CN106544392A - A kind of screening technique of histamine H 1 receptor antagonist - Google Patents
A kind of screening technique of histamine H 1 receptor antagonist Download PDFInfo
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Abstract
The present invention by monitoring the DMR response signals caused after histamine acts on A-431 cells or A-549 cells using n cell target spot pharmacological techniques, the DMR response signals caused after A-431 cells or A-549 cells are acted on histamine again desensitize in dose dependent, high expression H1 receptors on A-431 cells or A-549 cells are proved, and the DMR response signals caused after receptor agonism can be successfully monitored.DMR response signals are not caused with specificity histamine H 1 receptor antagonist pretreatment A-431 cells or A-549 cells, but the DMR response signals of histamine after histamine are added again by dose-dependently antagonism.Thus it is successfully established histamine H 1 receptor antagonist screening technique：Compound pretreatment A-431 cells to be screened or the DMR response signals caused by A-549 cells are monitored first, if compound to be screened does not cause DMR response signals, and add the DMR response signals of histamine after histamine by antagonism again, antagonistic activity of the compound to H1 receptors is judged by antagonism degree according to response signal.
Description
Technical field
The invention belongs to drug screening research field, and in particular to a kind of screening side of histamine H 1 receptor antagonist
Method.
Background technology
Histamine is generally present in mastocyte and basophilic granulocyte with its non-activated state, when by wound and
After the environmental stimulis such as antigen antibody reaction, extracellular, participation inflammatory reaction is discharged in the form of activating[1]。
Histamine is played its physiological action by combining with histamine receptor.Histamine receptor in human body can be divided into 4
Type：H1, H2, H3 and H4 receptor.This 4 type histamine receptor is g protein coupled receptor
(G-protein-coupled receptors, GPCRs), by G-protein transducer cell external signal to cell
The interior secondary signaling system[1,2].The alterative inflammation effect of histamine mediation causes skin mainly in conjunction with H1 receptors
The hyperemia of skin mucosa, edema and prurituss etc., in the case that dosage is larger, cause smooth muscle contraction, whole body capillary
Vasodilation and permeability increase, and show as asthma, anaphylactic shock, tachycardia, erubescence and nose
Hyperemia etc.[3].H1 receptor antagonists are by competitively tying with H1 receptors with H1 receptor bindings and blocking histamine
Close, so as to suppress histamine to play biological effect[4].This is the main machine that medicine plays anti-allergy action
System.The histamine H 1 receptor antagonist for clinically using is antiallergic and antipruritic important drugs, but generally existing
The side effect such as CNS inhibition, cholinolytic, cardiac toxicity, increased weight[5].Therefore, it is necessary to set up one kind
The high-throughput screening method of histamine H 1 receptor antagonist, so as to rapid effective for histamine H 1 receptor antagonist
New drug development provide lead compound.
The features such as n cell target spot pharmacological techniques have high sensitivity, unmarked and fanout free region, which is former
Reason is will by waveguide optical grating (resonant wave guide grating, the RWG) biosensor that resonates
Medicine acts on after membrane receptor that dynamic material resets (dynamic mass in trigger cell
Redistribution, DMR) signal is transformed into the wavelength shift signal of light[6-7].It has been widely used in receptor
The research of biology[8-9]With the discovery of medicine[10-11]。
In consideration of it, the present invention is based on n cell target spot pharmacological techniques, histamine H1-receptor antagonism is established
The new method of agent screening, high-flux fast screening histamine H 1 receptor antagonist.
List of references：
MacGlashan D Jr.Histamine:A mediator of inflammation.J Allergy Clin
Immunol,2003,112(4Suppl):S53-9.
Hill SJ,Ganellin CR,Timmerman H,et al.International Union of
Pharmacology.XIII.Classification of histamine receptors.Pharmacol
Rev,1997,49(3):253-78.
Simons FE.Advances in H1-antihistamines.N Engl J Med,2004,351(21):
2203-17.
Simons FE,Simons KJ.The pharmacology and use of
H1-receptor-antagonist drugs.N Engl J Med,1994,330(23):1663-70.
Simons FE,Simons KJ.Histamine and H1-antihistamines:celebrating
a century of progress.J Allergy Clin Immunol.2011,128(6):1139-50.
Fang Y,Ferrie AM,Fontaine NH,et al.Resonant waveguide grating
biosensor for living cell sensing.Biophys J,2006,91(5):1925-40.
