CN105891489A - Test paper for detecting HBcAb (hepatitis B core antibody) IgM (immunoglobulin m) in serum and preparation method of test paper - Google Patents
Test paper for detecting HBcAb (hepatitis B core antibody) IgM (immunoglobulin m) in serum and preparation method of test paper Download PDFInfo
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- CN105891489A CN105891489A CN201610218809.0A CN201610218809A CN105891489A CN 105891489 A CN105891489 A CN 105891489A CN 201610218809 A CN201610218809 A CN 201610218809A CN 105891489 A CN105891489 A CN 105891489A
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/576—Immunoassay; Biospecific binding assay; Materials therefor for hepatitis
- G01N33/5761—Hepatitis B
Abstract
The invention provides test paper for detecting HBcAb (hepatitis B core antibody) IgM (immunoglobulin m) in serum and a preparation method. The test paper comprises a bottom plate, a sample pad, a colloidal gold label pad, a detection reaction area and a water absorption pad, wherein an HBcAb specific antigen is adsorbed on the colloidal gold label pad, and a nitrocellulose membrane coated with HBcAb detection lines and quality control lines with mouse anti-human IgM antibodies printed are arranged in the detection reaction area. During preparation, a base and a panel are embedded together, a detection window on the panel corresponds to a test paper core detection reaction area, a sampling hole corresponds to a test paper core sample pad, an HBcAb in the serum of a human body is rapidly detected with an immune colloidal gold method, infection of the HBcAb in the serum is rapidly and conveniently diagnosed, the test paper preparation method is simple and easy to operate, and the test paper is stable in performance, high in specificity and high in sensitivity and can be popularized in the field of clinical examination.
Description
Technical field
The present invention relates to technical field of medical examination, be specifically related to hepatitis B core antibody in a kind of detection serum
(HBcAb) reagent paper of IgM and preparation method thereof.
Background technology
Hepatitis B core antibody is also anti-HBc, and english abbreviation is HBcAb.Including anti-HBcIgM and
Hepatitis B core antibody IgG.After HBV infection, majority's serum can detect this antibody, be a kind of sensitive
Serologic marker, be also the early sign of hepatitis B actute infection, the time is the longest present in the serum,
The hepatitis B core antibody of high titre is also that a kind of infection indicates.In diagnosis of hepatitis b and discriminating, paired sera
Antibody titer compares, and the antibody titer of convalescent serum has 4 times or the growth of more than 4 times than acute stage, can examine
Break as HBV infection.Hepatitis B core antibody also has important meaning in viral hepatitis classification diagnosis.
Hepatitis B core antibody (HBcAb) is the corresponding antibody of hepatitis B virus core antigen, and it is not that protectiveness resists
Body, its existence is affected by one of index of hepatitis B virus infringement on the contrary.It includes IgM, IgA, IgG tri-
Type.IgM is the important indicator judging acute hepatitis b, be after organism infection hepatitis B virus the most the earliest
The specific antibody occurred, Anti-HBc Serum lgM type, it is called " hepatitis B core antibody IgM ", usually points out
For acute HBV infection, or enter active stage, along with the course of disease at chronic viral hepatitis B acute attack, chronic viral hepatitis B
Prolongation or the improvement of the state of an illness, 1gM progressively disappears, and is replaced by IgG, therefore for acute hepatitis B
Virus the infected, detects the key factor that its lgM is therapeutic process.
At present, hepatitis B virus DNA detection is the conventional means judging hepatitis B virus duplication, owing to DNA examines
Survey technology requires strict, is easily subject to interference, and therefore, once result is not enough to describe the problem, and needs repeatedly to examine
Survey, detect cycle length and gradient of infection well can not be detected, the anti-HBcIgM of high titre
Being acute or the important indicator of recent infection, also can be positive at chronic hepatitis of active stage, mark hepatitis B virus exists
Replicating, being infectious, so checking the method for hepatitis B core antibody (HBcAb) IgM facing quickly and efficiently
Certain Research Significance is had on bed.
Summary of the invention
Present invention solves the technical problem that being just to provide one checks hepatitis B core in human serum rapidly and accurately
Test paper of antibody (HBcAb) IgM and preparation method thereof.
