CN109541204A - A kind of detection antihepatitis b e antibody kit and preparation method thereof - Google Patents

A kind of detection antihepatitis b e antibody kit and preparation method thereof Download PDF

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CN109541204A
CN109541204A CN201811329562.5A CN201811329562A CN109541204A CN 109541204 A CN109541204 A CN 109541204A CN 201811329562 A CN201811329562 A CN 201811329562A CN 109541204 A CN109541204 A CN 109541204A
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hbe
detection
magnetic microsphere
antibody
antihepatitis
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李根平
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Guangzhou Origin Health Technology Co Ltd
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    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/543Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
    • G01N33/54313Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals the carrier being characterised by its particulate form
    • G01N33/54326Magnetic particles
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • G01N33/6854Immunoglobulins
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • G01N33/6893Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids related to diseases not provided for elsewhere

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Abstract

The invention discloses a kind of detection antihepatitis b e antibody kits and preparation method thereof, are grouped as by following group: being coated with the magnetic microsphere of anti-HBe, anti-HBe calibration object, neutralize antigen, anti-HBe marker, analysis buffer, cleaning solution, enhancement solution and RFID card in conjunction with lanthanide series.This method is based on magnetic microsphere and combines with Timed-resolved fluoroimmunoassay, overcome the physical sorption reaction time of ELISA Plate longer, the slower drawback of testing result, the reaction time is greatly shortened, while also having that time resolution detection technique accuracy height, high sensitivity, high specificity, the range of linearity is wide, detection is stable and convenient advantage.Magnetic microsphere is coated with corresponding antigen or antibody and greatly increases the contact surface area of immune response because of the characteristic of magnetic microsphere solid, to greatly shorten detection time, detecting anti-HBe only needs 35 minutes can detect result.

Description

A kind of detection antihepatitis b e antibody kit and preparation method thereof
Technical field
The present invention relates to a kind of detection antihepatitis b e antibody kits and preparation method thereof, and in particular to Yi Zhongji The preparation method of detection antihepatitis b e antibody kit is combined with Timed-resolved fluoroimmunoassay in magnetic microsphere.
Background technique
Hepatitis B (abbreviation hepatitis B) is to lead to liver property inflammation and liver function energy loss caused by hepatitis type B virus (HBV) A kind of harmful communicable disease.Antihepatitis b e antibody (anti-HBe) is the mark of HBV infection, often comes across hepatitis B In viral surface antigen or the blood of the anti-HBs positive.It is reported that the detection of anti-HBe is generally found from B-mode liver It is scorching convalescence, slightly early compared with hepatitis B surface antibody appearance, it can be detected in 4 to 7 weeks after jaundice appearance, be a prognosis bona Sign.
The project of hepatitis B Serological testing at present generally comprises HBsAg, Anti-HBs, HBeAg, Anti-HBe, Anti- HBc, that is, " Hepatitis B virus " being commonly called as.The detection method of the existing anti-HBe of mainstream has enzyme linked immunological (ELISA), colloidal gold method, change Learn luminous (CLIA), electrochemical luminescence (ECL), time-resolved fluoroimmunoassay (TRFIA) etc..ELISA is examined as sxemiquantitative Survey method is now most widely used detection method, but its sensitivity, linear extent range are not up to higher level, it is difficult to be adapted to The demand of market development.Colloidal gold method and ELISA have the shortcomings that identical.Though and CLIA and ECL high sensitivity but presence inspection Measurement equipment cost is high, and corresponding marker research and development threshold is high, and the country can be under one's control in short-term, equally also limits Popularization at home.And time-resolved fluoroimmunoassay (TRFIA) sensitivity can reach the consistent level of CLIA, not only detect Equipment manufacturing cost is cheap, and labelling technique is quite mature at home, testing cost is low simultaneous.And traditional time resolution detection, Still based on the physical absorption of blank ELISA Plate, time-consuming for detection process.Shorten the reaction time, reduce testing cost, improves inspection Sensitivity and the range of linearity are surveyed, it will be with good market prospects.
Summary of the invention
Based on this, it is an object of the invention to overcome defect in the prior art and the relevant technologies testing cost is high lacks Point provides the preparation method of a kind of detection antihepatitis b e antibody kit and the kit.
