CN207976394U - Time-resolved fluoroimmunoassay chromatography detects the test strips and kit of AMH - Google Patents
Time-resolved fluoroimmunoassay chromatography detects the test strips and kit of AMH Download PDFInfo
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Abstract
The utility model is related to test strips and kit that a kind of quick, convenient, economic time-resolved fluoroimmunoassay chromatography detects AMH, and the test strips include PVC bottom plates, whole blood filter bed, antibody carrier film and blotting paper；The kit includes test strips, sample buffer and the ID card containing AMH standard curves of above-mentioned time-resolved fluoroimmunoassay chromatography detection AMH.Technical solutions of the utility model disclosure satisfy that the requirement of single part, small lot detection AMH, have the advantages that high sensitivity, specificity is high, blood using amount is few, detection time is short, easy to operate, testing result is accurate and reliable.
Description
Technical field
The utility model is related to hormone test technical fields, and in particular to a kind of time-resolved fluoroimmunoassay chromatography detection
The test strips and kit of AMH.
Background technology
Anti- Miao Le pipes hormone (Anti-M ü llerian hormone, AMH), also referred to as Miao Le pipes inhibitory hormone (M ü
Llerian-inhibiting hormone MIH), be in a kind of structure with transforming growth factor-β (TGF-β) superfamily member
Inhibin and the relevant glycoprotein hormones of activin, main function are regulation and control Growth and Differentiation and Folliculogenesis.In male embryo
In, for AMH by the testicular Sertoli cell SOX9 gene activations of male fetus, expression inhibiting female genital tract or Miao Leguan are (secondary
Ductus mesonephricus) development, to inhibit fallopian tubal, uterus and the generation of vagina.Specific period pair of the expression of AMH in embryonic development
Sex Differentiation is most important.In women, AMH is secreted by granular cell before sinus and small antral follicle count, until menopause gradually stops
Only.AMH adjusts ovarian follicular growth by inhibiting recruitment and the selection of dominant follicles of ovarian follicle.Entovarial small Antral follicles amount is got over
More, the concentration of AMH is just higher；Conversely, when ovarian follicle as age and various factors gradually use up, AMH concentration can also decrease.
Therefore AMH can be used as the molecular marker of prediction Ovary reserve.Recognized by more and more people with the effect of AMH,
Applications of the AMH in fields such as gynecological endocrine disease, reproductive medicine, sexual abnormalities increasingly attracts attention.Specifically, AMH
There is following purposes：
In terms of evaluation substantially fecundity, more personal AMH levels are given birth to the average level with age bracket in evaluation
It is helpful in terms of ability.It provides the guide of a determining female ovary deposit, and can identify may need to consider ovum
Son freezing or the women for attempting pregnancy as early as possible, rather than after waiting trial is considered further that when their long-term fecundity variation
Fertility.
Aspect in vitro fertilization, AMH are the tools for predicting poor responder in (IVF) in vitro fertilization.In addition, AMH water
The flat remaining ovum supply that can be used for one women of evaluation.According to NICE guides in vitro fertilization, AMH≤5.4pmol/l
(0.8ng/mL) implies ovarian hyperstimulation low reaction, and AMH >=25pmol/l (3.6ng/mL) indication height reacts.Secondly,
AMH levels are higher, and probability of being born after IVF is bigger.Therefore, AMH, which can be used for rationalizing, formulates Ol regimens, determines auxiliary life
The quantity of embryo is shifted in plantation technology, to improve Pregnancy Success rate to the maximum extent, while it is comprehensive to reduce ovarian hyperstimulation to the greatest extent
The risk that simulator sickness (OHSS) occurs.AMH predictions are respectively 82% and 76% for the sensibility and specificity of OHSS overreactions.
But AMH levels should combine Via vagina ovary scanning result comprehensive assessment ovarian follicle quantity and Ovarian Volume.
In terms of female tumor：Radiation and chemotherapy can damage ovarian reserve.In this case, AMH detections can before treating
Ovarian function loses degree after predicting long-term chemotherapy, decides whether to take fecundity conversation strategy, such as the freezing of egg mother cell
It preserves.And AMH detections often imply that fecundity declines after treating.Granular cell tumor ovarian secretion AMH, AMH test is being examined
Sensitivity in terms of these tumours of breaking is between 76 and 93%.