The technical scheme is that the reagent paper of hepatitis B core antibody (HBcAb) IgM in a kind of detection serum,
This reagent paper includes base plate, sample pad, colloid gold label pad, detection reaction zone and adsorptive pads, described gold colloidal mark
Note pad is adsorbed with HBcAb specific antigen；Described detection reaction zone is provided with and is coated with hepatitis B core antibody detection
The nitrocellulose filter of line, and the nature controlling line that mouse-anti human IgM antibody is printed；Described it is coated with hepatitis B core
Antibody is hepatitis B virus monoclonal antibody or polyclonal antibody.
Further, the HBcAb specific antigen preparation method of described colloid gold label is: in boiling
150mL0.02～0.03% chlorauric acid solution add freshly prepared 1.5% sodium citrate 5mL, it is thus achieved that diameter
For the colloidal gold solution of 20-30nm, with the potassium carbonate regulation pH value of 0.2mol/L to 8～9, it is placed in 1～5 DEG C
Preserve, standby；Add the colloid of pH8～9 than by antigen of hepatitis B virus to be marked with the labelling of 1:3000
In gold solution, add the PEG10000 of 30% after labelling 15min to final concentration of 0.08%, 5 DEG C, 1800～
3000rpm is centrifuged 30min, removes unconjugated colloid gold particle, 3 DEG C, 15000rpm be centrifuged 1h, remove
Supernatant, it is thus achieved that the colloid gold label thing protein mixture of preliminary purification, then with propylene glucosan S-400 post layer
Analysis carries out isolated and purified, it is thus achieved that the HBcAb specific antigen of colloid gold label.
Further, described base plate is made up of PVC material；Described sample pad glass fibre cotton is made；Institute
State adsorptive pads absorbent filter to make；Described detection reaction zone nitrocellulose filter (NC film) is made；Institute
State colloid gold label pad nylon membrane to make.
In a kind of detection serum, the preparation method of the reagent paper of hepatitis B core antibody (HBcAb) IgM is:
(1) prepared by colloid gold label pad:
A. in the 150mL0.02～0.03% chlorauric acid solution of boiling, freshly prepared 1.5% sodium citrate is added
5mL, it is thus achieved that the colloidal gold solution of a diameter of 20-30nm, with the potassium carbonate of 0.2mol/L regulate pH value to 8～
9, it is placed in 1～5 DEG C of preservation, standby；Add than by antigen of hepatitis B virus to be marked with the labelling of 1:3000
In the colloidal gold solution of pH8～9, after labelling 15min, the PEG10000 of addition 30% is to the most final concentration of
0.08%, 5 DEG C, 1800～3000rpm are centrifuged 30min, remove unconjugated colloid gold particle, 3 DEG C,
15000rpm is centrifuged 1h, removes supernatant, it is thus achieved that the colloid gold label thing protein mixture of preliminary purification, then
Carry out isolated and purified with propylene glucosan S-400 column chromatography, it is thus achieved that the HBcAb specificity of colloid gold label resists
Former；
B. the HBcAb specific antigen of good for labelling gold colloidal is uniformly put and be mapped on nylon membrane, lyophilization,
I.e. prepare colloid gold label pad.
(2) detection reaction zone celluloid film preparation: by the described polyclone being coated with hepatitis B core antibody
Antibody or monoclonal antibody and mouse-anti human IgM antibody are sprayed on the two ends zones of different of nitrocellulose filter respectively,
Form detection line and nature controlling line, obtain, containing detection line and the nitrocellulose filter of nature controlling line, being then placed in being dried
Room is dried；The described polyclonal antibody being coated with hepatitis B core antibody or MAb concentration are 3mg/ml；
Described mouse-anti human IgM antibody concentration is 2mg/ml.
(3) prepared by sample pad: soaked 1.5 hours in the buffer that temperature is 38 DEG C by glass fibre membrane,
Obtain sample pad, be then placed in hothouse being dried；Described buffer refers to phosphate buffer, first by phosphoric acid
Salt is dissolved in 500mL distilled water, corrects pH7.5-8 with 1mol/L sodium hydroxide solution.
