CN102010468A - Toxoplasma circulating antigen double antibody sandwich ELISA (Enzyme-Linked Immuno Sorbent Assay) detection method - Google Patents
Toxoplasma circulating antigen double antibody sandwich ELISA (Enzyme-Linked Immuno Sorbent Assay) detection method Download PDFInfo
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Abstract
The invention discloses a toxoplasma circulating antigen double antibody sandwich ELISA (Enzyme-Linked Immuno Sorbent Assay) detection reagent and a preparation method. Toxoplasma MIC10 is a protein secreted by toxoplasma when invading a host cell and has important actions in invasion and proliferation of the toxoplasma. In the invention, amplification and expression at different areas are carried out on an MIC10 gene to obtain recombinant fusion proteins MIC10A, MIC10B and MIC10C, the purified recombinant fusion proteins MIC10A and MIC10B respectively immunize rabbits, are used for preparing a polyclonal antibody and are respectively used as a coated antibody and a detection antibody, a double antibody sandwich ELISA is established with purified MIC10C as a standard antibody to detect a toxoplasma circulating antigen in serum and has the advantages of strong specificity, strong sensitivity, strong reproducibility and strong stability, wherein y is equal to 0.0304Ln(x)-0.0567, R2 is equal to 0.9643, and the minimum detectable amount is 50pg/ml.
Description
Technical field
The invention belongs to biological test quarantine technical field, specifically, make up recombinant protein, utilize the antibody of recombinant protein immunity preparation, set up a kind of arch insect circulating antigen double antibodies sandwich ELISA detection method.
Background technology
Toxoplasma gondii (toxoplasma) is a kind of special sexual cell endoparasitism protozoon, has host group and worldwide popular widely, causes that the people beast suffers from parasitosis altogether.Damnification of immunity function or inhibition persons such as AIDS patient, organ transplantation and malignant tumor patient, toxoplasma gondii can cause death as a kind of opportunistic infection factor.Toxoplasmosis is medically a kind of important diseases still not, the more important thing is that it is an important biomolecule factor of the human prenatal and postnatal care of influence.
Human infection toxoplasma, majority may be asymptomatic carrier, only a few peoples morbidity.This disease complicated clinical manifestation, the lighter is inapparent infection, weight person can show as the grievous injury of many organs, as the pathology of each system such as toxoplasma gondii encephalopathic, illness in eye, ephrosis, hepatopathy, tuberculosis and toxoplasma myocarditis.It should be noted that toxoplasma has very significant effects to gestation.Pregnant woman by toxoplasma infects no matter it has or not clinical symptom, often can pass to fetus with toxoplasma gondii by placenta, thereby directly influence the growth of fetus, makes the serious teratogenesis of fetus, even dead, also can miscarry, stillbirth, premature labor or increase pregnancy complication.Infection takes place more early, and fetus is impaired serious more.The second trimester of pregnancy infect, stillborn foetus, premature labor and serious brain, eye illness how can occur and suffer from; In third trimester of pregnancy, ripe gradually because of fetus, parent is as being infected at this moment, and fetus can grow normally, also can occur symptom just occurring after premature labor or the birth, shows as the damage in various degree of each system.It is reported that the fetus of accidental pregnancy period toxoplasma gondii infection just shows toxoplasmosis behind several years after the birth even adult.
The whole world 1/4th populations are subjected to the threat of toxoplasmosis according to statistics, and the U.S. has 400-4000 example congenital toxoplasmosis to take place every year.There are more than 6,100 ten thousand people's toxoplasma gondii infection diseases in China, wherein the women of child-bearing age have 1,300 ten thousand-1,500 ten thousand, toxoplasma gondii is one of pathogenic agent of intrauterine infection, annual have 90,000 be born by the newborn infant of toxoplasma gondii infringement approximately, pregnant woman's arch insect infection rate is 6.6%-32.9%, by the placental infection fetus, have a strong impact on embryo growth and development.Aspect livestock industry, miscarriage appears in pregnant animal, and the pig toxoplasma gondii infection can cause large quantities of death.