To achieve the above object, the present invention adopts the following technical scheme that:
A kind of detection antihepatitis b e antibody kit, is grouped as by following group: being coated with the magnetic microsphere, anti-of anti-HBe HBe calibration object, neutralize antigen, the anti-HBe marker with lanthanide series in conjunction with, analysis buffer, be concentrated washing lotion (cleaning solution) and Enhancement solution, RFID card.
Preferably, the magnetic microsphere in the magnetic microsphere for being coated with anti-HBe is by micron-sized Fe2O3Or Fe3O4Magnetic particle and high-molecular organic material carry out it is compound, formed have superparamagnetism, can be with immunising antigen or anti- The micron-sized microballoon that body combines, that is, be commonly referred to as " magnetic bead ".It is 0.1~5 μm that magnetic microsphere, which should be able to meet diameter, magnetic microsphere Various active functional group, including but not limited to hydroxyl (- OH), amino (- NH can be had by surface modification2), carboxyl (- COOH)。
Preferably, the anti-HBe in the magnetic microsphere for being coated with anti-HBe can be monoclonal antibody, can also To be polyclonal antibody.
Preferably, the magnetic microsphere for being coated with anti-HBe is prepared using following steps:
It is micro- with the magnetism after cleaning, activation by anti-HBe after 2~8 DEG C of superspeed refrigerated centrifuges are by buffer system replacement Treatment Ball mixing and constant-temperature incubation clean magnetic bead and abandon supernatant, then closed with magnetic bead confining liquid after incubation, cleaning magnetic bead is again The anti-HBe being coated with is saved in liquid in magnetic bead and is uprightly saved with 2~8 DEG C of refrigerators by secondary discarding supernatant.
It is 10 μ g that every milligram of magnetic microsphere of EDC and Sulfo-NHS, which is preferably loaded quality, in the activation of the magnetic microsphere ~1000 μ g.Activation method includes but is not limited to EDC, Sulfo-NHS one such and two kinds.
Preferably, the neutralization antigen is the HBeAg antigen of preferred proportion.
Preferably, the antibody in the anti-HBe marker in conjunction with lanthanide series is either monoclonal antibody also can be more It is anti-.Lanthanide series therein is to be completed in conjunction with anti-HBe by intermediate chelating agent, lanthanide series include but is not limited to europium (EU), Samarium (Sm), chelating agent include but is not limited to isothiocyanic acid phenyl-EDTA, isothiocyanic acid benzyl-DTTA, P- isothiocyano benzyl Base-DTTA, diethylene triamine pentaacetic acid aminophenyl-EDTA.
The present invention also provides a kind of methods for detecting anti-HBe using mentioned reagent box, and described method includes following steps:
(1) working solution is diluted to by washing lotion is concentrated with purified water;
(2) magnetic microsphere of the working concentration diluted is added in reaction cup;
(3) sample to be examined or calibration object are added in above-mentioned reaction cup;
(4) antigen will be neutralized to be added in above-mentioned reaction cup;
(5) reaction cup is incubated for for the first time at room temperature;
(6) magnetic microsphere after using the cleaning liquid in (1) to add magnetic in washing reaction cup;
(7) marker is diluted to working solution with analysis buffer;
(8) after washing, the marker working solution diluted in (7) is added in reaction cup for degaussing;
(9) reaction cup is incubated for for second at room temperature;
(10) magnetic microsphere after using the cleaning liquid in (1) to add magnetic in washing reaction cup;
(11) after washing, degaussing is added enhancement solution and is incubated for;
(12) after being incubated for, the fluorescent collecting for carrying out corresponding wavelength is detected and is analyzed.
Preferably, for the present invention in order to further reduce manual steps, self-produced SmartTRF grinds certainly in cooperation company The relevant parameter of detection method, operating procedure are all copied to RFID card by complete series Immunofluorescence test equipment In.In actual mechanical process, it is only necessary to RFID card is adapted to above-mentioned Immunofluorescence test equipment can be automatically finished it is above-mentioned The operating procedure of experiment.RFID(Radio Frequency Identification) technology, also known as radio frequency identification are one The kind communication technology can be identified specific objective by radio signals and read and write related data, without identifying system and specific mesh Mechanical or optical contact is established between mark.
