. 6, Fig. 6 is normal control.As shown in Figure 7, representing under Fig. 7 that whole centrum is tumor cell invasion, Fig. 7 is normal control.
6, the spinal compression symptom by observing mouse every day finds: the development along with the course of disease also can occur and clinical similar spinal compression symptom, along with PD occur that tail mops floor, instep lands gait successively, walking step state of sweeping the floor, paralysis, death etc. can be easy to the Node Events (shown in Fig. 8) observed, betide 1.43 ± 1.72 days, 3.71 ± 2.06 days, 8.29 ± 2.06 days, 11.57 ± 3.69 days, 14.29 ± 3.59 days after living imaging system discovery backbone metastasis respectively.
7, through finding the monitoring of Mouse Weight: after A549L6 is transferred to backbone, its body weight significantly declines, and when injecting A549L6 cell with left ventricle, tail mops floor, instep lands gait, walking step state of sweeping the floor, paralysis, dead corresponding body weight are respectively 17.84 ± 0.83g, 26.20 ± 1.85g, 24.96 ± 1.52g, 22.78 ± 1.44g, 21.02 ± 1.72g, 18.76 ± 1.73g.(as shown in Figure 9).
8, through woundhealing (by tumor cell inoculation in 12 orifice plates, when tumour cell 90% is polymerized, hungry 12 hours, use aseptic pipettor gun head along culture plate center cut, then in the migration situation of different timing node observation of cell), after 27 hours and 36 hours, the ability of migrating of A549L6 cell is 5.5 times and 11.5 times of source cell respectively, as shown in Figure 10, wherein, left figure is schematic diagram, and right figure is statistical graph.
9, through transwell, (tumour cell is resuspended in the DMEM of serum-free, concentration adjustment is 50,000/200ul, gets the upper strata that 200ul is inoculated in the cell that shuttles back and forth, and inserts the DMEM substratum 600ul containing 10% foetal calf serum in cell below.4% paraformaldehyde is used to fix at different time points, violet staining, and the tumour cell of statistics migration of taking pictures under the microscope) detect, after 36 hours, the ability of migrating of A549L6 cell is 22.4 times of source cell, as shown in figure 11, wherein, left figure is schematic diagram, and right figure is statistical graph.
10, A549L6 nutrient solution, go down to posterity, frozen, recovery condition is all identical with A549, in-vitro multiplication experiment (by cell with 1x10
4the density in/hole is inoculated in 6 orifice plates, respectively after inoculation the 2nd, 4,6,8,10 day is by cell dissociation, counting) to show its multiplication capacity similar to A549L0 and A549, as shown in figure 12, show that A549L6 and A549L0 and A549 multiplication capacity is in vitro similar.There is notable difference in cellular form and the A549L0 of A549L6, the cellular form that Figure 13 shows A549L6 and A549L0, A549 there is notable difference, A549L6 is almost the strong cell clone of light transmission, and A549L0, A549 are all only containing the cell clone that partial light permeability is strong.As represented the similar cell clone all only containing the strong cell clone of partial light permeability and light transmission difference of A549L0 and A549 on Figure 13 with in Figure 13, and A549L6 (under Figure 13) is almost the strong cell clone of light transmission.
Claims (2)
1. a Spinal highly-metastatic human lung adenocarcinoma cell strain, is characterized in that, described cell strain is preserved in China typical culture collection center, and preserving number is CCTCCNo:C201346, and the preservation time is on April 25th, 2013.
2. the application of Spinal highly-metastatic human lung adenocarcinoma cell strain as claimed in claim 1 in the anti-human adenocarcinoma of lung medicine of preparation.
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