As seen toxoplasma gondii is very harmful to the mankind, pregnant woman especially, before pregnant and pregnancy period (first trimester the is better) serology of preferably doing toxoplasma gondii detect.
ELISA is the most widely used technology of current diagnosis arch insect infection, and current commercial test kit mainly comprises detection antibody and detects circulating antigen two big classes.Wherein antibody test comprises two kinds of IgG detection kit and IgM detection kit again.Because IgM antibody is to occur in early days, and the residence time is shorter, detect the early diagnosis that IgM antibody is used to multiple disease in serum; And the IgG appearance is later, is the sign of chronic phase or previous infection.The disadvantage that IgG detects is to judge current infection or previous infection.With current infection good dependency is arranged although IgM detects, be not suitable for the detection of immunologic hypofunction crowd and pregnancy period, this is that detection sensitivity is low, causes false negative because these crowd's body's immunities are low.
Circulating antigen (CAG) is that toxoplasma gondii acute infection period polypide self drains and split product, in the intrusion and reproductive process of polypide, is secreted out by polypide, therefore detects the infection that CAG can diagnose toxoplasma gondii more exactly.And MIC10 is proved to be a kind of major protein in the excretory circulating antigen (CAG) in the polypide course of infection, polypide secretes this albumen in a large number in substratum during vitro culture, therefore detecting this albumen not only can be qualitative to infecting, but also can be by this protein quantification is determined infection intensity.
Summary of the invention
The purpose of this invention is to provide recombinant protein MIC-10.
Described recombinant protein MIC-10 is MIC-10A, MIC-10B or MIC-10C, its nucleotide sequence successively respectively as: shown in the SEQ ID No.1, shown in the SEQ ID No.2 or shown in the SEQ ID No.3;
The preparation method of recombinant protein MIC-10, it comprises:
1) synthetic primer;
Preparation recombinant protein MIC-10A primer is:
FW：GATCGGATCCGATTTTTTTAACGATTAC
RV：GATCAAGCTTTCATTCACGAAGTCCTCTTTTC
Preparation recombinant protein MIC-10B primer is:
FW：GATCGGATCCGCCGCTGAGCGTGAGGAGAAG
RV：GATCAAGCTTTCACATTGATTTCCTGCG
Preparation recombinant protein MIC-10C primer is:
FW：GATCGGATCCGATTTTTTTAACGATTAC
RV：GATCAAGCTTTCACATTGATTTCCTGCG
2) extracting RH strain toxoplasma cdna group DNA is template, carries out pcr amplification, obtains gene fragment, is cloned on the PMD-18T carrier;
3) the MIC10A gene fragment is inserted structure recombinant expression plasmid PET28A-MIC-10A, PET28A-MIC-10B or PET28A-MIC-10C among the expression vector PET28A;
4) recombinant expression plasmid PET28A-MIC10A changed among the BL21 (DE3) express, recombination fusion protein MIC-10A, MIC-10B or MIC-10C, use the ni-sepharose purification recombinant protein.
Anti-recombinant protein MIC-10 polyclonal antibody is prepared by following method:
With recombinant protein MIC-10 immunity 3 month female rabbit, subcutaneous 3 injections, immunizing dose 500ug//time, 21d immunity at interval, immunity is 3 times altogether, the 1st employing Freund's complete adjuvant, 2nd, adopt Freund's incomplete adjuvant, the most immune back 21 days, heart bloodletting 3 times, place after 1 hour for 37 ℃, 4 ℃ are spent the night, and leave the heart 15 minutes next day 3000, get supernatant, with the anti-MIC10 of protein A chromatography column purified rabbit, by specification carries out.IgG behind the purifying measures protein concentration with PBS dialysis 3 times with the BCA method, and-80 ℃ frozen;
Described recombinant protein MIC-10 is MIC-10A or MIC-10B.