Kenakin TP.Cellular assays as portals to seven-transmembrane
receptor-based drug discovery.Nat Rev Drug Discov,2009,8(8):
617-26.
Sun H,Wei Y,Deng H,et al.Label-free cell phenotypic profiling
decodes the composition and signaling of an endogenous ATP-sensitive
potassium channel.Sci Rep,2014,12(4):4934.
Deng H,Hu H,Fang Y.Multiple tyrosine metabolites are GPR35agonists.
Sci Rep,2012,2:373.
Fang Y.Label-free cell-based assays with optical biosensors in drug
discovery.Assay Drug Dev Technol,2006,4(5):583-95.
Fang Y.Guest Editor's Introduction:Label-free optical biosensors
to cell biology and drug discovery.J Recept Signal Transduct Res,
2009,29(3-4):127.
The content of the invention
It is an object of the invention to provide a kind of be based on n cell target spot pharmacological techniques, set up histamine H 1 and receive
The new method of body antagonist screening, high-flux fast screening histamine H 1 receptor antagonist.
The technical scheme for being adopted for：
The present invention relates to a kind of screening technique of histamine H 1 receptor antagonist, the present invention is by using unmarked thin
The DMR responses that born of the same parents' target spot pharmacological techniques monitoring histamine causes after acting on A-431 cells or A-549 cells
Signal, the DMR response signals caused after acting on A-431 cells or A-549 cells with histamine again are in agent
Amount dependency desensitization, it was demonstrated that high expression H1 receptors on A-431 cells or A-549 cells, and after receptor agonism
The DMR response signals for causing can be successfully monitored.With specificity histamine H 1 receptor antagonist pretreatment A-431
Cell or A-549 cells do not cause DMR response signals, but add the DMR response letters of histamine after histamine again
Number by dose-dependently antagonism.Thus it is successfully established histamine H 1 receptor antagonist screening technique：Monitor first
DMR response signals caused by compound pretreatment A-431 cells to be screened or A-549 cells, if treated
SCREENED COMPOUND does not cause DMR response signals, and adds the DMR response signals of histamine after histamine short of money again
It is anti-, antagonistic activity of the compound to H1 receptors is judged by antagonism degree according to response signal.
What the present invention was caused after monitoring H receptor stimulating agent histamine to act on A-431 cells or A-549 cells
DMR response signals, it is found that histamine can cause DMR response signals on A-431 cells or A-549 cells, and
In dose dependent；After with the histamine pretreatment A-431 cells or A-549 cells of variable concentrations, add again
The histamine of fixed concentration, the caused DMR response signals of monitoring find that the concentration of histamine during pretreatment is higher
It is more obvious that second step adds the DMR response signals caused by histamine to be desensitized, in dose dependent.A Simi
Azoles, Loratadine are specificity histamine H 1 receptor antagonist, with the astemizole or Loratadine of variable concentrations
After pretreatment A-431 cells or A-549 cells, the histamine of fixed concentration added again, and monitoring is caused
DMR response signals, find astemizole and Loratadine difference dose-dependently caused by antagonizing histamine
DMR response signals.
Histamine H1-receptor is expressed in A-431 cells and A-549 cells, therefore the above results prompting, using nothing
Labelling cell target spot pharmacological techniques can successfully monitor on A-431 cells or A-549 cells H1 receptors by histamine
Histamine is caused to desensitize after DMR response signals and Agonists of Histamine Receptor or antagonist pretreatment caused by excitement
Or the DMR response signals of antagonism.
Thus it is successfully established histamine H 1 receptor antagonist screening technique：Compound pretreatment to be screened is monitored first
Caused DMR response signals after A-431 cells or A-549 cells, if compound to be screened does not cause DMR
Response signal, then continuously add histamine, whether monitors the DMR response signals caused by histamine by antagonism, sentences
Antagonistic activity of the seco compound to histamine H1-receptor.
A-431 cells of the present invention refer to people's epidermis cancerous cell；
A-549 cells of the present invention refer to Non-small cell lung carcinoma cell.
