CN106405074A - ELISA kit for detection of human cytomegalovirus IgG and IgM antibodies based on PP65 protein - Google Patents
ELISA kit for detection of human cytomegalovirus IgG and IgM antibodies based on PP65 protein Download PDFInfo
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Abstract
The present invention relates to a kit for detection of human cytomegalovirus (HCMV) IgG and IgM antibodies based on PP65 protein, the kit includes an elisa plate coated with a recombinant HCMV PP65322-561 protein antigen and enzyme-labeled second antibodies, and the enzyme-labeled second antibodies include a HRP-anti human IgG antibody for detection of IgG and a HRP-anti human IgM antibody for detection of IgM. Reagents have good specificity, sensitivity, stability and repeatability, the preparation process is simple, and the standard is easy to control.
Description
Technical field
The present invention relates to a kind of detection human cytomegalic inclusion disease virus（HCMV）IgG and the ELISA detection kit of IgM antibody.
Background technology
Human cytomegalic inclusion disease virus（Human cytomegalovirus, HCMV）Oral cavity, reproductive tract, Placenta Hominiss, lactication can be passed through
And the multipath propagation such as blood transfusion or organ transplantation.The distinguishing feature of population infection HCMV is that infection rate is high, adult normal HCMV
Antibody positive rate is 76.7%-95.8%, and China is also one of HCMV infection high prevalence country.
HCMV primary infection can non-evident sympton, but virus is hidden for a long time from this in vivo.When body's immunity is low,
Latent viruss start to replicate and breed in a large number, and secondary Active infection simultaneously shows various clinical symptom.HCMV infection is to lead to
One of key factor of raw defect, and have research display HCMV infection and diabetes, atherosclerosiss and some are pernicious swollen
Morbidity closely related.Clinical Activity HCMV infection is more common in anemia of pregnant woman, neonate, blood transfusion patients, malignant tumor and acquired immune deficiency syndrome (AIDS)
Patient and organ transplantation etc. use the patient of immunosuppressant.Anemia of pregnant woman's Active infection can be passed by Placenta Hominiss, birth canal or lactication
To fetus, lead to miscarry, stillborn fetuses, fetus form deformity, organ dysfunction.Can be with Huang during the neonate birth of congenital infection
Subcutaneous ulcer, ecchymosis, chorioretinitises, microcephalus etc., and asymptomatic part neonate, are just gradually found to listen with developing
Feel, visual disturbance, mental retardation, dyskinesia etc..Weaken in immunologic functions such as malignant tumor, acquired immune deficiency syndrome (AIDS), organ transplantations
Or the Active infection Chang Bingfa interstitial pneumonia of downtrod patient HCMV, retinitiss, esophagitis, colitis and meninges brain
The various clinical syndromes of the multiple organ organ injury such as inflammation, are that patient is lethal or the major reason of transplantation failure.
, viral massive duplication, morning of symptom in the either secondary Active infection of primary infection or latent viruss
Phase body can produce IgM type antibody.The feature with respect to IgG for the IgM is that generation is early, it is fast to disappear.So the detection of IgM represents in the recent period
Infection, body may still be in viral massive duplication state.In addition the detection of IgM is how quickly cheap using simplicity
ELISA method, so while existing, the time period being detected is shorter, is still the inspection whether clinical conventional preliminary judgement is infected
Survey method.And detect the reagent of HCMV IgM how using HCMV totiviruss crack protein as envelope antigen at present.Its antigenic component
The possibility of complicated and potential host cell proteins pollution, also potential other antiviral antibodies of combination herpetoviridae generation false positive letter
Number possibility, and the manufacturing process of reagent because need culture virus and complex operation.Minority HCMV IgM antibody detectable
Though with the recombiant protein of HCMV as envelope antigen, reagent makes economical and convenient, the less stable of reagent, sensitivity and spy
The opposite sex also has much room for improvement.So developing, manufacturing process is simple, low cost, Sensitivity and Specificity more preferable HCMV IgM antibody
Detectable still has practical significance.
Detection IgG antibody understands crowd infection rate, the system to the prevention of HCMV relevant disease, management control and relevant policies
Fixed significant.And clearly individuality HCMV Infection Status are then beneficial to clinical determination individual treatment scheme, such as AIDS and device
Official transplantation patient, needs to detect HCMV IgG antibody, determines whether to need to follow the tracks of its IgM antibody level, so that in time
Anti- HCMV treatment.The so although detection of HCMV IgG antibody can only represent once infection, hiding in vivo has HCMV, and non-viral work
What dynamic sexuality contaminated accuses of, but still has the market demand.The reagent of presently commercially available detection HCMV IgG antibody is how complete with HCMV
Virolysis thing as envelope antigen, its antigenic component complicated and potential host cell proteins pollution possibility, also potential combination
Other antiviral antibodies of herpetoviridae produce the possibility of false positive signal, and the manufacturing process of reagent is because of needs culture virus
Complex operation.Then with the recombiant protein of HCMV as envelope antigen, this reagent makes warp to minority HCMV IgG antibody detectable
The less stable of the easy still reagent of Ji, Sensitivity and Specificity also has much room for improvement, and such reagent selling at exorbitant prices of import.
