CN102928415A - Magnetic particle chemiluminescence kit for detecting estradiol, and applications thereof - Google Patents
Magnetic particle chemiluminescence kit for detecting estradiol, and applications thereof Download PDFInfo
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- CN102928415A CN102928415A CN 201110227587 CN201110227587A CN102928415A CN 102928415 A CN102928415 A CN 102928415A CN 201110227587 CN201110227587 CN 201110227587 CN 201110227587 A CN201110227587 A CN 201110227587A CN 102928415 A CN102928415 A CN 102928415A
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Abstract
The present invention relates to a magnetic particle chemiluminescence kit for detecting estradiol. The kit comprises a luminous marker, a fluorescein marker, a standard substance, a quality control substance and a separation reagent, wherein the luminous marker is isoluminol luminous marker labeled estradiol hapten, the fluorescein marker is fluorescein labeled estradiol monoclonal antibody or fluorescein derivative labeled estradiol monoclonal antibody, and the separation reagent is sheep anti-FITC monoclonal antibody coated paramagnetic nanometer microbeads. The present invention further relates to a method for detecting estradiol in foods of animal origin by using the kit, wherein the method provides characteristics of high sensitivity, high specificity and rapid detection for estradiol detection.
Description
Technical field
The present invention relates to a kind of chemiluminescence detection kit and method of testing thereof, be specifically related to a kind of magnetic granule chemoluminescence detection kit that detects estradiol in the samples such as animal tissue, feed, urine.
Background technology
Estradiol (Estradiol, E2) is widely used steroid class anabolic hormone, by a large amount of growths that are used for promoting the ruminant animal, because it can promote animal and improve livestock and poultry lean meat ratio, once uses in brutish aquaculture.But since estradiol can be in animal tissue high residue, healthy by the food chain harm humans, be that Female Children is grown in advance, the male children mammogenesis is womanlike; Male reproductive system dysplasia or cause pathology; And cause women with breast cancer and endometriosis incidence to rise, and there is obvious carcinogenicity in estradiol, and is American-European etc., and developed country forbids in succession or strictly ban use of.The regulation estradiol must not detect in feed in No. 226 file of the State Council of the PRC, " feed and feed addictive management rules ".
At present, the detection method of estradiol common are high performance liquid chromatography (HPLC), GC-MS (GC-MS) and euzymelinked immunosorbent assay (ELISA) (ELISA) etc.
1, high performance liquid chromatography (HPLC) has accuracy of detection height, characteristics that false positive rate is low.The major defect of HPLC method is that instrument is expensive, operate more loaded down with trivial details, length consuming time, testing cost is expensive, and testing staff's specialty is had relatively high expectations, Sample pretreatment is complicated.
2, gas chromatography-mass spectrography (GC-MS) sensitivity is very high, and false positive rate is low.The major defect of the method is that sample needs derivatization to process complicated like this early stage.Although can provide more structural information through the sample of derivatization treatment in mass spectrum, derivatization can produce a plurality of different products and cause the partial loss of sample, causes the deviation of experimental result.
3, euzymelinked immunosorbent assay (ELISA) (ELISA) is that 20 century 70s occur, be used for the detection of micro substance, be applied to the earliest the clinical detection such as infectious disease, tumor markers, hormonal readiness, begin the nineties to apply in China food safety detection field, relying at present the kit of elisa technique, the leading products that test strips has become food security fast detecting field, also is simultaneously the major product type that my company researches and develops and sells.The enzyme linked immunosorbent detection method is based on the specific reaction of Ag-Ab, detection sensitivity is higher, specificity is better, technical operation is simple and easy, grasp easily, really solved the qualitative and quantitative analysis work of a large amount of former insoluble many micro-small-molecule substances such as microbiotic, hormone, residues of pesticides etc., positive facilitation has been played in the development of food security Fast Detection Technique.But there are many defectives that self can't overcome in the ELISA method, and is mainly manifested in:
1) heterogeneous reaction: be separated free thing and bond in the testing process, need multistep to wash the plate process, time and effort consuming is difficult to improve the automaticity of operation.
2) enzymic catalytic reaction: by substrate for enzymatic activity colour developing, assaying reaction liquid absorbance.There are considerable influence in time and the temperature of reaction to enzyme activity, reagent stability.