A kind of arch insect circulating antigen double antibodies sandwich ELISA detection kit, it comprises: coated antibody, biotin labeled detection antibody, positive control, washings, confining liquid, enzyme mark avidin, substrate, stop buffer and sample diluent is characterized in that: described coated antibody is that the anti-recombinant protein MIC-10AIgG of rabbit, detection antibody are that anti-recombinant protein MIC-10BIgG of rabbit or coated antibody are the anti-recombinant protein MIC-10BIgG of rabbit, detect antibody for exempting from anti-recombinant protein MIC-10AIgG; Described positive control is recombinant protein MIC-10C;
Described confining liquid is the TBS that contains 1%BSA, 10% horse serum, and substrate is TMB, and stop buffer is a 2M sulfuric acid.
A kind of arch insect circulating antigen double antibodies sandwich ELISA detection kit using method, it comprises:
1) the anti-reorganization of the 5ug/ml rabbit MIC-10AIgG of adding 100 μ l purifying wraps by the ELISA Sptting plate 4 ℃ of overnight incubation;
2) wash 5 times each 5 minutes with TBST;
3) every hole adds the sealing of 300 μ l confining liquids, hatches 30 minutes for 37 ℃;
4) wash 5 times each 5 minutes with TBST;
5) every hole adds the different dilution serum samples to be checked of 100 μ l, hatches 1 hour for 37 ℃;
6) wash 5 times each 5 minutes with TBST;
7) every hole adds the anti-MIC-10BIgG of the biotin labeled rabbit of 100 μ l, hatches 1 hour for 37 ℃;
8) wash 5 times each 5 minutes with TBST;
9) add enzyme mark avidin, hatched 30 minutes for 37 ℃;
10) wash 5 times with TBST, each 5 minutes, hatched 10 minutes;
11) every hole adds the 100ul substrate, room temperature lucifuge colour developing 30 minutes; Every hole adds 50ul 2M sulfuric acid termination reaction.
12) read plate, record OD value; According to typical curve, calculate the MIC10 content in the serum sample to be checked;
Described coated antibody is that the anti-MIC-10BIgG of rabbit, detection antibody are the anti-recombinant protein MIC-10AIgG of rabbit.
A kind of arch insect circulating antigen double antibodies sandwich of the present invention ELISA detection reagent and preparation method.Toxoplasma gondii microneme protein 10 (MIC10) is toxoplasma gondii a kind of albumen of excretory when invading host cell, and the intrusion and the propagation of toxoplasma gondii is had important effect.The present invention is the different zones amplification to the MIC10 gene, insert among the expression vector PET28A and make up recombinant expression plasmid, changing recombinant expression plasmid over to prokaryotic cell prokaryocyte expresses, obtain recombination fusion protein MIC10A, MIC10B and MIC10C, with recombination fusion protein MIC10A, MIC10B behind purifying difference immunizing rabbit, the high polyclonal antibody of tiring of preparation.With the anti-MIC10A IgG of the rabbit behind the purifying as coated antibody, with the anti-MIC10B IgG of biotin labeled rabbit as detecting antibody, with the MIC10C of purifying as standard antigen, with the serum of the humans and animals of certain extension rate as detecting sample, set up double antibodies sandwich ELISA,, have very strong specificity, susceptibility, repeatability, stability as detection method to arch insect circulating antigen, y=0.0304Ln (x)-0.0567, R
2=0.9643, minimum detectable activity is 50pg/ml.
Description of drawings
Fig. 1 is the pcr amplification qualification result of MIC-10A and MIC-10B gene, 1:MIC10-A wherein, 2:MIC10-B.
Fig. 2 cuts qualification result for the enzyme of MIC-10A and MIC-10B recombinant plasmid, 1:MIC10-A wherein, 2:MIC10-B.
Fig. 3 is the graph of a relation of recombinant protein MIC-10A, MIC-10B, MIC-10C and MIC10 full-length gene (its base preface is shown in SEQ IDNo.4).