The present invention also provides a kind of preparation methods for detecting anti-HBe kit, and described method includes following steps:
(1) preparation is coated with the magnetic microsphere of anti-HBe;
(2) anti-HBe marker of the preparation in conjunction with lanthanide series;
(3) calibration object is prepared;
(4) preparation neutralizes antigen;
(5) analysis buffer, concentration washing lotion and enhancement solution are prepared;
(6) each component is dispensed respectively into corresponding storage container;
(7) RFID card duplicate copy;
(8) coding, labelling;
(9) it is assembled into finished product kit.
Compared with prior art, the beneficial effects of the invention are that:
This method is based on magnetic microsphere and combines with Timed-resolved fluoroimmunoassay, that is, when overcoming the physical adsorption reaction of ELISA Plate Between it is longer, the slower drawback of testing result greatly shortens the reaction time, while also having time resolution detection technique accuracy Height, high sensitivity, high specificity, the range of linearity are wide, detection is stable and convenient advantage.Magnetic microsphere be coated with corresponding antigen or Antibody greatly increases the contact surface area of immune response because of the characteristic of magnetic microsphere solid, so that detection time is greatly shortened, Detecting anti-HBe only needs 35 minutes can detect result.In addition, being no longer limited by traditional enzyme due to the fluid behaviour of magnetic microsphere The frame of target limits, and detection can be completed in arbitrary reaction cup and small test tube, while can reduce detecting instrument equipment Volume size and equipment cost, also can be such as ELISA Plates, the reflective inspection of chemistry to meet the full-automatic detection demand in two, three line cities Survey equally carries out big flux detection, meets the full-automatic detection demand in a line city.
Detailed description of the invention
Fig. 1 is to be used to store the magnetic microsphere for being coated with anti-HBe and group of the lanthanides member using detection kit of the present invention The schematic top plan view of the reagent strip of anti-HBe marker, neutralization antigen, analysis buffer, cleaning solution and enhancement solution that element combines.
Fig. 2 is to be used to store the magnetic microsphere for being coated with anti-HBe and group of the lanthanides member using detection kit of the present invention The stereochemical structure mark of anti-HBe marker, the reagent strip for neutralizing antigen, analysis buffer, cleaning solution and enhancement solution that element combines is anticipated Diagram is intended to.
Specific embodiment
The present invention is further illustrated with attached drawing with reference to embodiments, and it is special that technology of the invention is better described Point, technical solution.Following embodiment does not cause any restrictions to the present invention.HBeAg antigen, antibody come in following embodiment Derived from Wuhan AudioCodes Bo Tai Biotechnology Co., Ltd;Magnetic microsphere derives from GE company;Europium label is purchased from Finland Wallac Company;Antihepatitis b e antibody National reference is provided by National Institute for Food and Drugs Control;Contrast agents box is Abbott Laboratories Antihepatitis b e antibody assay kit (chemiluminescence particulate immunodetection).
Embodiment 1
A kind of detection antihepatitis b e antibody kit, the kit includes: the magnetic microsphere for being coated with anti-HBe, anti- HBe calibration object neutralizes antigen, the anti-HBe marker of europium label, analysis buffer, concentration washing lotion and enhancement solution, RFID card.
The present invention also provides the preparation methods of above-mentioned detection antihepatitis b e antibody kit, and the method includes such as Lower step:
(1) preparation is coated with the magnetic microsphere of anti-HBe: anti-HBe is set buffer system through 2~8 DEG C of superspeed refrigerated centrifuges After changing processing, mixes simultaneously constant-temperature incubation 1~3 hour with 1 μm of carboxyl magnetic microsphere of diameter after cleaning, activation, used after incubation Magnetic bead cleaning solution cleaning magnetic bead simultaneously abandons supernatant, is then closed with magnetic bead confining liquid, and cleaning magnetic bead abandons supernatant again The anti-HBe magnetic microsphere being coated with is saved in liquid in magnetic bead and is uprightly saved with 2~8 DEG C of refrigerators by liquid.It is again that coating is good anti- HBe magnetic microsphere saves liquid with magnetic bead and is diluted to working solution concentration, is distributed into 10mL/ bottles.Preferably, magnetic microsphere and anti-HBe Coated mass ratio is one of 10:1,20:1,30:1,40:1;Preferably, the displacement buffer and magnetic bead cleaning solution It is the MES buffer of 0.05~0.5M PH, 5.8~PH 7.0;Preferably, EDC and Sulfo-NHS in the activation of magnetic microsphere It is 10 μ of μ g~1000 g that every milligram of magnetic microsphere, which is preferably loaded quality,;Preferably, the confining liquid of magnetic microsphere and preservation liquid are The Tris-HCl buffer of 0.1~0.5M PH 7.0~8.5 containing 0.1%~8% BSA;
(2) prepare europium label anti-HBe marker: by anti-HBe be placed in molecular cut off be 10000 ultra-filtration centrifuge tubes in, 10000rpm is centrifuged 7~10min, discards filtrate.Add 9.6 carbonate buffer solution 10000rpm of 0.05M PH centrifugation 7~ 10min 2~3 times repeatedly, centrifuge tube filter membrane reversion 3000rpm is centrifuged 6min, collects 200 μ L solution being finally concentrated.And it will Itself and in advance with the solvent DTTA-EU of carbonate buffer solution3+Mixing, anti-HBe and europium mass ratio are 2:1, and 2~8 DEG C of oscillations mix 48 ± 2 hours.Sephadex TM G-75 gel column that label solution is balanced through 7.8 Tris-Hcl buffer of 0.05M PH ( 1.0*50cm) chromatographic purifying monitors in A280 and collects first peak.By the anti-HBe marker of the europium being collected into label with 0.2% 7.8 Tris-Hcl buffer of BSA, 0.05M PH is diluted to 1/20 times of optium concentration, dispenses to 1.0mL/ bottles.