Summary of the invention
Technical matters to be solved by this invention is to provide a kind of magnetic granule chemiluminescence kit that detects the estradiol medicine, not only possesses higher sensitivity when adopting this kit to carry out the detection of estradiol medicine, specificity, and have higher reaction velocity.
Another object of the present invention is to provide a kind of method of testing of estradiol medicine, and the method is simple to operate, and is highly sensitive, and specificity is good.
Realize above-mentioned purpose, the invention provides a kind of estradiol drug test kit, its main agents that comprises has: luminous marker, fluorescein-labelled thing, standard items, quality-control product, separation agent.
Described separation agent is the paramagnetism nano microsphere that is coated with goat-anti FITC antibody.
Described paramagnetism nano microsphere, surface contain-OH ,-COOH or-NH
2The microballon of reactive group is used for coated goat-anti FITC monoclonal antibody, and its inner core is Fe
3O
4Or γ-Fe
2O
3, so that microballon has paramagnetism.
Described luminous marker is the estradiol haptens of different Derivative of Luminol mark.Described different Derivative of Luminol is ABEI, AHEI or ABEN.
Described kit also comprises standard items, quality-control product and concentrated washing lotion.
Described fluorescein-labelled thing is FITC mark estradiol monoclonal antibody.
The present invention also provides a kind of method of utilizing kit to detect estradiol, comprises the following steps:
1) draw respectively 20 μ l-100 μ l standard items or samples, then add 20 μ l-100 μ l luminous markers, add again the fluorescein-labelled thing of 20 μ l-100 μ l, behind the abundant mixing, 37 ℃ of incubation 15min;
2) add the paramagnetism nano microsphere 80-150 μ l be coated with goat-anti FITC monoclonal antibody, behind the mixing at 37 ℃ of incubation 5min;
3) separate 5min with the magnetic separator frame, abandon behind the supernatant with cleaning fluid 300-500 μ l flushing compound precipitation;
4) above-mentioned cleaning step repeats 2-4 time;
5) 4) compound of gained separator well is directly put into the measurement camera bellows, add and excite substrate 1 and excite substrate 2, postpone to detect the relative light intensity (RLU) that sends behind the 3-5s, content in the sample and RLU proportion relation can be gathered the concentration that calibration curve method calculates estradiol by RLU.
The principal ingredient of described luminous substrate is NaOH and H
2O
2
The used analyser of analysis test method is among the present invention: comprise power circuit, automatic injection pump 1 and 2, measuring chamber, illuminated chamber, photomultiplier counter and output system, also dispose simultaneously the Windows control software of computing machine and Chinese interface, can carry out that data typing, result gather, quality control, the result stores and the function such as result queries, can finish the programming of multiple analytical model, quantitative or qualitative reporting the result, automatic generation and storage, update functions, automatically revise typical curve at 2, the system that adopts is the CI-2008 system.
Separation agent of the present invention is coated goat-anti FITC monoclonal antibody, the content of its surface group is 0.1-0.3eqm/g, and it is stored in and contains the 3-6% bovine serum albumin(BSA), the 0.2-0.4% Tween-20,0.2% sodium azide is in the phosphate buffer of pH7.0-7.4.Described FITC is fluorescein isothiocynate.Described percentage composition is the quality percentage composition.
Described luminous marker is the estradiol haptens of different Derivative of Luminol ABEI, AHEI or ABEN mark, and it is stored in and contains pH7.2-7.6, and the 0.1-0.3% Tween-20 is in the phosphate buffer of 0.1% sodium azide.Described percentage composition is the quality percentage composition.
The estradiol monoclonal antibody that described fluorescein-labelled thing is the FITC mark, it is stored in pH7.2-7.6, contains the 3-5% bovine serum albumin(BSA), the phosphate buffer of 0.2-0.4% sodium azide.Described percentage composition is the quality percentage composition.
Estradiol standard solution (0ng/ml, 0.03ng/ml, 0.1ng/ml, 0.3ng/ml, 1.0ng/ml, 3.0ng/ml), the standard items dilution is pH7.2,0.3% sodium azide, 0.1mol/L phosphate buffer.Described percentage composition is the quality percentage composition.
Estradiol quality-control product solution concentration is respectively 0.05ng/ml, 2.0ng/ml, quality-control product dilution pH7.2,0.3% sodium azide, 0.1mol/L phosphate buffer.Described percentage composition is the quality percentage composition.