Fig. 4 is the typical curve of double antibodies sandwich ELISA method.
Embodiment
The preparation of embodiment 1 recombinant protein MIC-10A
1) synthetic following primer:
FW：GATC
GGATCCGATTTTTTTAACGATTAC
RV：GATC
AAGCTTTCATTCACGAAGTCCTCTTTTC
2) extracting RH strain toxoplasma cdna group DNA is template, carries out pcr amplification, and amplification condition is: 95 ℃ of pre-sex change 2 minutes, and 94 ℃ of sex change 50 seconds, 50 ℃ of annealing 1 minute, 72 ℃ were extended 1 minute, 35 circulations, last 72 ℃ are extended 6min., obtain the MIC-10A gene fragment, see accompanying drawing 1, to be cloned on the PMD-18T carrier, enzyme is cut evaluation, sees accompanying drawing 2, and its nucleotide sequence is shown in SEQ ID No.1;
3) the MIC10A gene fragment is inserted structure recombinant expression plasmid PET28A-MIC-10A among the expression vector PET28A;
4) recombinant expression plasmid PET28A-MIC10A changed among the BL21 (DE3) express, recombinant protein MIC-10A, use the ni-sepharose purification recombinant protein.
The preparation of embodiment 2 recombinant protein MIC-10B
1) synthetic following primer:
FW：GATCGGATCCGCCGCTGAGCGTGAGGAGAAG
RV：GATCAAGCTTTCACATTGATTTCCTGCG
2) extracting RH strain toxoplasma cdna group DNA is template, carries out pcr amplification, and amplification condition is: 95 ℃ of pre-sex change 2 minutes, 94 ℃ of sex change 50 seconds, 50 ℃ of annealing 1 minute, 72 ℃ were extended 1 minute, 35 circulations, last 72 ℃ are extended 6min, obtain the MIC-10B gene fragment, see accompanying drawing 1, be cloned on the PMD-18T carrier, enzyme is cut evaluation, sees accompanying drawing 2, and its nucleotide sequence is shown in SEQ ID No.2;
3) the MIC10B gene fragment is inserted structure recombinant expression plasmid PET28A-MIC-10B among the expression vector PET28A; Enzyme is cut evaluation, sees accompanying drawing 2;
4) recombinant expression plasmid PET28A-MIC-10A changed among the BL21 (DE3) express, recombination fusion protein MIC-10B, use the ni-sepharose purification recombinant protein.
The preparation of embodiment 3 recombination fusion protein MIC-10C
1) synthetic following primer:
FW：GATCGGATCCGATTTTTTTAACGATTAC
RV：GATCAAGCTTTCACATTGATTTCCTGCG
2) extracting RH strain toxoplasma cdna group DNA is template, carries out pcr amplification, and amplification condition is: 95 ℃ of pre-sex change 2 minutes, 94 ℃ of sex change 50 seconds, 50 ℃ of annealing 1 minute, 72 ℃ were extended 1 minute, 35 circulations, last 72 ℃ were extended 6 minutes, and obtained the MIC10C gene fragment; Be cloned on the PMD-18T carrier, enzyme is cut evaluation, and its nucleotide sequence is shown in SEQ IDNo.2;
3) the MIC10C gene fragment is inserted structure recombinant expression plasmid PET28A-MIC-10C among the expression vector PET28A;
4) recombinant expression plasmid PET28A-MIC-10C changed among the BL21 (DE3) express, recombination fusion protein MIC-10C, use the ni-sepharose purification recombinant protein.
The preparation of embodiment 4 anti-MIC-10A and MIC-10B rabbit polyclonal antibody
Immunity: use MIC-10A and MIC-10B immunity 3 month female New Zealand large ear rabbits respectively, subcutaneous 3 injections, immunizing dose 500ug//time, 21d immunity at interval, immunity is 3 times altogether, the 1st employing Freund's complete adjuvant, 2nd, adopt Freund's incomplete adjuvant 3 times, back 21 days of last immunity, the heart bloodletting is placed after 1 hour for 37 ℃, 4 ℃ are spent the night, left the heart 15 minutes in the 2nd day 3000, and got supernatant ,-80 ℃ frozen standby.