(4) prepared by reagent paper: by described sample pad, colloid gold label pad, nitrocellulose filter and adsorptive pads from
Top to bottm it is sequentially fixed on base plate, cuts and obtain hepatitis B core antibody (HBcAb) IgM in a kind of detection serum
Reagent paper, then reagent paper is put into together with desiccant aluminium foil bag and packs, 5 DEG C of storages.
The operation principle of reagent paper of the present invention is: use colloidal gold immunochromatographimethod technology, when containing second in measuring samples
During liver core antibody (HBcAb) IgM, due to capillary action HBcAb IgM under the traction of adsorptive pads
Antibody first combines with the HBcAb specific antigen of colloid gold label on nylon membrane and forms complex, is then combined
Thing is moved forward by chromatography effect, when being up to detect line, runs into mouse-anti human IgM antibody, forms two
The gold mark complex of one antigen of antibodies, because there being gold grain to deposit at this, is gathered on detection band, therefore inspection
The aobvious redness of survey line, can observe by the naked eye colour developing result.
The reagent paper using method of the present invention: take blood by Clinical Laboratory routine and separate serum, takes 100-150ul blood
Clearly, in the sample pad of the reagent paper being added drop-wise to preparation with suction pipe, 8-10 minute sentence read result: contain as in sample
(HBcAb) IgM antibody, then it is through colloid gold label pad, can be combined with HBcAb specific antigen,
And then the mouse-anti human IgM antibody capture on tested survey line, it is gathered in that detection band is upper forms red precipitation, i.e.
For the positive, otherwise it is then negative.
Beneficial effects of the present invention is embodied in: owing to hepatitis B core antibody (HBcAb) IgM is to judge acute second
The important indicator of liver, is the specific antibody occurred the most the earliest after organism infection hepatitis B virus, passes through glue
Whether body gold immunochromatographyassay assay human serum exists (HBcAb) IgM, is possible not only to fast direct ground connection
Judge whether infected hepatitis B virus pathogen, and also can be positive at chronic hepatitis of active stage, indicate second
Hepatovirus is replicating, and is infectious, it is possible to the gradient of infection of hepatitis B virus in detection therapeutic process, this method can
With detection hepatitis B core antibody (HBcAb) IgM the most rapidly and efficiently, it is not required to professional training, easy to operate,
Quickly, it is suitable for clinical expansion and utilization.
Detailed description of the invention
Embodiment 1: the reagent paper of hepatitis B core antibody (HBcAb) IgM, this reagent paper bag in a kind of detection serum
Including base plate, sample pad, colloid gold label pad, detection reaction zone and adsorptive pads, described colloid gold label pad adsorbs
There is HBcAb specific antigen；Described detection reaction zone is provided with the nitric acid being coated with hepatitis B core antibody detection line
Cellulose membrane, and the nature controlling line that mouse-anti human IgM antibody is printed；The described hepatitis B core antibody that is coated with is second
Hepatovirus polyclonal antibody.
Wherein, the HBcAb specific antigen preparation method of described colloid gold label is: in boiling
150mL0.02% chlorauric acid solution adds freshly prepared 1.5% sodium citrate 5mL, it is thus achieved that a diameter of
The colloidal gold solution of 20nm, with the potassium carbonate regulation pH value of 0.2mol/L to 8, is placed in 1 DEG C of preservation, standby；
With the labelling of 1:3000 ratio antigen of hepatitis B virus to be marked added in the colloidal gold solution of pH8, labelling
Add the PEG10000 of 30% after 15min to final concentration of 0.08%, 5 DEG C, 1800rpm be centrifuged 30min,
Remove unconjugated colloid gold particle, 3 DEG C, 15000rpm be centrifuged 1h, remove supernatant, it is thus achieved that the purest
The colloid gold label thing protein mixture changed, then carry out isolated and purified with propylene glucosan S-400 column chromatography, obtain
Obtain the HBcAb specific antigen of colloid gold label.
The reagent paper of hepatitis B core antibody (HBcAb) IgM in a kind of detection serum, described PVC base plate is purchased from
Shanghai Jinbiao Bio-Tech Co., Ltd.；Described sample pad glass fibre cotton is made；Described adsorptive pads absorbs water
Filter paper is made；Described detection reaction zone nitrocellulose filter (NC film) is made；Described colloid gold label pad
Make with nylon membrane.