Described concentrated washing lotion is pH7.1-7.5,0.2-0.4% Tween-20,0.2-0.4% sodium azide, 0.1-0.2mol/L phosphate buffer.Described percentage composition is the quality percentage composition.Beneficial effect of the present invention is as follows:
1) sensitivity of kit of the present invention is higher, can reach 0.03ng/ml to the detection sensitivity of estradiol.
2) detection of kit of the present invention is quick, is lower than 20min detection time.
Embodiment
The preparation of the concrete component of embodiment 1 kit
1) estradiol is haptenic synthetic
Estradiol and bromo ethyl butyrate are synthesized the estradiol haptens by reaction.
2) preparation of estradiol artificial antigen
Adopt the water-soluble carbodiimide method to carry out coupling estradiol haptens and hemocyanin and obtain immunogene.
3) preparation of monoclonal antibody
Animal immune: with immunogene the Balb/c mouse is carried out immunity, immunizing dose is 100 μ g/, makes it produce antiserum.
Fusion of Cells and cloning: get immune Balb/c mouse boosting cell, merge in 9: 1 ratios and SP2/0 myeloma cell, obtain the hybridoma cell strain of monoclonal antibody.
Cell cryopreservation and recovery: hybridoma is made 1 * 10 with cryopreserving liquid
9The cell suspension of individual/ml is in the medium-term and long-term preservation of liquid nitrogen.Take out cryopreservation tube during recovery, put into immediately 37 ℃ of water-bath middling speeds and melt, behind the centrifugal removal cryopreserving liquid, move in the culture flask and cultivate.
The preparation and purification of monoclonal antibody: the Balb/c mouse peritoneal is only injected sterilization paraffin oil 0.5ml/, 7 days pneumoretroperitoneum injection hybridomas 5 * 10
7Individual/as only, to gather ascites after 7 days.Carry out purifying, obtain monoclonal antibody with sad-saturated ammonium sulfate method.
4) preparation of luminous marker
Get 4.5mmol/L ABEI, be dissolved in the 4ml distilled water, the 5.0mmol/L N-hydroxy-succinamide is dissolved in the 0.5ml DMF, room temperature reaction 3-4h behind the two abundant mixing.Get the estradiol haptens 15mg of above-mentioned preparation, to 1.5ml, then add the ABEI solution of above-mentioned activation with the pH7.4PBS adjusted volume, fully room temperature reaction spends the night behind the mixing, crosses G-25 gel column purifying.
5) fluorescent marker preparation
Use 0.025mol/L, the carbonate buffer solution of pH9.0 is diluted to 1% mass percent concentration with the estradiol monoclonal antibody; Total protein according to wanting mark adds 0.01mg FITC by every milligram of immunoglobulin (Ig), accurately takes by weighing required FITC powder with analytical balance.With the solution that FITC is made into 0.1mg/ml, by 3-5 times of above-mentioned antibody-solutions volume, sneak into the FITC dilution with same damping fluid; Electromagnetic agitation, mark 30-48h under 4～℃ lucifuge condition; With label solution 3000r/min, the centrifugal 20min of room temperature removes wherein a small amount of sediment, and in the bag filter of packing into, the PBS damping fluid of pH7.4 dialysis is 2-3 days again, during change at least dislysate 3 times; Get the label of dialysed overnight, by SephadexG-25 or G-50 post, the separated free fluorescein is collected the fluorescent marker of mark and is identified that packing is stored in 4 ℃ of refrigerators.
6) separation agent preparation
A) magnetic bead activation
The magnetic bead of surface-COOH group (be purchased from DYNAL, particle diameter is 2.8 μ m), its content is 0.15eq/g; Get 100 μ l magnetic beads, with 100 μ l 25mmol/L, pH5.0,0.05%Tween-20MES solution washing twice, magnetic removes supernatant after separating; Before the use, dispose respectively EDC, the NHS solution of 50mmol/L with the 25mmol/L MES solution of 4 ℃ of storages; Add respectively the new EDC that disposes of 50 μ l and NHS solution in the centrifuge tube that magnetic bead is housed, vortex mixing, room temperature activation 30min; Centrifuge tube places on the magnetic separator frame magnetic to separate 4min, removes supernatant, adds 100 μ l, and 25mmol/L, pH5.0, MES clean and get final product to get the magnetic bead of surperficial activated carboxylic after 2-3 time.