The histamine H 1 receptor antagonist screening technique comprises the steps,
A. cell culture：
A-431 cells are with containing 10% hyclone, 100U/ml ampicillin, 100 μ g/ml sulfate chains
The DMEM culture medium of mycin, in 37 DEG C, 5%CO2Incubator in cultivate；
A-549 cells are with containing 10% hyclone, 100U/ml ampicillin, 100 μ g/ml sulfate chains
The F12K culture medium of mycin, in 37 DEG C, 5%CO2Incubator in cultivate；
B. A-431 cells or A-549 cells that step A is cultivated are harvested, 384 hole biological inductors are inoculated into
In minitype plate, 37 DEG C, 5%CO are placed in2Incubator in cultivate 24～48h；
C. the A-431 cells of step B culture continue with the culture medium Nature enemy 16～24 without hyclone
H, the A-549 cells of step B culture are not required to Nature enemy；
D. the cell of step C culture is washed 2 times with HBSS buffer, is added HBSS buffer, is placed in high pass
1～2h of balance incubation on amount, unmarked screening system；
E. set up the baseline of a 2min after balancing first on high flux, unmarked screening system, then
Compound to be screened described in 10 μ L is added, continues 0.5～1h of monitoring, the histamine for being subsequently adding 10 μ L continues
0.5～1h of monitoring；
F. the maximum DMR responses drafting amount effect relation curve that caused by compound pretreatment dosage and histamine, in terms of
Calculate the IC of the compound to be screened50Value, the comparison compound to be screened are lived to the antagonism of histamine H1-receptor
Property.
In step B, the inoculum density of A-431 cells or A-549 cells is 2 × 104～2.5 × 104Individual/hole.
Another aspect of the present invention is that above-mentioned histamine H 1 receptor antagonist screening technique is in histamine receptor antagonism
Purposes in agent screening.Specifically, it is, using said method Jing high flux screenings, to find with antagonizing histamine
The high activity drug candidate of H1 receptor actives.
The present invention advantage and is had the beneficial effect that compared with other histamine H 1 receptor antagonist screening techniques：
The present invention adopts n cell target spot pharmacological techniques, acts on work by monitoring probe molecule/medicine
After cell, the dynamic change procedure of trigger cell inner material to be carrying out the screening of histamine H 1 receptor antagonist, with
Other histamine H 1 receptor antagonist screening techniques are compared, and n cell target spot pharmacological method has high flux
(can be determined using 384 hole biological inductor minitype plates), high intension (can reflect pharmaceutically-active multidimensional category
Property), the characteristic of highly sensitive (response signal that medicine causes under nM concentration can be monitored), reflection
Be drug molecule stimulate cell to cause ligand binding, receptor activation, albumen to raise, receptor internalisation etc. and cause
Living cells internal dynamic material redistribution (dynamic mass redistribution, DMR), analysis
Be multi signal path comprehensive effect and non-orphaned mono signal transduction.The method set up by the present invention is not only
The screening of histamine H 1 receptor antagonist can be carried out, moreover it is possible in target specificity, effectiveness and binding mode (i.e.
Agonism, antagonism or reverse excitement effect) in terms of carry out molecule pharmacology it is qualitative.
Description of the drawings
The amount effect relation curve of the DMR signals that Fig. 1 a. histamine causes on A-431 cells；Histamine locates agent in advance
The amount effect relation curve of the DMR signals that amount is caused with 8 μM of histamine：
Histamine can cause DMR response signals on A-431 cells, and in dose dependent, EC50For 1.29
μM；So that the histamine of fixed concentration, the DMR response signals of histamine are added after the histamine pretreatment of variable concentrations
Desensitized, during pretreatment, the DMR response signals of the higher second step histamine of the concentration of histamine are desensitized more obvious, are in
Dose dependent, IC50For 0.59 μM.
The amount effect relation curve of the DMR signals that Fig. 1 b. histamine causes on A-549 cells；Histamine locates agent in advance
The amount effect relation curve of the DMR signals that amount is caused with 8 μM of histamine：
Histamine can cause DMR response signals on A-549 cells, and in dose dependent, EC50For 1.77
μM；So that the histamine of fixed concentration, the DMR response signals of histamine are added after the histamine pretreatment of variable concentrations
Desensitized, during pretreatment, the DMR response signals of the higher second step histamine of the concentration of histamine are desensitized more obvious, are in
Dose dependent, IC50For 0.84 μM.