(3) anti-HBe calibration object is prepared: using 2% BSA, 0.5% Tween-20,0.02% Proclin300, pH 7.5 Anti- HBe is diluted to lower concentration values point 2PEI U/mL, high concentration value point 32PEI U/mL by 0.02M Tris-HCl buffer.
(4) preparation neutralizes antigen: using 2%BSA, 0.5%Tween-20,0.02%Proclin300, pH7.5 0.02M By HBeAg antigen, 1:5000 dilutes Tris-HCL buffer in proportion, dispenses to 1.0ml/ bottles.
(5) it prepares analysis buffer: containing Tween-20, Proclin300, EDTA, BSA, Tris-HCl buffer, dividing It is filled to 30~40mL/ bottles.
(6) preparation concentration washing lotion: containing Tween-20, Proclin300, Tris-Hcl buffer, dispense to 30~ 40mL/ bottles.
(7) it prepares enhancement solution: containing sodium acetate, β-NTA, TOPO, glacial acetic acid, dehydrated alcohol, Triton X-100, dividing It is filled to 30~40mL/ bottles.
(8) it prepares RFID card: blank RFID card is subjected to relative parameters setting by detection method in the present embodiment of the present invention;
(9) coding, labelling assemble kit.
The present invention also provides the detection method of above-mentioned detection antihepatitis b e antibody kit, the method is specifically grasped Make as follows:
(1) reagent prepares
Kit restores in being placed at room temperature for room temperature;
The anti-HBe marker of europium label is diluted to working solution concentration using analysis buffer, is mixed stand-by;
Cleaning solution: purified water is added by 1:25 in concentration washing lotion and is diluted to work cleaning solution;
Upright light rolling is coated with the magnetic microsphere of anti-HBe before experiment, mixes stand-by;
(2) experimental implementation
The magnetic microsphere for drawing 50 μ L mixing is added in reaction cup;
Sample to be tested or 50 μ L of calibration object are added into each reaction cup;
It is added into each reaction cup and neutralizes 50 μ L of antigen;
Room temperature blending incubation 20 minutes;
After incubation, cleaned 4 times using cleaning solution;
The anti-100 μ L of HBe marker working solution diluted is added into each reaction cup again;
Room temperature blending incubation 15 minutes;
After incubation, cleaned 4 times using cleaning solution;
The addition 100 μ L of enhancement solution into each reaction cup, room temperature blending incubation 3 minutes;Fluorescent collecting is completed in 30 minutes And carry out data analysis.
In the present embodiment, actual laboratory operating procedures and relevant parameter are copied in matched RFID card in advance, real It tests after operating process only needs to be ready to by reagent preparation process, RFID card is adapted to fully-automatic equipment can be completed from information Read, be loaded onto the overall process of detection.
Embodiment 2
A kind of detection antihepatitis b e antibody kit, essentially identical with detection kit described in embodiment 1, difference exists In:
(1) the detection antihepatitis b e antibody kit component only includes: reagent strip, anti-HBe calibration object, RFID card.