B) magnetic bead coupling goat-anti FITC monoclonal antibody
50-100 μ g goat-anti FITC monoclonal antibody is dissolved into 60 μ l, and 25mmol/L among the pH5.0MES, regulates cumulative volume to 100 μ l with described MES solution, soft mixing magnetic bead and antibody; At least coupling 30min or 4 ℃ of coupling 2h under the room temperature condition can utilize the vortex instrument to make magnetic bead keep the mixing state during this period; Centrifuge tube places magnetic separation 3-5min on the magnetic separator frame, removes supernatant; For cancellation unreacted-COOH, can add 100 μ l, pH7.4, TRIS reaction 15min or 100 μ l, pH8.0 contains the PBS sealing magnetic bead of 50mmol/L monoethanolamine; With 100 μ l, 0.1-0.3%BSA, the PBS of 0.1%Tween-20 or TRIS clean the good magnetic bead of sealing 3-5 time; At last magnetic bead is redissolved in containing 0.1-0.5%BSA, 0.01-0.1%Tween-20,0.02%NaN
3PBS or the TRIS damping fluid in, 2-8 ℃ of preservation.
The establishment of embodiment two kits
Set up the magnetic granule chemoluminescence detection kit that detects estradiol, make it contain following component:
The fluorescent marker of the estradiol monoclonal antibody of FITC mark
The haptenic luminous marker of the estradiol of ABEI mark
The separation agent of the paramagnetism nano microsphere of pan coating goat-anti FITC monoclonal antibody
Estradiol standard solution (0ng/ml, 0.03ng/ml, 0.1ng/ml, 0.3ng/ml, 1.0ng/ml, 3.0ng/ml), the standard items dilution is pH7.2,0.3% sodium azide, 0.1mol/L phosphate buffer.Described percentage composition is the quality percentage composition.
Estradiol quality-control product solution concentration is respectively 0.05ng/ml, 2.0ng/ml, quality-control product dilution pH7.2,0.3% sodium azide, 0.1mol/L phosphate buffer.Described percentage composition is the quality percentage composition.
Concentrated washing lotion is pH7.2,0.3% Tween-20,0.3% sodium azide, 0.2mol/L phosphate buffer.Described percentage composition is the quality percentage composition.
The detection of estradiol in embodiment three actual samples
1, Sample pretreatment
(1) tissue (chicken/liver, pork/liver, shrimp, fish etc.) pre-treating method
With homogenizer homogeneous structure sample; Take by weighing the equal pledge of 2.0 ± 0.05g to 50ml polystyrene centrifuge tube, add 10ml acetonitrile-0.1M sodium hydroxide solution, with the oscillator 10min that vibrates, more than the 3000g, the centrifugal 10min of room temperature (20-25 ℃); Get the 1ml supernatant to the clean glass test tube of 10ml, flow down in 50～60 ℃ of water-bath nitrogen and dry up; Dissolve with the 0.5ml methenyl choloride; Add 2ml 1M sodium hydroxide solution and strip, violent fully vibration 5min, more than the 3000g, the centrifugal 10min of room temperature (20-25 ℃) gets the 1ml supernatant and adds 100 μ l 6M phosphoric acid solution mixings; Add 5ml acetonitrile-methenyl choloride mixed liquor, with the oscillator 10min that vibrates, more than the 3000g, the centrifugal 10min of room temperature (20-25 ℃), go to whole upper stratas, get whole lower floors organic phase to the clean glass test tube of 10ml, flow down in 50～60 ℃ of water-bath nitrogen and dry up; With the dry residue of 1ml redissolution working fluid dissolving; Getting 50 μ l waters is used for analyzing sample extension rate: 10.
(2) urine pre-treating method
Get the 2ml urine in centrifuge tube, more than the 3000g, the centrifugal 10min of room temperature (20-25 ℃) is until limpid; Pipette the limpid urine specimen of 1ml to 50ml polystyrene centrifuge tube, add 10 μ l Glucuronidase/ Arylsulfatase (glucuronidase/aryl sulfatase), at 37 ℃ of hydrolysis 3h; Add the 5ml methenyl choloride with the oscillator 10min that vibrates, more than the 3000g, the centrifugal 10min of room temperature (20-25 ℃); Take off in layer glass test tube that organic phase 1ml to 10ml is clean, flow down in 50～60 ℃ of water-bath nitrogen and dry up; With the dry residue of 1ml redissolution working fluid dissolving; Getting 50 μ l waters is used for analyzing sample extension rate: 5.