In sum, need that a kind of manufacturing process is simple, low cost, Sensitivity and Specificity more preferable HCMV IgG and
HCMV IgM antibody detectable.
Content of the invention
The technical problem to be solved in the present invention is to provide a kind of reagent based on PP65 Protein Detection HCMV IgG and IgM
Box, at least has the characteristics that specificity, sensitivity, stability are higher.
According to an aspect of the invention, it is provided a kind of test kit based on PP65 Protein Detection HCMV IgG and IgM,
Including
It is coated with restructuring HCMV PP65322-561The ELISA Plate of proteantigen；
ELIAS secondary antibody, including for detecting the HRP- anti-human IgG antibodies of IgG and being used for detecting that the anti-human IgM of HRP- of IgM resists
Body.
As preferred technical scheme, described is coated with restructuring HCMV PP65322-561The system of the ELISA Plate of proteantigen
Preparation Method includes step：Prokaryotic expression purification HCMV is by phospholamban PP65322-561；It is coated liquid dilution restructuring PP65322-561Egg
Bai Kangyuan is 10 g/ml, and every hole 100 μ l adds in ELISA Plate, 37 DEG C 1 hour, then 4 DEG C be coated 12 ~ 24 hours；Discard in the hole
Liquid, PBST pats dry after washing, and using the every hole of the PBS confining liquid 100 μ l containing 1%BSA, 37 DEG C of closing 2h, PBST pat dry after washing.
Mentioned reagent box can be used for detecting HCMV IgG antibody and HCMV IgM antibody.
As preferred technical scheme, described HRP- anti-human IgG antibodies are selected from HRP- goat anti-human igg antibody, HRP- rabbit
One of anti-human IgG antibodies, HRP- Mus anti-human γ chain antibody.Described HRP- anti-human IgM antibodies are selected from HRP- goat-anti people IgM
One of antibody, HRP- rabbit anti-human IgM antibodies, the anti-human chain antibody of HRP- Mus.
Recombiant protein containing the strongly immunogenic epitope of HCMV can replace totiviruss to be used for the inspection of HCMV specific antibody
Survey.HCMV PP65 albumen be in the numerous structural protein of HCMV can with specificity excite B cell, cytotoxic T cell (
CTL) and complementary T cell (Th) immunity major target proteins, in different HCMV Strain, HCMV pp65 gene height
Conservative, PP65 used by the present invention332～561Fragment is enriched in PP65 the 322 to 561st bit amino of t cell epitope and B cell epi-position
Acid sequence.PP65 albumen be HCMV by phospholamban rather than glycosylation albumen, the glycosylation without eukaryotic expression and folding process,
It is achieved with high immunological activity protein fragments in procaryotic cell expression.
According to a further aspect in the invention, there is provided a kind of method of detection human cytomegalic inclusion disease virus IgG antibody, employ
State test kit.
As preferred technical scheme, the method for described detection human cytomegalic inclusion disease virus IgG antibody includes step：Dilute treats
Inspection serum, is loaded 37 DEG C of incubations in ELISA Plate in the hole, washes plate；Plus 37 DEG C of incubations of ELIAS secondary antibody HRP- anti-human IgG antibodies, wash plate；
Plus substrate nitrite ion, color development at room temperature；Plus terminate liquid, microplate reader detection OD450Value.
According to a further aspect in the invention, there is provided a kind of method of detection human cytomegalic inclusion disease virus IgM antibody, employ
State test kit.
As preferred technical scheme, the method for described detection human cytomegalic inclusion disease virus IgM antibody includes step：By IgG-
RF adsorbent is mixed with test serum, and room temperature is placed；Dilute the test serum crossed through sorbent treatment, add in ELISA Plate in the hole
Sample, after 37 DEG C of incubations, PBST washes plate, pats dry；Plus 37 DEG C of incubations of ELIAS secondary antibody HRP- anti-human IgM antibodies, wash plate；Plus substrate colour developing
Liquid, color development at room temperature；Plus terminate liquid, microplate reader detection OD450Value.The specific IgM being directed to envelope antigen in test serum sample is normal
Often exist with specific IgG, the latter can disturb the mensure of IgM simultaneously.Meanwhile, rheumatoid factor（RF）Can also interfere with detection knot
Really.Therefore before blood serum sample mensure, IgG-RF adsorbent must be added to eliminate IgG, RF interference.The present invention experiment verify by
IgG-RF adsorbent and test serum 1:1 mixing, room temperature is placed 30 min and be can remove interference.