Stein-Leventhal syndrome (PCOS) is a kind of endocrine system disease being most commonly in Women of Childbearing Age, its main feature is that it is few or
No-clay weak interbed, androgen increase, polycystic ovary.The AMH of this kind of patient is higher by two to three times of normal value.
More and more people think that AMH is a kind of tool or biomarker diagnosed or prompt PCOS.
AMH is a brand-new Testing index, and compared to existing sex hormone detection project, it has the following advantages：AMH
Level is not influenced by the variation in the menstrual cycle between the period；AMH can take a blood sample in any time in the menstrual cycle；
AMH is not influenced by hormonal contraceptive, is convenient for Clinical practice；AMH can earlier, more accurately reflect age correlation ovarian reserve work(
The decline of energy.
Method currently used for detecting AMH mainly has enzyme-linked immunosorbent assay (enzyme-linked immunosorbent
Assay, ELISA), Electrochemiluminescince (electro-chemiluminescence immunoassay, ECLI), chemistry hair
Light method (chemiluminescent immunoassay, CLI) etc., but these methods all have the following defects：Detection device is wanted
Ask high, it is of high cost；Disturbing factor is more, and repeatability is bad；Detection time is long.Therefore these methods are unsuitable for the clinic of AMH
Quick diagnosis.
The test strips for being usually used in quick diagnosis at present are based on colloidal gold or fluorescein-labeled method more.Colloidal gold method is fixed
There are drawbacks in amount and sensitivity.Fluorescein marking sensitivity is 10-10mol/L；Background signal is strong, has non-specific fluorescence, specifically
Property is low；Fluorescein is marked to photo-labile, is easy to happen photobleaching phenomenon, and luminous efficiency is low, makes analysis result reliability and again
Renaturation reduces；Background signal is strong, and absorption spectrum is relatively narrow, and emission spectrum is wider, and Strokes displacements are small, is easy to happen the friendship of spectrum
It is folded, influence result precision.
Time-resolved fluoroimmunoassay chromatography (time-resolved fluorescence Immunoassay, TRFIA)
It uses lanthanide series (trivalent rare earth ions and its chelate) as tracer, labelled antigen or antibody, when immune response occurs, uses
Time-resolved fluorescence assay instrument measures the fluorescence intensity of immune response end product, further according to fluorescence intensity and relative intensity of fluorescence
Ratio judges the concentration of analyte in reaction system, achievees the purpose that quantitative analysis.The high sensitivity of TRFIA is up to 10-18mol/
L；Specificity fluorescent and non-specific fluorescence resolution are come by time delay, theoretical background are made to reach 0 by high specificity；It is glimmering
The light service life is extremely long, extremely long fluorescence fall time, and luminous efficiency is high, highly stable；Emission band is relatively narrow, excitation spectrum band compared with
Width, Strokes displacements are big, therefore interfere few, high sensitivity, and result precision is high.
However, being directed to the dry type fluorescence immunity analyzer using TRFIA in the prior art, lacking one kind can quickly, just
The kit and method that prompt, economy uses can meet the requirement of single part, small lot detection AMH.
Utility model content
The purpose of this utility model is in view of the deficiencies of the prior art, in conjunction with dry type fluorescence immunity analyzer, to provide one kind
Quickly, the test strips and kit of convenient, economic time-resolved fluoroimmunoassay chromatography detection AMH and, so as to meet
The requirement of single part, small lot detection AMH, with high sensitivity, specificity is high, blood using amount is few, detection time is short, operation is simple
Just, the accurate and reliable advantage of testing result.
To achieve the above object, the first aspect of the utility model provides a kind of time-resolved fluoroimmunoassay chromatography
The test strips of AMH are detected, the test strips include PVC bottom plates, whole blood filter bed, antibody carrier film and blotting paper；Whole blood filter bed resists
Body carrier film and blotting paper overlap successively to be pasted onto on PVC bottom plates；The antibody carrier film include on basilar memebrane and basilar memebrane according to
The secondary microballoon line, detection line and the control line that are mutually parallel being coated with, wherein the microballoon line is close to whole blood filter bed, control line is close
Blotting paper；
The control line is made of rabbit anti-mouse IgG, and detection line is made of AMH monoclonal antibodies, and microballoon line is by combining AMH
The time-resolved fluorescence microballoon of monoclonal antibody forms.