In a kind of detection serum, the preparation method of the reagent paper of hepatitis B core antibody (HBcAb) IgM is:
(1) prepared by colloid gold label pad:
A. in the 150mL0.02% chlorauric acid solution of boiling, freshly prepared 1.5% sodium citrate 5mL is added,
Obtain the colloidal gold solution of a diameter of 20, with the potassium carbonate regulation pH value of 0.2mol/L to 8, be placed in 1 DEG C of guarantor
Deposit, standby；The gold colloidal that antigen of hepatitis B virus to be marked adds pH8 with the labelling of 1:3000 ratio is molten
In liquid, after labelling 15min, add the PEG10000 to final concentration of 0.08% of 30%, 5 DEG C, 1800rpm
Centrifugal 30min, removes unconjugated colloid gold particle, 3 DEG C, 15000rpm be centrifuged 1h, remove supernatant,
Obtain the colloid gold label thing protein mixture of preliminary purification, then carry out point with propylene glucosan S-400 column chromatography
From purification, it is thus achieved that the HBcAb specific antigen of colloid gold label；
B. the HBcAb specific antigen of good for labelling gold colloidal is uniformly put and be mapped on nylon membrane, lyophilization,
I.e. prepare colloid gold label pad.
(2) detection reaction zone celluloid film preparation: by the described polyclone being coated with hepatitis B core antibody
Antibody and mouse-anti human IgM antibody are sprayed on the two ends zones of different of nitrocellulose filter respectively, formed detection line and
Nature controlling line, obtains containing detection line and the nitrocellulose filter of nature controlling line, is then placed in hothouse being dried；Institute
Stating and being coated with the polyclonal antibody of hepatitis B core antibody is 3mg/ml；Described mouse-anti human IgM antibody concentration is
2mg/ml。
(3) prepared by sample pad: soaked 1.5 hours in the buffer that temperature is 38 DEG C by glass fibre membrane,
Obtain sample pad, be then placed in hothouse being dried；Described buffer refers to phosphate buffer, first by phosphoric acid
Salt is dissolved in 500mL distilled water, corrects pH7.5 with 1mol/L sodium hydroxide solution.
(4) prepared by reagent paper: by described sample pad, colloid gold label pad, nitrocellulose filter and adsorptive pads from
Top to bottm it is sequentially fixed on base plate, cuts and obtain hepatitis B core antibody (HBcAb) IgM in a kind of detection serum
Reagent paper, then reagent paper is put into together with desiccant aluminium foil bag and packs, 5 DEG C of storages.
Embodiment 2: the reagent paper of hepatitis B core antibody (HBcAb) IgM, this reagent paper bag in a kind of detection serum
Including base plate, sample pad, colloid gold label pad, detection reaction zone and adsorptive pads, described colloid gold label pad adsorbs
There is HBcAb specific antigen；Described detection reaction zone is provided with the nitric acid being coated with hepatitis B core antibody detection line
Cellulose membrane, and the nature controlling line that mouse-anti human IgM antibody is printed；The described hepatitis B core antibody that is coated with is second
Hepatovirus polyclonal antibody.
Wherein, the HBcAb specific antigen preparation method of described colloid gold label is: in boiling
150mL0.025% chlorauric acid solution adds freshly prepared 1.5% sodium citrate 5mL, it is thus achieved that a diameter of
The colloidal gold solution of 25nm, with the potassium carbonate regulation pH value of 0.2mol/L to 8.5, is placed in 2.5 DEG C of preservations,
Standby；With the labelling of 1:3000 ratio antigen of hepatitis B virus to be marked added in the colloidal gold solution of pH8.5,
The PEG10000 of 30% is added to final concentration of 0.08% after labelling 15min, 5 DEG C, 2400rpm is centrifuged
30min, removes unconjugated colloid gold particle, 3 DEG C, 15000rpm be centrifuged 1h, remove supernatant, it is thus achieved that
The colloid gold label thing protein mixture of preliminary purification, then carry out separating pure with propylene glucosan S-400 column chromatography
Change, it is thus achieved that the HBcAb specific antigen of colloid gold label.