(3) feed pre-treating method
Take by weighing 2.0 ± 0.05g feed sample, add the 8ml acetonitrile, with the oscillator 10min that vibrates, more than the 3000g, 15 ℃ of centrifugal 10min; Get the 2ml supernatant to the clean glass test tube of 10ml, flow down in 50～60 ℃ of water-bath nitrogen and dry up; Add the 0.5ml chloroform, with vortex instrument whirling motion 20s, add 2ml 1M sodium hydroxide solution, with vortex instrument whirling motion 30s, more than the 3000g, 15 ℃ of centrifugal 5min; Get 1ml, add 100 μ l 6M phosphoric acid solutions, with vortex instrument whirling motion 10s; Sample solution with redissolve working fluid and dilute (25 μ l sample extraction liquid+975 μ l redissolve working fluid), usefulness vortex instrument whirling motion mixing by 1: 39 volume ratio; Getting 50 μ l waters is used for analyzing sample extension rate: 160.
2, detect and interpretation of result with kit
Draw respectively 20 μ l-100 μ l standard items or samples, then add 20 μ l-100 μ l luminous markers, add again the fluorescein-labelled thing of 20 μ l-100 μ l, behind the abundant mixing, 37 ℃ of incubation 15min; Add the paramagnetism nano microsphere 80-150 μ l be coated with goat-anti FITC monoclonal antibody, behind the mixing at 37 ℃ of incubation 5min; Separate 5min with the magnetic separator frame, abandon behind the supernatant with cleaning fluid 300-500 μ l flushing compound precipitation; The compound of gained separator well is directly put into the measurement camera bellows, add and excite substrate 1 and excite substrate 2, postpone to detect the relative light intensity (RLU) that sends behind the 3-5s, the content of estradiol and RLU proportion relation in the sample can be by the concentration of RLU combined standard curve method calculating estradiol.
Exciting substrate 1 is NaOH, and exciting substrate 2 is H
2O
2Before substrate loads, clean the substrate pump 10-20 time with distilled water, behind the interior remaining water mark of emptying pipe, more corresponding substrate is directly put into instrument, pump line is inserted in the substrate bottle; With substrate flushing pipe 5 times, and measure the RLU value of substrate, under normal circumstances, the RLU value of substrate should be above 1200.If surpass 1200, need again with distilled water pipeline and substrate pump to be carried out more times cleaning, until blank value is down in the zone of reasonableness.Do not do experiment if surpass three days, need unloading substrate bottle, and cover lid, with vaporization prevention.Use subsequently the distilled water pipe blow-through, and emptying, in order to avoid strong base solution corrosion substrate pump.
The present invention adopts 6 estradiol standard items (0ng/ml, 0.03ng/ml, 0.1ng/ml, 0.3ng/ml, 1.0ng/ml, 3.0ng/ml) to carry out plotting curves.Instrument detects according to described method and obtains a calibration curve that RLU value is relevant with estradiol concentration, and after this in the measurement, the estradiol concentration in each sample is compared with typical curve and drawn estradiol content in the sample.
The mensuration of embodiment four kit quality
1, the sensitivity of kit
Kit sensitivity is defined as: measure 20 times the zero standard product, the mean value of mensuration adds 3 times of standard deviations.The sensitivity of this kit is 0.03ng/ml.
2, the accuracy of sample and precision
Accuracy refers to the matching degree between measured value and true value, and the kit accuracy recovery commonly used represents.Precision claims again repeatability, and the coefficient of variation commonly used represents.
Sample extraction method according to embodiment three, estradiol with 0.6ng/g (ml), two concentration of 1.2ng/g (ml) adds the chicken sample, estradiol with 0.3ng/g, two concentration of 0.6ng/g adds recovery to urine specimen, estradiol with 3.2ng/g, two concentration of 6.4ng/g adds recovery to the feed sample, every kind of each 4 of each concentration of sample are parallel, measure with three batches of kits, calculate average recovery rate and the precision of sample.Experimental result sees the following form.