Further, interlaced 1.8-2.3mm overlap joints are pasted successively for the whole blood filter bed, antibody carrier film and blotting paper
On PVC bottom plates.It is above-mentioned interlaced to be preferably dimensioned to be 2mm.
Further, 4~6mm of the microballoon line-spacing whole blood filter bed.
Further, the microballoon line and detection line spacing are 4~10mm；And the detection line and the spacing of control line are
4~10mm.It is preferred that above range is respectively 4~6mm, more preferable 5~6mm.
Pass through the setting of above-mentioned spacing so that being optimal of sensitivity and specificity of the ELISA test strip AMH.
Further, the microballoon line, detection line and control line are solid threadiness.
Further, the basilar memebrane of the antibody carrier film is nitrocellulose filter.
Nitrocellulose is also known as nitrocellulose, cellulose nitrate, is the product of cellulose and nitric acid esterification.With
Cotton fiber is that the nitrocellulose of raw material is known as nitrocotton.Nitrocellulose is a kind of white fibrous polymer, water-fast, resistance to dilute
Sour, resistance to weak base and various oils.It is especially suited as the substrate membrane material of the utility model antibody carrier film, can be obtained
The theoretical Tomography Velocity of the antibody carrier film obtained reaches 15min or so.
Further, the AMH monoclonal antibodies are mouse anti human AMH monoclonal antibodies.
Further, the test strips further include shell, and shell is by whole blood filter bed, antibody carrier film, blotting paper and the bottoms PVC
Plate is packed into wherein, and shell includes upper casing and lower casing, and whole blood filter bed, antibody carrier film and blotting paper are pressed on PVC bottom plates by upper casing
On, and upper casing is respectively equipped with well and observation window in the part of corresponding whole blood filter bed and antibody carrier film.
The test strips can be used for the detection of dry type fluorescence immunity analyzer.
The second aspect of the utility model provides the test strips of above-mentioned time-resolved fluoroimmunoassay chromatography detection AMH
Preparation method, the preparation method includes the following steps：
(1) pretreatment of time-resolved fluorescence microballoon
After time-resolved fluorescence microballoon is handled 1min with ultrasonic disperse, 200 μ l, 17500rpm high speed centrifugations 20min are taken
After remove supernatant, 10~100mmol/L of sediment, the MES buffer solutions that pH is 6.0 wash；Carbodiimide and amber is added
Acid imide, it is 0.1mg/ml to make the two final concentration, after reacting at room temperature 10-30min, 17500rpm high speed centrifugation 20min, and precipitation
Object is washed with the pH MES solutions for being 6.0 and is resuspended to 1ml, obtains time-resolved fluorescence microspheres solution.
(2) preparation of the time-resolved fluorescence microballoon of AMH monoclonal antibodies is combined
50~200 μ g AMH monoclonal antibodies, mixing, room are added in every above-mentioned time-resolved fluorescence microspheres solutions of 250 μ l
Temperature reaction 1.5~2.5 hours closes liquid chamber with 10~50mmol/L, pH containing the 1-2%BSA Tris-HCl for being 7.5~8.5
After temperature closing 2 hours, 17500rpm high speed centrifugation 20min, with containing 0.2-1%BSA, 0.2-0.3%Tween20,0.1%NaN3
10~50mmol/L, pH be 7.5~8.5 Tris-HCl preserve liquid wash and be resuspended to 250 μ l, be kept in dark place, obtain in 4 DEG C
Obtain the time-resolved fluorescence microspheres solution in conjunction with AMH monoclonal antibodies.
(3) preparation of antibody carrier film
Rabbit anti-mouse IgG and mouse anti human AMH Dan Ke are diluted respectively with the 10mmol/L PBS buffer solution containing 1% sucrose
Grand antibody takes the 100 μ l of time-resolved fluorescence microspheres solution of above-mentioned combination AMH monoclonal antibodies to 0.5~1.5mg/ml of concentration,
17500rpm high speed centrifugation 20min, with 200 μ l-500 μ l containing 0.5-1%BSA, 0.2-0.3%Tween20,10% sucrose 10
The Tris-HCl microballoon re-suspension liquids that~50mmol/L, pH is 7.5~8.5 are resuspended；Existed with the amount of 1~5 μ l/cm with quantitative spray film instrument
The parallel spray painting control lines of 4~6mm, detection line and microballoon line are spaced on nitrocellulose filter；It is put into baking oven, 35~38 DEG C are protected from light baking
Dry, addition drier is sealed up for safekeeping spare.