The reagent paper of hepatitis B core antibody (HBcAb) IgM in a kind of detection serum, described PVC base plate is purchased from
Shanghai Jinbiao Bio-Tech Co., Ltd.；Described sample pad glass fibre cotton is made；Described adsorptive pads absorbs water
Filter paper is made；Described detection reaction zone nitrocellulose filter (NC film) is made；Described colloid gold label pad
Make with nylon membrane.
In a kind of detection serum, the preparation method of the reagent paper of hepatitis B core antibody (HBcAb) IgM is:
(1) prepared by colloid gold label pad:
A. in the 150mL0.025% chlorauric acid solution of boiling, freshly prepared 1.5% sodium citrate is added
5mL, it is thus achieved that the colloidal gold solution of a diameter of 25nm, with the potassium carbonate regulation pH value of 0.2mol/L to 8.5,
It is placed in 2.5 DEG C of preservations, standby；PH8.5 is added than by antigen of hepatitis B virus to be marked with the labelling of 1:3000
Colloidal gold solution in, add the PEG10000 of 30% after labelling 15min to final concentration of 0.08%, 5 DEG C,
2400rpm is centrifuged 30min, removes unconjugated colloid gold particle, 3 DEG C, 15000rpm be centrifuged 1h, remove
Supernatant, it is thus achieved that the colloid gold label thing protein mixture of preliminary purification, then with propylene glucosan S-400 post layer
Analysis carries out isolated and purified, it is thus achieved that the HBcAb specific antigen of colloid gold label；
B. the HBcAb specific antigen of good for labelling gold colloidal is uniformly put and be mapped on nylon membrane, lyophilization,
I.e. prepare colloid gold label pad.
(2) detection reaction zone celluloid film preparation: by the described polyclone being coated with hepatitis B core antibody
Antibody and mouse-anti human IgM antibody are sprayed on the two ends zones of different of nitrocellulose filter respectively, formed detection line and
Nature controlling line, obtains containing detection line and the nitrocellulose filter of nature controlling line, is then placed in hothouse being dried；Institute
Stating and being coated with the Anti-TNF-α bulk concentration of hepatitis B core antibody is 3mg/ml；Described mouse-anti human IgM antibody concentration
For 2mg/ml.
(3) prepared by sample pad: soaked 1.5 hours in the buffer that temperature is 38 DEG C by glass fibre membrane,
Obtain sample pad, be then placed in hothouse being dried；Described buffer refers to phosphate buffer, first by phosphoric acid
Salt is dissolved in 500mL distilled water, corrects pH7.75 with 1mol/L sodium hydroxide solution.
(4) prepared by reagent paper: by described sample pad, colloid gold label pad, nitrocellulose filter and adsorptive pads from
Top to bottm it is sequentially fixed on base plate, cuts and obtain hepatitis B core antibody (HBcAb) IgM in a kind of detection serum
Reagent paper, then reagent paper is put into together with desiccant aluminium foil bag and packs, 5 DEG C of storages.
Embodiment 3: the reagent paper of hepatitis B core antibody (HBcAb) IgM, this reagent paper bag in a kind of detection serum
Including base plate, sample pad, colloid gold label pad, detection reaction zone and adsorptive pads, described colloid gold label pad adsorbs
There is HBcAb specific antigen；Described detection reaction zone is provided with the nitric acid being coated with hepatitis B core antibody detection line
Cellulose membrane, and the nature controlling line that mouse-anti human IgM antibody is printed；The described hepatitis B core antibody that is coated with is second
Hepatovirus monoclonal antibody.
Wherein, the HBcAb specific antigen preparation method of described colloid gold label is: in boiling
150mL0.03% chlorauric acid solution adds freshly prepared 1.5% sodium citrate 5mL, it is thus achieved that a diameter of
The colloidal gold solution of 30nm, with the potassium carbonate regulation pH value of 0.2mol/L to 9, is placed in 5 DEG C of preservations, standby；
With the labelling of 1:3000 ratio antigen of hepatitis B virus to be marked added in the colloidal gold solution of pH9, labelling
Add the PEG10000 of 30% after 15min to final concentration of 0.08%, 5 DEG C, 3000rpm be centrifuged 30min,
Remove unconjugated colloid gold particle, 3 DEG C, 15000rpm be centrifuged 1h, remove supernatant, it is thus achieved that the purest
The colloid gold label thing protein mixture changed, then carry out isolated and purified with propylene glucosan S-400 column chromatography, obtain
Obtain the HBcAb specific antigen of colloid gold label.