Table 1 accuracy and precision are measured ng/g (ml)
As seen from the table, the average recovery rate scope that two concentration of estradiol are all added in chicken, urine, the feed sample between 85.6-96.2%, in batch, batch between all the coefficient of variation less than 15%.
3, specificity
Cross reacting rate refers to the ability of the antigenic determinant generation combination that antibody is different from structure.
Table 2 kit cross reacting rate
| Medicine name | Cross reacting rate |
| Estradiol | 100％ |
| Estradiol-3 methyl ether | 13％ |
| Ethinyloestradiol | 3％ |
| Estrone | Less than 0.1% |
| Oestradiol benzoate | Less than 0.1% |
| Estriol | Less than 0.1% |
4, correlativity
X＝CI-2008，Y＝RIA
Y＝1.24X-2.58
R＝0.9534
X is CI-2008 system measurement result, and Y is the ria-determination result.
Claims (8)
1. magnetic granule chemiluminescence kit that detects estradiol, the reagent that comprises has: luminous marker, fluorescein-labelled thing, standard items, quality-control product, separation agent.
2. kit according to claim 1, it is characterized in that: described separation agent is the paramagnetism nano microsphere that is coated with goat-anti FITC monoclonal antibody.
3. kit according to claim 2 is characterized in that: described paramagnetism nano microsphere is that the inside is coated with Fe
3O
4Or γ-Fe
2O
3, the surface contains-OH ,-COOH or-NH
2The microballon of reactive group.
4. each described kit according to claim 1-3, it is characterized in that: described luminous marker is the estradiol haptens of different Derivative of Luminol mark.
5. kit according to claim 4, it is characterized in that: described testosterone coupled isoluminol thing is ABEI, AHEI or ABEN.
6. each described kit according to claim 1-3, it is characterized in that: described kit also comprises titer, quality-control product and concentrated washing lotion.
7. one of according to claim 1-3 described kit is characterized in that: the estradiol monoclonal antibody that described fluorescein-labelled thing is the FITC mark.
8. a method of utilizing each described kit of claim 1-7 to detect estradiol comprises the following steps:
1) draw respectively 20 μ l-100 μ l standard items or samples, then add 20 μ l-100 μ l luminous markers, add again the fluorescein-labelled thing of 20 μ l-100 μ l, behind the abundant mixing, 37 ℃ of incubation 15min;
2) add the paramagnetism nano microsphere 80-150 μ l be coated with goat-anti FITC monoclonal antibody, behind the mixing at 37 ℃ of incubation 5min;
3) separate 5min with the magnetic separator frame, abandon behind the supernatant with cleaning fluid 300-500 μ l flushing compound precipitation;
4) above-mentioned cleaning step repeats 2-4 time;
5) 4) compound of gained separator well is directly put into the measurement camera bellows, add and excite substrate 1 and excite substrate 2, postpone to detect the relative light intensity (RLU) that sends behind the 3-5s, the content of estradiol and RLU proportion relation in the sample can be gathered the concentration that calibration curve method calculates estradiol by RLU.
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|---|---|---|---|
| CN 201110227587 CN102928415A (en) | 2011-08-09 | 2011-08-09 | Magnetic particle chemiluminescence kit for detecting estradiol, and applications thereof |
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|---|---|---|---|
| CN 201110227587 CN102928415A (en) | 2011-08-09 | 2011-08-09 | Magnetic particle chemiluminescence kit for detecting estradiol, and applications thereof |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103308677A (en) * | 2013-06-14 | 2013-09-18 | 南京医科大学第二附属医院 | Chemiluminescent immune quantitative detection kit of estradiol nanometer magnetic particles and preparation method thereof |
| CN103344774A (en) * | 2013-07-17 | 2013-10-09 | 江阴泽成生物技术有限公司 | Chemiluminescence immune assay kit and method for magnetic particle of human estradiol (E2) |
-
2011
- 2011-08-09 CN CN 201110227587 patent/CN102928415A/en active Pending
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103308677A (en) * | 2013-06-14 | 2013-09-18 | 南京医科大学第二附属医院 | Chemiluminescent immune quantitative detection kit of estradiol nanometer magnetic particles and preparation method thereof |
| CN103344774A (en) * | 2013-07-17 | 2013-10-09 | 江阴泽成生物技术有限公司 | Chemiluminescence immune assay kit and method for magnetic particle of human estradiol (E2) |
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Application publication date: 20130213 |