The purifying of embodiment 5 anti-MIC10A and MIC10B rabbit polyclonal antibody
With protein A anti-MIC-10AIgG of (GE company) chromatography column purified rabbit and MIC-10B IgG, by specification carries out, and the IgG behind the purifying measures protein concentration with PBS dialysis 3 times with the BCA method.
The biotinylation of the anti-MIC-10B lgG of embodiment 6 rabbits
Anti-MIC-10A of rabbit or B IgG with embodiment 5 makes use biotin labeling, and labelling kit adopts
(Pierce, USA), by specification carries out Sulfo-NHS-LC-Biotinylation Kit.
The foundation of the typical curve of embodiment 7 double antibodies sandwich ELISA methods
1. the anti-MIC-10A of rabbit (5ug/ml) IgG that adds 100 μ l purifying wraps by the ELISA Sptting plate 4 ℃ of overnight incubation;
2, wash 5 times each 5 minutes with TBST;
3, every hole adds 300 μ l confining liquids (TBS of 1%BSA, 10% horse serum), hatches 30 minutes for 37 ℃;
4. wash 5 times each 5 minutes with TBST
5. every hole adds the different dilution MIC-10C (standard model) of 100 μ l, hatches 1 hour for 37 ℃;
6. wash 5 times each 5 minutes with TBST
7. every hole adds the anti-MIC-10BIgG of the biotin labeled rabbit of 100 μ l, hatches 1 hour for 37 ℃;
8. wash 5 times each 5 minutes with TBST;
9. add enzyme mark avidin, hatched 30 minutes for 37 ℃;
10. wash 5 times with TBST, each 5 minutes, hatched 10 minutes;
Annotate: wash with TBS (not containing Tween20) for last 1 time
11. every hole adds 100ul substrate (TMB), room temperature lucifuge colour developing 30 minutes; Every hole adds 50ul 2M sulfuric acid termination reaction.
12.450nm read plate, record OD value, the drawing standard curve is seen Fig. 4.
Prove that by experiment MIC10C concentration and absorbance that present method is measured exist tangible linear relationship, coefficient R in the scope of 50-5000pg/ml
2=0.9643, y=0.0304Ln (x)-0.0567, minimum detectable activity are 50pg/ml.Show that present method has very high susceptibility, satisfy the needs that detect clinical sample fully.
8 one kinds of arch insect circulating antigen double antibodies sandwiches of embodiment ELISA detection kit
It comprises: coated antibody, biotin labeled detection antibody, positive control, washings, confining liquid, enzyme mark avidin, substrate (TMB), stop buffer, described coated antibody are that the anti-recombinant protein MIC-10AIgG of rabbit, detection antibody are that anti-MIC-10BIgG of biotin labeled rabbit or coated antibody are that the anti-recombinant protein MIC-10BIgG of rabbit, detection antibody are biotin labeled recombinant protein MIC-10A; Described positive control is recombinant protein MIC-10C;
Described confining liquid is the TBS that contains 1%BSA, 10% horse serum, and substrate is TMB, and stop buffer is a 2M sulfuric acid.