Fig. 2 a. histamine H 1 receptor antagonists on A-431 cells locate the DMR letters that dosage and 16 μM of histamine cause in advance
Number amount effect relation curve：
Astemizole, Loratadine are histamine H 1 receptor antagonist, with the astemizole of variable concentrations or chlorine thunder
After he processes A-431 cells surely, add 16 μM of histamine, the DMR response signals that histamine causes by antagonism,
And the concentration of antagonist more high antagonistic is stronger, in dose dependent, the antagonistic activity Ah of two kinds of antagonisies
Department imidazoles (IC50=0.01 μM) it is better than Loratadine (IC50=0.18 μM).
Fig. 2 b. histamine H 1 receptor antagonists on A-549 cells locate the DMR letters that dosage and 8 μM of histamine cause in advance
Number amount effect relation curve：
Astemizole, Loratadine are histamine H 1 receptor antagonist, with the astemizole of variable concentrations or chlorine thunder
After he processes A-549 cells surely, add 8 μM of histamine, the DMR response signals that histamine causes by antagonism, and
The concentration of antagonist more high antagonistic is stronger, in dose dependent, the antagonistic activity Ah Si of two kinds of antagonisies
Imidazoles (IC50=0.04 μM) it is better than Loratadine (IC50=0.25 μM).
Fig. 3 a. DMR signals that aretigenin pretreatment dosage and 16 μM of histamine cause on A-431 cells
Amount effect relation curve：
With aretigenin (64 μM of highest working concentration, 2 times of dilutions, 7 Concentraton gradient) pretreatment A-431
After cell, 16 μM of histamine (EC are added80), the DMR response signals that histamine causes are by antagonism, and are in dosage
Dependency.
Fig. 3 b. DMR signals that aretigenin pretreatment dosage and 8 μM of histamine cause on A-549 cells
Amount effect relation curve：
After on aretigenin (64 μM, 32 μM) pretreatment A-549 cells, 8 μM of histamine are added,
The DMR response signals that histamine causes are by antagonism, and are in dose dependent.
Specific embodiment
Following non-limiting examples can make one of ordinary skill in the art that the present invention is more fully understood,
But the present invention is limited never in any form.The present invention is made further in the way of specific embodiment below
It is bright.
Various chemical reagent used in the embodiment of the present invention if no special instructions, by routine business way
Footpath obtains, but is not intended to limit the scope of method claimed by the present invention.Histamine, astemizole, Loratadine
Purchased from Sigma companies；DMSO is bought from Bio Basic companies；HBSS and the purchase of McCoy ' s 5A culture fluid
From Gibco companies；384 hole biological inductor minitype plates are purchased from Corning companies；A-431 people's table
Skin cancer cell, A-549 Non-small cell lung carcinomas cell are from American Type Culture Collection committee of Chinese Academy of Sciences cell
Storehouse obtains (China, Shanghai).
The foundation of 1 histamine H 1 receptor antagonist screening technique of embodiment
1. method
1.1 cell culture
A-431 people's epidermis cancerous cell, A-549 Non-small cell lung carcinomas cell are from Chinese Academy of Sciences's type culture
Preservation committee cell bank obtains (China, Shanghai).A-431 cells are with containing 10% hyclone, 100U/ml
Ampicillin, (GIBCO, article No. 12800017, contains the DMEM culture medium of 100 μ g/ml streptomycin sulfates
D-Glucose 4.5g/L, add NaHCO31.5g/L), in 37 DEG C, 5%CO2Incubator in cultivate；
A-549 cells are with containing 10% hyclone, 100U/ml ampicillin, 100 μ g/ml streptomycin sulfates
F12K culture medium, in 37 DEG C, the incubator of 5%CO2 cultivate.
1.2 n cell target spot pharmacological experiments
A-431 cells or A-549 cells are with 2 × 104The density in individual/hole is inoculated into384 hole biological responses
In device minitype plate, 37 DEG C, 5%CO are placed in2Incubator in cultivate 24h.Training of the A-431 cells without FBS
Foster base Nature enemy 24h, A-549 cells are not required to Nature enemy.Washed 2 times with HBSS buffer before detection, then
30 μ L HBSS buffer are added in every hole, is placed inBalance incubation 1h in system.After balance first
The baseline of a 2min is set up in system, 10 μ L testing compounds are subsequently adding, continues monitoring 30
Min, the histamine for being subsequently adding 10 μ L continue monitoring 30min.By monitoring the DMR response signals that histamine causes
Whether pretreated the testing compound that adds when suppressed judging that compound is lived to the antagonism of histamine H1-receptor
Property.