(4) assembling of test strips
Overlap joint pastes whole blood filter bed, antibody carrier film and blotting paper successively on PVC bottom plates so that microballoon line is close to whole blood
Filter bed, control line obtain test paper plate close to blotting paper, then test paper plate cut to obtain the time-resolved fluoroimmunoassay chromatography
Detect the test strips of AMH.
The preparation method of the test strips further includes, with shell by whole blood filter bed, antibody carrier film, blotting paper and the bottoms PVC
Plate is packed into wherein, and whole blood filter bed, antibody carrier film and blotting paper are pressed on PVC bottom plates, and the upper casing in shell is right
Whole blood filter bed and the part of antibody carrier film is answered to be respectively equipped with well and observation window.
A kind of reagent of time-resolved fluoroimmunoassay chromatography detection AMH is provided in terms of the third of the utility model
Box, the kit include the test strips of above-mentioned time-resolved fluoroimmunoassay chromatography detection AMH, sample buffer and contain
The ID cards of AMH standard curves.
The preparation of the sample buffer includes：0.5% is dissolved in the Tris-HCl buffer solutions of pH 7.5~8.5
NaCl, 0.5%BSA and 1%Tween20,100 μ l/ pipes are dispensed into centrifuge tube.The sample buffer is used for chromatography samples.
The firing of the ID cards containing AMH standard curves includes：The schools AMH of various concentration are measured by the test strips
Quasi- product, with a concentration of X-axis of AMH calibration objects, detection line, control line fluorescence intensity ratio be Y-axis, be depicted as standard curve, write
Enter and generate bar code information to be stored in D cards.When detecting AMH concentration, dry type fluorescence immunity analyzer can be read outside test strips
Corresponding bar code information on shell.
4th aspect of the utility model provides a kind of method detecting AMH using mentioned reagent box, the method
Include the following steps：
(1) sample to be detected and detection reagent are warmed to room temperature again；
(2) 100 μ l whole bloods to be measured is taken to be added in 100 μ l sample buffers (as previously mentioned, sample buffer has been distributed into
100 μ l/ pipes), mixing obtains mixing sample；
(3) above-mentioned 100 μ l of mixing sample are drawn, are added in the well of test strips, room temperature is protected from light 15 minutes；
(4) dry type fluorescence immunity analyzer is opened, the ID cards of corresponding detection AMH are inserted into after initializing self-test；
(5) by the test strips socket of test strips inserting instrument, instrument is run, is calculated automatically by analysis software to be checked
AMH concentration in test sample sheet.
The testing principle of the kit of time-resolved fluoroimmunoassay chromatography detection AMH described in the utility model is dual anti-folder
Heart method.Wherein, the microballoon line is the AMH monoclonal antibodies of time-resolved fluorescence microballoon label；The detection line is coated with small
The anti-human AMH monoclonal antibodies of mouse；The control line is coated with rabbit anti-mouse IgG antibody.It is added dropwise and waits on whole blood filter bed when test
Sample dilution is detected, is acted on by chromatography, sample to be detected is moved to blotting paper end, is flowed through when microballoon line that time resolution is glimmering
The AMH monoclonal antibodies of light microballoon label are redissolved, if containing determined antigen in sample to be detected, that is, form microballoon antibody-antigene
Compound forms microballoon antibody-antigen-antibody compound when moving to detection line, and microballoon antibody is fixed, extra microballoon
Labelled antibody moves to control line by rabbit anti-mouse antibody capture.It is detected with dry type fluorescence immunity analyzer after being protected from light,
Obtain the power and its ratio of detection line and control line fluorescence intensity, information in instrument software combination ID cards, by actually detected value
The concentration of determinand in sample can be analyzed by substituting into preset standard curve.
Technical solutions of the utility model have the following advantages that：
The utility model kit can accurate quantitative analysis detection people's whole blood in AMH content, utilize time-resolved fluorescence layer
Analysis technology can avoid the fluorescence interference of sample itself, have the characteristics that high specificity, high sensitivity, accuracy are high；Standard curve
It is preset in ID cards, it can be achieved that single part, small lot detection, standard curve is all made without detecting every time；It is complete to detect sample
Blood, not examined place limitation, detection is quick, easy to operate.