(2) in the present embodiment, reagent strip is by being coated with the magnetic microsphere of anti-HBe, europium label in the detection kit Anti- HBe marker, neutralize antigen, analysis buffer, sealer in cleaning solution and enhancement solution, packing to the corresponding hole of reagent strip After form.Wherein cleaning solution is that concentration washing lotion adds 25 times of purified water dilution to form in embodiment 1;Remaining each component and embodiment 1 In it is identical.
In the present embodiment, just as shown in Figure 1 and Figure 2, each hole bit function of reagent strip is described as follows in the detection kit:
Reagent strip is from left to right arranged successively, and title is followed successively by the 1st~13 hole.1st, 2 holes are instrument connection, the 3rd, 4 holes be Fluorescent marker hole, the 5th hole be analysis buffer hole, the 6th, 7 holes be cleaning fluid apertures, the 8th, 9 be Sample Dilution fluid apertures, the 10th, 11 It is enhancing fluid apertures for neutralizer hole, the 12nd, the 13rd is preparation hole.1st and 2 holes are that storage magnetic microsphere is immunoreacted Reacting hole, it is 800 μ L that maximum, which can store liquid volume,;3rd, 4 holes can be disassembled into from entire reagent strip and be independent component, Convenient for carrying out packing storage to fluorescent marker.3rd, 4,5 holes can store maximum liquid volume be 400 μ L;6th, 7 holes can be with Storage maximum liquid volume is 3000 μ L;It is 400 μ L that 8th ~ 12 hole, which can store maximum liquid volume,;13rd hole can store most Big liquid volume is 600 μ L.
In the present embodiment, the reagent strip in the detection kit is made after carrying out sealer as follows:
300 μ L are coated with anti-HBe magnetic microsphere, 50 μ L europiums mark anti-HBe marker, 200 μ L analysis buffers, 3000 μ L It is dispensed to the 1st of reagent strip the, 3,5,6,10,12 holes respectively in cleaning solution, 100 μ L with antigen, 200 μ L enhancement solutions, after sealer The reagent strip is made.
The present invention also provides the preparation methods of above-mentioned detection antihepatitis b e antibody kit, are obtained in the above describe manner After reagent strip, kit is constituted with anti-HBe calibration object, RFID card.In addition to each component packing mode and storage container are different Outside sample, remaining is in the same manner as in Example 1.
The present invention also provides the detection methods of above-mentioned detection antihepatitis b e antibody kit, it is only necessary to by above-mentioned examination Agent item is inserted into the reagent clamp bar slot of SmartTRF serial equipment after gently shaking mixing, and equipment reads RFID card relevant information can be complete It is automatically performed detection process.The related information parameters and detecting step of RFID card are in the same manner as in Example 1.
Embodiment 3
The performance evaluation of detection antihepatitis b e antibody kit of the present invention:
By the kit and detection method prepared in embodiment, B-mode liver of the detection buying from National Institute for Food and Drugs Control Scorching virus e antibody National reference, and collect the clinical sample that anti-HBe is detected from Abbott Laboratories, hospital Architect i2000 This 300.
Embodiment 1, embodiment 2 detect antihepatitis b e antibody National reference, as a result as follows:
(1) negative match-rate: 15 parts of negative National reference N1~N15, embodiment 1, embodiment 2 detect coincidence rate (-/-) It is 15/15;
(2) positive coincidence rate: 10 parts of positive National reference P1~P10, embodiment 1, embodiment 2 detect coincidence rate (+/+) It is 10/10;
(3) accuracy: accuracy reference material repeats detection 10 times, and embodiment 1, embodiment 2 detect accuracy C.V≤20%;
(4) limit of identification: detection sensitivity reference material, embodiment 1, the detection of embodiment 2 meet≤1IU/mL.
Embodiment 1, embodiment 2 detect the anti-HBe300 clinical sample of Abbott Laboratories, as a result as follows:
The comparison of 1 embodiment of table, 1 clinical sample
Detection kit described in the embodiment of the present invention 1 detects hepatitis B e antigen and contrast agents negative match-rate 100%, sun Property coincidence rate 100%.
What concentration washing lotion (cleaning solution) referred in embodiment 1 is the high concentration cleaning solution to be diluted to working solution;Cleaning What liquid referred in example 2 is the working solution for not needing any processing.