1.3 data analysiss
DMR data are recorded by Epic Imager softwares (Corning Incorporated, the U.S.), and Jing Imager Beta 3.7
Software (Corning Incorporated, the U.S.) process is obtained.Using Microsoft Excel 2010 and GraphPad Prism
Software carries out statistical analysiss.All of DMR signals all Jing blank corrections are obtained.The EC of body part description50With
IC50Obtained based on maximum DMR signal of change in compound 30min is added.All data from 2 independent experiments,
4 repetitions every time.
2. result
The dose-effect relationship of the DMR response signals that 2.1 histamine cause
The dose-effect relationship of the DMR response signals that 2.1.1 histamine causes on A-431 cells
Histamine (128 μM of highest working concentration, 2 times of dilutions, 13 Concentraton gradient) is on A-431 cells
DMR response signals can be caused, and in dose dependent, EC50For 1.29 μM；With above-mentioned Concentraton gradient
8 μM of histamine is added after histamine pretreatment A-431 cells, caused DMR response signals are monitored,
It was found that the higher second step of the concentration of histamine adds the DMR response signals caused by histamine to be desensitized more during pretreatment
Substantially, in dose dependent, IC50For 0.59 μM (Fig. 1 a).There is histamine on this explanation A-431 cell
Receptor, combined with histamine it is rear DMR response signals are caused by excitement, and after adding histamine again, histamine swashs
Action is with being desensitized.
The dose-effect relationship of the DMR response signals that 2.1.2 histamine causes on A-549 cells
Histamine (128 μM of highest working concentration, 2 times of dilutions, 13 Concentraton gradient) is on A-549 cells
DMR response signals can be caused, and in dose dependent, EC50For 1.77 μM；With above-mentioned Concentraton gradient
8 μM of histamine is added after histamine pretreatment A-549 cells, caused DMR response signals are monitored,
It was found that the higher second step of the concentration of histamine adds the DMR response signals caused by histamine to be desensitized more during pretreatment
Substantially, in dose dependent, IC50For 0.84 μM (Fig. 1 b).There is histamine on this explanation A-549 cell
Receptor, combined with histamine it is rear DMR response signals are caused by excitement, and after adding histamine again, histamine swashs
Action is with being desensitized.
The impact of the DMR response signals that 2.2 histamine H 1 receptor antagonists cause to histamine
The impact of the DMR response signals that 2.2.1 histamine H 1 receptor antagonist causes to histamine on A-431 cells
Astemizole, Loratadine are histamine H 1 receptor antagonist, and with variable concentrations, (highest work is dense respectively
Degree 16 μM, 2 times dilution, 19 Concentraton gradient) astemizole or Loratadine process A-431 cells
Afterwards, 16 μM of histamine (EC are added80), the DMR response signals that histamine causes are suppressed, and antagonist
The higher inhibitory activity of concentration it is stronger, in dose dependent, the inhibitory activity astemizole of two kinds of antagonisies
(IC50=0.01 μM) it is better than Loratadine (IC50=0.18 μM), see Fig. 2 a.This explanation H1 is received
Body antagonist can the DMR response signals that cause of antagonizing histamine, H1 receptors can be set up based on this method short of money
Anti-agent screening model, for the high flux screening of H1 receptor antagonists.
The impact of the DMR response signals that 2.2.2 histamine H 1 receptor antagonist causes to histamine on A-549 cells
Astemizole, Loratadine are histamine H 1 receptor antagonist, and with variable concentrations, (highest work is dense respectively
Degree be respectively 8 μM and 64 μM, 2 times dilute, 14 Concentraton gradient) astemizole or Loratadine at
After reason A-549 cells, 8 μM of histamine are added, the DMR response signals that histamine causes are suppressed, and short of money
The higher inhibitory activity of concentration of anti-agent is stronger, in dose dependent, inhibitory activity Ah 's miaow of two kinds of antagonisies
Azoles (IC50=0.04 μM) it is better than Loratadine (IC50=0.25 μM), see Fig. 2 b.This explanation H1
Receptor antagonist can the DMR response signals that cause of antagonizing histamine, H1 receptors can be set up based on this method
Antagonist screening model, for the high flux screening of H1 receptor antagonists.