The reagent paper of hepatitis B core antibody (HBcAb) IgM in a kind of detection serum, described PVC base plate is purchased from
Shanghai Jinbiao Bio-Tech Co., Ltd.；Described sample pad glass fibre cotton is made；Described adsorptive pads absorbs water
Filter paper is made；Described detection reaction zone nitrocellulose filter (NC film) is made；Described colloid gold label pad
Make with nylon membrane.
In a kind of detection serum, the preparation method of the reagent paper of hepatitis B core antibody (HBcAb) IgM is:
(1) prepared by colloid gold label pad:
A. in the 150mL0.03% chlorauric acid solution of boiling, freshly prepared 1.5% sodium citrate 5mL is added,
Obtain the colloidal gold solution of a diameter of 30nm, with the potassium carbonate regulation pH value of 0.2mol/L to 9, be placed in 5 DEG C
Preserve, standby；Add the gold colloidal of pH9 than by antigen of hepatitis B virus to be marked with the labelling of 1:3000
In solution, after labelling 15min, add the PEG10000 to final concentration of 0.08% of 30%, 5 DEG C, 3000rpm
Centrifugal 30min, removes unconjugated colloid gold particle, 3 DEG C, 15000rpm be centrifuged 1h, remove supernatant,
Obtain the colloid gold label thing protein mixture of preliminary purification, then carry out point with propylene glucosan S-400 column chromatography
From purification, it is thus achieved that the HBcAb specific antigen of colloid gold label；
B. the HBcAb specific antigen of good for labelling gold colloidal is uniformly put and be mapped on nylon membrane, lyophilization,
I.e. prepare colloid gold label pad.
(2) detection reaction zone celluloid film preparation: by the described monoclonal being coated with hepatitis B core antibody
Antibody and mouse-anti human IgM antibody are sprayed on the two ends zones of different of nitrocellulose filter respectively, formed detection line and
Nature controlling line, obtains containing detection line and the nitrocellulose filter of nature controlling line, is then placed in hothouse being dried；Institute
Stating and being coated with the MAb concentration of hepatitis B core antibody is 3mg/ml；Described mouse-anti human IgM antibody concentration
For 2mg/ml.
(3) prepared by sample pad: soaked 1.5 hours in the buffer that temperature is 38 DEG C by glass fibre membrane,
Obtain sample pad, be then placed in hothouse being dried；Described buffer refers to phosphate buffer, first by phosphoric acid
Salt is dissolved in 500mL distilled water, corrects pH8 with 1mol/L sodium hydroxide solution.
(4) prepared by reagent paper: by described sample pad, colloid gold label pad, nitrocellulose filter and adsorptive pads from
Top to bottm it is sequentially fixed on base plate, cuts and obtain hepatitis B core antibody (HBcAb) IgM in a kind of detection serum
Reagent paper, then reagent paper is put into together with desiccant aluminium foil bag and packs, 5 DEG C of storages.
Clinical statistics is tested:
Clinical patients hepatitis B core antibody (HBcAb) IgM detects: collects the 80 doubtful hepatitis B virus infections of example and suffers from
Person's serum sample, 20 example hepatitis B virus infection patients diagnosed, the hepatitis B virus DNA detection method conventional with tradition
Relatively, test strips method of the present invention has 1 example suspected case IgM antibody to fail to detect.Its sensitivity is 99.0%,
Specificity is 100%.Testing result is as shown in table 1.
Table 1:DNA method is added up with ELISA test strip result of the present invention
Result of the test shows: the test strip of the present invention has higher sensitivity and specificity, is especially suitable for
Clinical practice.
Last it is noted that above example is only in order to illustrate technical scheme, it is not intended to limit；
Although being described in detail the present invention with reference to previous embodiment, those of ordinary skill in the art should manage
Solve: the technical scheme described in previous embodiment still can be modified by it, or to wherein portion of techniques
Feature carries out equivalent；And these amendments or replacement, do not make the essence of appropriate technical solution depart from this
The spirit and scope of bright embodiment technical scheme.