It should be understood that detecting the detection method of anti-HBe kit in embodiment 1 and preparation method is to meet Big flux testing goal and invent, instrument and equipment bulky and cost is relatively high, for further satisfaction two, three line cities The detection demand in city, correspondingly, we have further made some detection methods and kit preparation on the basis of embodiment 1 Modification in method, as in embodiment 2.
It should be understood that difference of the present invention in example 2 with detection method and preparation method in embodiment 1 exists In, embodiment 2 be by the magnetic microsphere for being coated with anti-HBe in embodiment 1, the anti-HBe marker in conjunction with lanthanide series, Antigen, analysis buffer, cleaning solution and enhancement solution is neutralized to dispense into special reagent strip (as shown in Figure 1 and Figure 2), therefore simultaneously The component of kit only has reagent strip, RFID card, anti-HBe calibration object in embodiment 2.Wherein RFID card, anti-HBe calibration object with It is consistent in embodiment 1.
It should be understood that the detection method of the present invention in example 2 is after reagent strip is inserted into detection device, to use simultaneously Corresponding RFID card is adapted to equipment, and equipment full automatic working step is consistent with embodiment 1;
It should be understood that the preparation method of the detection kit of the present invention in example 2 is with the difference in embodiment 1, Concentration washing lotion becomes cleaning solution in step (5), and step (6) becomes " dispensing to the corresponding aperture of reagent strip each component respectively, dividing Sealer coding immediately after installing ".

Claims (7)

1. a kind of detection antihepatitis b e antibody kit, it is characterised in that be grouped as by following group: being coated with anti-HBe's Magnetic microsphere, neutralizes antigen, the anti-HBe marker in conjunction with lanthanide series, analysis buffer, cleaning at anti-HBe calibration object Liquid, enhancement solution and RFID card.
2. detection antihepatitis b e antibody kit as described in claim 1, which is characterized in that described being coated with is anti- Magnetic microsphere in the magnetic microsphere of HBe is by micron-sized Fe2O3Or Fe3O4Magnetic particle and high-molecular organic material carry out It is compound, form micron-sized microballoon that there is superparamagnetism, can combining with immunising antigen or antibody.
3. detection antihepatitis b e antibody kit as claimed in claim 2, which is characterized in that described
Magnetic microsphere diameter is 0.1~5 μm, and magnetic microsphere has various active functional group by surface modification, including but not It is limited to hydroxyl, amino or carboxyl.
4. detection antihepatitis b e antibody kit as described in claim 1, which is characterized in that described being coated with is anti- The magnetic microsphere of HBe is prepared using following steps:
It is micro- with the magnetism after cleaning, activation by anti-HBe after 2~8 DEG C of superspeed refrigerated centrifuges are by buffer system replacement Treatment Ball mixing and constant-temperature incubation clean magnetic bead and abandon supernatant, then closed with magnetic bead confining liquid after incubation, cleaning magnetic bead is again The anti-HBe being coated with is saved in liquid in magnetic bead and is uprightly saved with 2~8 DEG C of refrigerators by secondary discarding supernatant.
5. detection antihepatitis b e antibody kit as described in claim 1, which is characterized in that the neutralization antigen For HBeAg antigen.
6. detection antihepatitis b e antibody kit as described in claim 1, which is characterized in that described first with group of the lanthanides The antibody in anti-HBe marker that element combines is monoclonal antibody or resists more;Lanthanide series is by intermediate chelating agent in conjunction with anti-HBe It completes, lanthanide series is europium or samarium, and chelating agent is isothiocyanic acid phenyl-EDTA, isothiocyanic acid benzyl-DTTA, the different sulphur cyanogen of P- Base benzyl-DTTA or diethylene triamine pentaacetic acid aminophenyl-EDTA.
7. detecting the preparation method of antihepatitis b e antibody kit described in claim 1, it is characterised in that including walking as follows It is rapid:
(1) preparation is coated with the magnetic microsphere of anti-HBe;
(2) anti-HBe marker of the preparation in conjunction with lanthanide series;
(3) calibration object is prepared;
(4) preparation neutralizes antigen;
(5) analysis buffer, concentration washing lotion and enhancement solution are prepared;
(6) each component is dispensed respectively into corresponding storage container;
(7) RFID card duplicate copy;
(8) coding, labelling;
(9) it is assembled into finished product kit.
CN201811329562.5A 2018-11-09 2018-11-09 A kind of detection antihepatitis b e antibody kit and preparation method thereof Pending CN109541204A (en)

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