The application of 2 histamine H 1 receptor antagonist screening technique of embodiment
The histamine H1-receptor antagonism of 40 testing compounds is lived using n cell target spot pharmacological techniques
Property is screened.First, by 10 μ L testing compounds (64 μM of highest working concentration, 2 times dilution,
2 Concentraton gradient) add and be inoculated with A-431 cells or A-549 cells384 hole biological inductors
In minitype plate,30min is monitored in system, (8 μM) of 10 μM of histamine is subsequently adding and is continued prison
30min is surveyed, the maximum DMR response signals caused in 30min after histamine are added by antagonism journey according to second step
Spend to judge testing compound to histamine H1-receptor antagonistic activity, wherein aretigenin shows that H1 receptors are short of money
Resistant activity.With aretigenin (64 μM of highest working concentration, 2 times of dilutions, 7 Concentraton gradient) pretreatment
After A-431 cells, 16 μM of histamine (EC are added80), the DMR response signals that histamine causes are suppressed,
And be in dose dependent (Fig. 3 a).With aretigenin (64 μM, 32 μM) pretreatment A-549 cells
Afterwards, 8 μM of histamine are added, the DMR response signals that histamine causes are suppressed, and (the figure in dose dependent
3b).May infer that aretigenin may be histamine H 1 receptor antagonist by the above results.This research is based on nothing
Labelling cell target spot pharmacological techniques, have been successfully established the high-throughput screening method of histamine H 1 receptor antagonist.
Claims (4)
1. a kind of screening technique of histamine H 1 receptor antagonist, it is characterised in that：The histamine H1-receptor is short of money
The screening technique of anti-agent adopts n cell target spot pharmacological techniques high flux screening histamine H1-receptor antagonism
Agent；
People's epidermis cancerous cell (A-431 cells) or people's non-small cell of compound pretreatment to be screened are monitored first
Lung carcinoma cell (A-549 cells) DMR response signals caused afterwards, if compound to be screened does not cause DMR
Response signal, then continuously add histamine, whether monitors the DMR response signals caused by histamine by antagonism, sentences
Break antagonistic activity of the compound to be screened to histamine H1-receptor.
2. screening technique according to claim 1, it is characterised in that：
The histamine H 1 receptor antagonist screening technique comprises the steps,
A. cell culture：
A-431 cells are with containing 10% hyclone, 100U/ml ampicillin, 100 μ g/ml sulphuric acid
The DMEM culture medium of streptomycin, in 37 DEG C, 5%CO2Incubator in cultivate；
A-549 cells are with containing 10% hyclone, 100U/ml ampicillin, 100 μ g/ml sulphuric acid
The F12K culture medium of streptomycin, in 37 DEG C, 5%CO2Incubator in cultivate；
B. A-431 cells or A-549 cells that step A is cultivated are harvested, 384 hole biological inductors are inoculated into
In minitype plate, 37 DEG C, 5%CO are placed in2Incubator in cultivate 24～48h；
C. the A-431 cells of step B culture continue with the culture medium Nature enemy 16～24 without hyclone
H, the A-549 cells of step B culture are not required to Nature enemy；
D. the cell of step C culture is washed 2 times with HBSS buffer, is added HBSS buffer, is placed in high pass
1～2h of balance incubation on amount, unmarked screening system；
E. set up the baseline of a 2min after balancing first on high flux, unmarked screening system, then
Compound to be screened described in 10 μ L is added, continues 0.5～1h of monitoring, the histamine for being subsequently adding 10 μ L continues
0.5～1h of monitoring；
F. the maximum DMR responses drafting amount effect relation curve that caused by compound pretreatment dosage and histamine, in terms of
Calculate the IC of the compound to be screened50Value, the comparison compound to be screened are lived to the antagonism of histamine H1-receptor
Property.
3. screening technique according to claim 2, it is characterised in that：In step B A-431 cells or
The inoculum density of A-549 cells is 2 × 104～2.5 × 104Individual/hole.
4. screening technique according to claim 2, it is characterised in that：The high flux, unmarked sieve
System is selected to be healthy and free from worrySystem.
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| CN113005089A (en) * | 2019-12-20 | 2021-06-22 | 泰州医药城国科化物生物医药科技有限公司 | Cell screening model of unlabeled cell membrane receptor GPR88 and application |
| CN113604557A (en) * | 2021-08-04 | 2021-11-05 | 杭州浙大迪迅生物基因工程有限公司 | Human histamine receptor HRH1mRNA detection primer probe set, kit and application |
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