Description of the drawings
Fig. 1 is the side structure signal that the time-resolved fluoroimmunoassay chromatography of the utility model detects the test strips of AMH
Figure；
Wherein：1, bottom plate；2, whole blood filter bed；3, antibody carrier film；4, blotting paper；5, microballoon line；6, detection line；7, it controls
Line.
Fig. 2 is the Facad structure signal that the time-resolved fluoroimmunoassay chromatography of the utility model detects the test strips of AMH
Figure.
Fig. 3 is the standard curve stored in the ID cards of detection AMH in the utility model embodiment 2.
Fig. 4 is the kit prepared using the utility model and Roche AMH kits (Electrochemiluminescince) testing result
The comparison of correlation.
Specific implementation mode
The utility model is further described with reference to specific embodiment, but the scope of protection of the utility model
It is not limited to that.
Embodiment 1：Time-resolved fluoroimmunoassay chromatography detects the preparation of the kit of AMH
Time-resolved fluoroimmunoassay chromatography detects the kit of AMH, using double-antibody method immunochromatography principle, detection
AMH contents in people's whole blood.The kit is detected test strips, the sample buffer of AMH by time-resolved fluoroimmunoassay chromatography
With the ID cards composition containing AMH standard curves.In this embodiment, as depicted in figs. 1 and 2：It overlaps successively on PVC bottom plates 1
Whole blood filter bed 2, antibody carrier film 3 (nitrocellulose filter is as basilar memebrane) and blotting paper 4；Antibody carrier film 3 is equipped with microballoon
Line 5, detection line 6 and control line 7；Whole blood filter bed 2 is used as sample application zone, for drawing whole blood sample to be checked.
Mouse anti human AMH monoclonal antibodies are coated with using the polystyrene ball containing rare-earth fluorescent dyestuff on microballoon line, Gu
Content 1% (contains 10mg microsphere particles) in 1ml solution, the average grain diameter of microballoon is 200nm, excitation wavelength 360nm, transmitted wave
Long 615nm；Peridium concentration is the μ l fluorescent microspheres of 50 μ g antibody/250；Mouse anti human AMH monoclonal antibodies coating is dense in detection line
Degree is 1mg/ml；Rabbit anti-mouse IgG antibody peridium concentration is 0.5mg/ml in nature controlling line.Microballoon line is with quantitatively spray film instrument with 3 μ l
Coating liquid measure/cm is sprayed on nitrocellulose filter；Detection line and control line are sprayed with quantitatively spray film instrument with 1 μ l coatings liquid measure/cm
It is applied on nitrocellulose filter.From Bangs Lab companies of the U.S., mouse anti human AMH is mono- for time-resolved fluorescence microballoon buying used
Clonal antibody and rabbit anti-mouse IgG antibody are purchased from Hangzhou Bo Yin Bioisystech Co., Ltd.
In this embodiment, wherein the test strips of the time-resolved fluoroimmunoassay chromatography detection AMH are using as follows
It is prepared by method：
(1) pretreatment of time-resolved fluorescence microballoon
After time-resolved fluorescence microballoon is handled 1min with ultrasonic disperse, 200 μ l, 17500rpm high speed centrifugations 20min are taken
After remove supernatant, sediment 1ml 50mmol/L, the MES buffer solutions that pH is 6.0 wash；20 μ l carbodiimides and 20 are added
μ l succinimides, it is 0.1mg/ml to make the two final concentration, after reacting at room temperature 30min, 17500rpm high speed centrifugation 20min,
Sediment is washed with the pH MES solutions for being 6.0 and is resuspended to 1ml, obtains time-resolved fluorescence microspheres solution；
(2) preparation of the time-resolved fluorescence microballoon of AMH monoclonal antibodies is combined
50 μ g AMH monoclonal antibodies, mixing, room temperature reaction are added in every above-mentioned time-resolved fluorescence microspheres solutions of 250 μ l
2 hours, after being closed 2 hours with 50mmol/L, pH containing 1%BSA Tris-HCl confining liquid room temperatures for being 8.0,17500rpm high
Speed centrifugation 20min, with containing 0.1%BSA, 0.2%Tween20,0.1%NaN350mmol/L, pH be 8.0 Tris-HCl protect
Liquid storage is washed and is resuspended to 250 μ l, is kept in dark place in 4 DEG C, is obtained and is combined the time-resolved fluorescence microballoon of AMH monoclonal antibodies molten
Liquid；
(3) preparation of antibody carrier film
Rabbit anti-mouse IgG and mouse anti human AMH Dan Ke are diluted respectively with the 10mmol/L PBS buffer solution containing 1% sucrose