Claims (4)
1. a reagent paper of hepatitis B core antibody (HBcAb) IgM in detection serum, this reagent paper include base plate,
Sample pad, colloid gold label pad, detection reaction zone and adsorptive pads, it is characterised in that: described colloid gold label pad
It is adsorbed with HBcAb specific antigen；Described detection reaction zone is provided with and is coated with hepatitis B core antibody detection line
Nitrocellulose filter, and the nature controlling line that mouse-anti human IgM antibody is printed；Described it is coated with hepatitis B core antibody
For hepatitis B virus monoclonal antibody or polyclonal antibody.
2. the examination of hepatitis B core antibody (HBcAb) IgM in a kind of detection serum as claimed in claim 1
Paper, it is characterised in that the HBcAb specific antigen preparation method of described colloid gold label is: in boiling
150mL0.02～0.03% chlorauric acid solution add freshly prepared 1.5% sodium citrate 5mL, it is thus achieved that colloid
Gold solution, regulates pH value with potassium carbonate, and cryopreservation is standby；With the labelling ratio of 1:3000 by be marked
Antigen of hepatitis B virus add preparation colloidal gold solution in, after labelling 15min add 30% PEG10000
To final concentration of 0.08%, it is centrifuged off unconjugated colloid gold particle, is centrifuged off supernatant, it is thus achieved that be preliminary
The colloid gold label thing protein mixture of purification, then carry out isolated and purified with propylene glucosan S-400 column chromatography,
Obtain the HBcAb specific antigen of colloid gold label.
3. the examination of hepatitis B core antibody (HBcAb) IgM in a kind of detection serum as claimed in claim 1
Paper, it is characterised in that described base plate is made up of PVC material；Described sample pad glass fibre cotton is made；Institute
State adsorptive pads absorbent filter to make；Described detection reaction zone nitrocellulose filter is made；Described gold colloidal mark
Note pad nylon membrane is made.
4. the examination of hepatitis B core antibody (HBcAb) IgM in a kind of detection serum as claimed in claim 1
The preparation method of paper is:
(1) prepared by colloid gold label pad:
A. in the 150mL0.02～0.03% chlorauric acid solution of boiling, freshly prepared 1.5% sodium citrate is added
5mL, it is thus achieved that colloidal gold solution, regulates pH value with potassium carbonate, and cryopreservation is standby；With 1:3000's
Labelling adds than by antigen of hepatitis B virus to be marked in the colloidal gold solution of preparation, adds after labelling 15min
The PEG10000 of 30% to final concentration of 0.08%, is centrifuged off unconjugated colloid gold particle, is centrifuged off
Supernatant, it is thus achieved that the colloid gold label thing protein mixture of preliminary purification, then with propylene glucosan S-400 post layer
Analysis carries out isolated and purified, it is thus achieved that the HBcAb specific antigen of colloid gold label；
B. the HBcAb specific antigen of good for labelling gold colloidal is uniformly put and be mapped on nylon membrane, lyophilization,
I.e. prepare colloid gold label pad.
(2) detection reaction zone celluloid film preparation: by the described B-type hepatitis being coated with hepatitis B core antibody
Poison polyclonal antibody or monoclonal antibody and mouse-anti human IgM antibody are sprayed on the two ends of nitrocellulose filter not respectively
Same region, forms detection line and nature controlling line, obtains containing detection line and the nitrocellulose filter of nature controlling line, then
Put in hothouse and be dried.
(3) prepared by sample pad: soaked 1.5 hours in the buffer that temperature is 38 DEG C by glass fibre membrane,
Obtain sample pad, be then placed in hothouse being dried.
(4) prepared by reagent paper: by described sample pad, colloid gold label pad, nitrocellulose filter and adsorptive pads from
Top to bottm it is sequentially fixed on base plate, cuts and obtain hepatitis B core antibody (HBcAb) IgM in a kind of detection serum
Reagent paper, then reagent paper is put into together with desiccant aluminium foil bag and packs, 5 DEG C of storages.
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| CN109239337A (en) * | 2018-09-27 | 2019-01-18 | 天津欣普赛尔生物医药科技有限公司 | The test paper and preparation method of bovine serum albumin(BSA) in a kind of detection biological products |
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