9 one kinds of arch insect circulating antigen double antibodies sandwiches of embodiment ELISA detection kit using method
1) the anti-MIC-10AIgG of rabbit (5ug/ml) that adds 100 μ l purifying wraps by the ELISA Sptting plate 4 ℃ of overnight incubation;
2) wash 5 times each 5 minutes with TBST;
3) every hole adds the TBS sealing of 300 μ l 1%BSA, 10% horse serum, hatches 30 minutes for 37 ℃;
4) wash 5 times each 5 minutes with TBST
5) every hole adds the different dilution serum samples to be checked of 100 μ l, hatches 1 hour for 37 ℃;
6) wash 5 times each 5 minutes with TBST
7) every hole adds the anti-MIC-10BIgG of the biotin labeled rabbit of 100 μ l, hatches 1 hour for 37 ℃;
8) wash 5 times each 5 minutes with TBST;
9) add enzyme mark avidin, hatched 30 minutes for 37 ℃;
10) wash 5 times with TBST, each 5 minutes, hatched 10 minutes;
Annotate: last wash for 1 time also can be with TBS (not containing Tween20)
11) every hole adds 100ul substrate (TMB), room temperature lucifuge colour developing 30 minutes; Every hole adds 50ul 2M sulfuric acid termination reaction.
12) 450nm reads plate, record OD value; According to typical curve, calculate the MIC10 content in the serum sample to be checked.
10 1 kinds of arch insect circulating antigen double antibodies sandwiches of embodiment ELISA detection kit using method
Coated antibody is that the anti-recombinant protein MIC-10BIgG of rabbit, detection antibody are biotin labeled recombinant protein MIC-10AIgG, and other is with embodiment 9.
Claims (7)
1. recombinant protein MIC-10 is characterized in that: it is MIC-10A, MIC-10B or MIC-10C, its nucleotide sequence successively respectively as: shown in the SEQ ID No.1, shown in the SEQ ID No.2 or shown in the SEQ ID No.3.
2. the preparation method of recombinant protein MIC-10, it comprises:
1) synthetic primer;
The primer of preparation recombinant protein MIC-10A gene is:
FW：GATCGGATCCGATTTTTTTAACGATTAC
RV：GATCAAGCTTTCATTCACGAAGTCCTCTTTTC
The primer of preparation recombinant protein MIC-10B gene is:
FW：GATCGGATCCGCCGCTGAGCGTGAGGAGAAG
RV：GATCAAGCTTTCACATTGATTTCCTGCG
The primer of preparation recombinant protein MIC-10C gene is:
FW：GATCGGATCCGATTTTTTTAACGATTAC
RV：GATCAAGCTTTCACATTGATTTCCTGCG
2) extracting RH strain toxoplasma cdna group DNA is template, carries out pcr amplification, obtains gene fragment, is cloned on the PMD-18T carrier;
3) the MIC10A gene fragment is inserted structure recombinant expression plasmid PET28A-MIC-10A, PET28A-MIC-10B or PET28A-MIC-10C among the expression vector PET28A;
4) recombinant expression plasmid PET28A-MIC-10A changed among the BL21 (DE3) express, recombination fusion protein MIC-10A, MIC-10B or MIC-10C, use the ni-sepharose purification recombinant protein.
3. anti-recombinant protein MIC-10 polyclonal antibody is prepared by following method:
With recombinant protein MIC-10 immunity 3 month female rabbit, subcutaneous 3 injections, immunizing dose 500ug//time, 21d immunity at interval, immunity is 3 times altogether, the 1st employing Freund's complete adjuvant, 2nd, adopt Freund's incomplete adjuvant 3 times, back 21 days of the 3rd immunity, the heart bloodletting is placed after 1 hour for 37 ℃, 4 ℃ are spent the night, leave the heart 15 minutes next day 3000, get supernatant, with the anti-MIC10 of protein A chromatography column purified rabbit, by specification carries out, IgG behind the purifying measures protein concentration with PBS dialysis 3 times with the BCA method, and-80 ℃ frozen;
Described recombinant protein MIC-10 is MIC-10A or MIC-10B.
4. arch insect circulating antigen double antibodies sandwich ELISA detection kit, it comprises: coated antibody, biotin labeled detection antibody, positive control, washings, confining liquid, enzyme mark avidin, substrate, stop buffer and sample diluent is characterized in that: described coated antibody is that the anti-recombinant protein MIC-10AIgG of rabbit, detection antibody are that anti-MIC-10BIgG of rabbit or coated antibody are that the anti-recombinant protein MIC-10BIgG of rabbit, detection antibody are anti-recombinant protein MIC-10AIgG; Positive control is recombinant protein MIC-10C.