Grand antibody takes the 100 μ l of time-resolved fluorescence microspheres solution of above-mentioned combination AMH monoclonal antibodies to concentration 1mg/ml,
17500rpm high speed centrifugation 20min, with 200 μ l-500 μ l containing 0.5-1%BSA, 0.2-0.3%Tween20,10% sucrose 10
The Tris-HCl microballoon re-suspension liquids that~50mmol/L, pH is 7.5~8.5 are resuspended；With quantitative spray film instrument with the amount of 1 μ l/cm in nitre
Parallel spray painting control line and detection line on acid cellulose film are spaced 4mm, with quantitative spray film instrument with the amount of 3 μ l/cm in cellulose nitrate
Parallel spraying microballoon line on plain film, interval detection line 4mm；It is put into 37 DEG C of baking ovens and is protected from light drying 10 hours, drier is added and seals up for safekeeping
It is spare；
(4) assembling of test strips
Interlaced 2mm overlap joints paste whole blood filter bed (size 30* successively on PVC bottom plates (size 80*300mm)
300mm, glass fibre cotton material), antibody carrier film (size 25*300mm, nitrocellulose material) and blotting paper (size
For 28*300mm), wherein microballoon line cuts into 4mm close to blotting paper close to whole blood filter bed, control line to obtain test paper plate
Test strips.
The test strips of detection AMH are assembled in the plastic shell made of plastics upper casing and plastics lower casing fastening, on plastics
Shell is equipped with well and observation window, and well corresponds to the whole blood filter bed 2 of AMH test strips, and as a result observation window corresponds to
The detection line 6 and control line 7 of AMH test strips.
The sample buffer that the embodiment kit includes is to dissolve in 0.5% in the Tris-HCl buffer solutions of pH 8.0
NaCl, 0.5%BSA and 1%Tween20 are sub-packed in 100 μ l/ pipes in centrifuge tube.The sample buffer is used for chromatography samples.
In the embodiment kit, an ID card containing AMH standard curves, the product mark of same batch are matched per box
Directrix curve is identical, the calibration object of various concentration is measured by the AMH test strips, with a concentration of X-axis of calibration object, detection line, control
The ratio of line fluorescence intensity processed is Y-axis, is depicted as standard curve, is written and generates corresponding bar code information and is stored in ID cards, and
Matched bar code is printed to be pasted onto on test strips shell.When detectable concentration, ID card inserting instrument ID card plugs, dry type fluorescence is exempted from
Epidemic disease analyzer reads corresponding bar code information on test strips shell, obtains corresponding standard curve.
Embodiment 2：Time-resolved fluoroimmunoassay chromatography detection AMH concentration
In the test strips prepared by embodiment 1 be added various concentration people's AMH antigens calibration object (16.0ng/ml,
8.0ng/ml, 4.0ng/ml, 2.0ng/ml, 1.0ng/ml, 0.5ng/ml, 0.1ng/ml, 0.0ng/ml totally eight concentration, each
Three repetitions are arranged in concentration, are diluted with sample buffer by 1.0mg/ml people's AMH antigens), after chromatographing 15min, pass through
Law Firm Suzhou Jiangsu and the FIC-S1 type dry type fluorescence immunity analyzers of precision instrument Biology Pharmacy Co., Ltd advanced in years production read detection
The fluorescence intensity ratio (detection line detected value/control line detected value) of line, control line.
With a concentration of X-axis of AMH calibration objects, equation is established as Y-axis using sample detection line fluorescence intensity/control line fluorescence intensity
And it is fitted to standard curve, standard curve is as shown in Figure 3.R values are 0.9982.
The process specifically includes the following steps：
(1) sample to be detected and detection reagent are warmed to room temperature again；
(2) it takes 100 μ l whole bloods to be measured that (as previously mentioned, being distributed into 100 μ l/ pipes) is added in 100 μ l sample buffers, mixes
It is even, obtain mixing sample；
(3) 100 μ l of sample after the above-mentioned mixing of absorption, are added in the well of test strips, room temperature is protected from light 15 minutes；
(4) dry type fluorescence immunity analyzer is opened, the ID cards of corresponding detection AMH are inserted into after initializing self-test；
(5) by the test strips socket of test strips inserting instrument, instrument is run, is calculated automatically by corresponding analysis software
Go out the concentration of the AMH in sample to be tested.
Clinical sample detects
60 parts of whole blood sample for acquiring hospital detection AMH, with the kit and Roche Electrochemiluminescince of the utility model
The kit of detection AMH is compared.In the utility model kit, takes 100 μ l of whole blood that sample buffer is added, taken after mixing
100 μ l are added in detection card well, the FIC- that chromatography is produced after 15 minutes by Suzhou and precision instrument Co., Ltd advanced in years
S1 type dry type fluorescence immunity analyzers read concentration, and same blood sample is using the detection AMH examinations of comparison system Roche Electrochemiluminescince
Agent box carries out Concentration Testing.Two methods testing result carries out linear analysis, as shown in figure 4, the fine R=0.983 of its correlation,
P>0.05, mean relative deviation is less than 10%, as a result meets clinical analysis requirement, is suitable for clinical detection.
Although above having made detailed description to the utility model with generality explanation and specific embodiment,
On the basis of the utility model, it can be made some modifications or improvements, this is apparent to those skilled in the art
's.Therefore, these modifications or improvements on the basis of without departing from the spirit of the present invention, belong to the utility model and want
Seek the range of protection.
Claims (6)
1. a kind of test strips of time-resolved fluoroimmunoassay chromatography detection AMH, which is characterized in that the test strips include PVC
Bottom plate, whole blood filter bed, antibody carrier film and blotting paper；The whole blood filter bed, antibody carrier film and blotting paper overlap stickup successively
On PVC bottom plates；The antibody carrier film include the microballoon line being mutually parallel being coated with successively on basilar memebrane and the basilar memebrane,
Detection line and control line, wherein the microballoon line is close to whole blood filter bed, the control line is close to blotting paper；
The control line is made of rabbit anti-mouse IgG, and the detection line is made of AMH monoclonal antibodies, and microballoon line is by combining AMH
The time-resolved fluorescence microballoon of monoclonal antibody forms.
2. the test strips of time-resolved fluoroimmunoassay chromatography detection AMH according to claim 1, which is characterized in that institute
State 4~6mm of microballoon line-spacing whole blood filter bed；The microballoon line and detection line spacing are 4~6mm；And the detection line and control line
Spacing be 4~6mm.
3. the test strips of time-resolved fluoroimmunoassay chromatography detection AMH according to claim 2, which is characterized in that institute
It is solid threadiness to state microballoon line, detection line and control line.
4. the test strips of time-resolved fluoroimmunoassay chromatography detection AMH according to claim 3, which is characterized in that institute
The basilar memebrane for stating antibody carrier film is nitrocellulose filter.
5. the test strips of time-resolved fluoroimmunoassay chromatography detection AMH according to claim 4, which is characterized in that institute
It further includes shell to state test strips, and whole blood filter bed, antibody carrier film, blotting paper and PVC bottom plates are packed into wherein by the shell, described
Shell includes upper casing and lower casing, and whole blood filter bed, antibody carrier film and blotting paper are pressed on PVC bottom plates by upper casing, and upper casing exists
The part of corresponding whole blood filter bed and antibody carrier film is respectively equipped with well and observation window.
6. a kind of kit of time-resolved fluoroimmunoassay chromatography detection AMH, which is characterized in that the kit includes right
It is required that the time-resolved fluoroimmunoassay chromatography detection test strips of AMH, sample buffer described in any one of 1-5 and containing
The ID cards of AMH standard curves.
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| CN108195815A (en) * | 2018-01-31 | 2018-06-22 | 李翀 | Time-resolved fluoroimmunoassay chromatography detects test strips, kit and the method for AMH |
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Effective date of registration: 20210609 Address after: Floor 17, building B8, Zhihui Yunjin incubation base of Guizhou great health pharmaceutical industry, Wudang District, Guiyang City, Guizhou Province Patentee after: Guizhou lizhijian Biotechnology Co.,Ltd. Address before: 100101 Beijing city Chaoyang District Datun Road No. 15 Chinese Academy of Sciences Institute of Biophysics Patentee before: Li Li |