5. a kind of arch insect circulating antigen double antibodies sandwich ELISA detection kit according to claim 4, it is characterized in that: confining liquid is the TBS that contains 1%BSA and 10% horse serum, and substrate is TMB, and stop buffer is a 2M sulfuric acid.
6. arch insect circulating antigen double antibodies sandwich ELISA detection kit using method, it comprises:
1) the anti-MIC-10AIgG of 5ug/ml rabbit that adds 100 μ l purifying wraps by the ELISA Sptting plate 4 ℃ of overnight incubation;
2) wash 5 times each 5 minutes with TBST;
3) every hole adds the sealing of 300 μ l confining liquids, hatches 30 minutes for 37 ℃;
4) wash 5 times each 5 minutes with TBST;
5) every hole adds 100 μ l serum sample to be checked, hatches 1 hour for 37 ℃;
6) wash 5 times each 5 minutes with TBST;
7) every hole adds the anti-MIC-10BIgG of the biotin labeled rabbit of 100 μ l, hatches 1 hour for 37 ℃;
8) wash 5 times each 5 minutes with TBST;
9) add enzyme mark avidin, hatched 30 minutes for 37 ℃;
10) wash 5 times with TBST, each 5 minutes, hatched 10 minutes;
11) every hole adds the 100ul substrate, room temperature lucifuge colour developing 30 minutes; Every hole adds 50ul 2M sulfuric acid termination reaction;
12) read plate, record OD value; According to typical curve, calculate the MIC10 content in the serum sample to be checked.
7. a kind of arch insect circulating antigen double antibodies sandwich ELISA detection kit using method according to claim 6 is characterized in that: described coated antibody is for exempting from anti-recombinant protein MIC-10BIgG, detecting antibody for exempting from anti-recombinant protein MIC-10AIgG.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104965086A (en) * | 2015-05-21 | 2015-10-07 | 华南农业大学 | Dog toxoplasma gondii antibody indirect ELISA detection kit |
| CN106053817A (en) * | 2016-05-03 | 2016-10-26 | 吉林大学 | A double-antibody sandwich kit for detecting a toxoplasma gondii circulating antigen and a preparing method thereof |
| CN106556693A (en) * | 2016-11-30 | 2017-04-05 | 吉林省畜牧兽医科学研究院 | A kind of toxoplasma series connection multi-epitope gene ELISA detection kit |
-
2010
- 2010-08-27 CN CN 201010264255 patent/CN102010468B/en not_active Expired - Fee Related
Non-Patent Citations (3)
| Title |
|---|
| 《Experimental Parasitology》 20071001 George Dautu Toxoplasma gondii: Detection of MIC10 antigen in sera of experimentally infected mice 362-371 1-5 第118卷, * |
| 《Journal of Experimental Medicine》 20021231 Barragan, A Transepithelial migration of Toxoplasma gondii is linked to parasite motility and virulence 1625-1633 1-7 第195卷, * |
| 《Veterinary Research》 19981231 Buxton, D Protozoan infections (Toxoplasma gondii, Neospora caninum and Sarcocystis spp) in sheep and goats: recent advances 289-310 1-7 第29卷, * |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104965086A (en) * | 2015-05-21 | 2015-10-07 | 华南农业大学 | Dog toxoplasma gondii antibody indirect ELISA detection kit |
| CN106053817A (en) * | 2016-05-03 | 2016-10-26 | 吉林大学 | A double-antibody sandwich kit for detecting a toxoplasma gondii circulating antigen and a preparing method thereof |
| CN106556693A (en) * | 2016-11-30 | 2017-04-05 | 吉林省畜牧兽医科学研究院 | A kind of toxoplasma series connection multi-epitope gene ELISA detection kit |
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