The present invention is using restructuring PP65332～561Albumen establish detection HCMV IgG antibody and HCMV IgM antibody indirect
ELISA detection method is it was demonstrated that reagent has preferable specificity, sensitivity, stability and repeatability, and reagent manufacturing process
Simply, standard is easily controllable.Above-mentioned detection method only obtains intermediate result, and this result is no straight with the diagnosis of disease or health status
Connect necessary connection.For IgG detection method, the positive can only explain and once infected, and is mainly used in Epidemiological study, is beneficial to
Disease control department's policies reference.For IgM detection method, if the positive is in addition it is also necessary to combine other necessary indexs ability really
Determine whether to be in Infection Status, detection colony IgM positive rate is also commonly used for the Epidemiological study of crowd's recent infection present situation.
Using widely used, industry think Sensitivity and Specificity all preferably company of Italian DESAY detectable as
Relatively reference reagent, testing result shows, detection HCMV IgG antibody and its coincidence rate are 95%, detection HCMV IgM antibody with
Its coincidence rate is 81%, and the positive rate of the present invention is higher than this reagent, and sensitivity is higher.For clinical quick detection HCMV
IgG and HCMV IgM antibody, specifies previous infection and latent infection provides a kind of new selection, also for crowd's HCMV infection state
Epidemiological study and routine monitoring provide new alternative.
Specific embodiment
It should be noted that in the case of not conflicting, the embodiment in the application and the feature in embodiment can phases
Mutually combine.To describe the present invention below with reference to specific embodiment in detail.
Embodiment 1
Based on PP65322-561The reagent of Protein Detection IgG, including be coated with restructuring HCMV PP65322-561The enzyme of proteantigen
Target, HRP- anti-human igg, PBST cleaning mixture, substrate nitrite ion and terminate liquid.
Wherein, HCMV PP65322-561The preparation method of the ELISA Plate of proteantigen is prokaryotic expression purification HCMV envelope
Phosphoprotein PP65322-561；It is coated liquid dilution restructuring PP65322-561Proteantigen is 10 g/ml, and every hole 100 μ l adds ELISA Plate
In, 37 DEG C 1 hour, then 4 DEG C be coated 12 ~ 24 hours；Discard in the hole liquid, PBST washes 3 times, pats dry, using the PBS containing 1%BSA
The every hole of confining liquid 100 μ l, 37 DEG C of closing 2h, PBST wash 3 times, pat dry.
The formula of PBST cleaning mixture is：0.15M KH2PO40.2g, Na2HPO4·12H2O 2.9g, NaCl 8.0g, KCl
0.2g, 0.05% Tween-20 0.5ml, plus distilled water is to 40ml；
Substrate nitrite ion is to be mixed by substrate solution A and each half of substrate buffer solution B before use.Substrate solution A Main Ingredients and Appearance
It is tetramethyl benzidine（TMB)：TMB 20mg, dehydrated alcohol 10ml, plus distilled water is to 100ml；Substrate buffer solution B Main Ingredients and Appearance
It is the phosphate citrate acid of PH5.0：0.2M Na2HPO425.7ml, 0.1M citric acid 24.3ml, plus distilled water is to 100ml；
Terminate liquid Main Ingredients and Appearance is 2M H2SO4：Distilled water 178.3ml, is added dropwise over 98% concentrated sulphuric acid 21.7ml；
Shrouding film is overlay of the same size with the plate face of ELISA Plate.
Above-described HRP- anti-human IgG antibodies are selected from HRP- goat anti-human igg antibody, HRP- rabbit anti-human igg's antibody, HRP-
One of Mus anti-human γ chain antibody, but not limited to this.
Embodiment 2
Based on PP65322-561The test kit of Protein Detection IgM, including be coated with restructuring HCMV PP65322-561Proteantigen
ELISA Plate, HRP- anti-human IgM, PBST cleaning mixture, substrate nitrite ion and terminate liquid.
Wherein, HCMV PP65322-561The preparation method of the ELISA Plate of proteantigen is prokaryotic expression purification HCMV envelope
Phosphoprotein PP65322-561；It is coated liquid dilution restructuring PP65322-561Proteantigen is 10 g/ml, and every hole 100 μ l adds ELISA Plate
In, 37 DEG C 1 hour, then 4 DEG C be coated 12 ~ 24 hours；Discard in the hole liquid, PBST washes 3 times, pats dry, using the PBS containing 1%BSA
The every hole of confining liquid 100 μ l, 37 DEG C of closing 2h, PBST wash 3 times, pat dry.
The formula of PBST cleaning mixture is：0.15M KH2PO40.2g, Na2HPO4·12H2O 2.9g, NaCl 8.0g, KCl
0.2g, 0.05% Tween-20 0.5ml, plus distilled water is to 40ml；
Substrate nitrite ion is to be mixed by substrate solution A and each half of substrate buffer solution B before use.Substrate solution A Main Ingredients and Appearance
It is tetramethyl benzidine（TMB)：TMB 20mg, dehydrated alcohol 10ml, plus distilled water is to 100ml；Substrate buffer solution B Main Ingredients and Appearance
It is the phosphate citrate acid of PH5.0：0.2M Na2HPO425.7ml, 0.1M citric acid 24.3ml, plus distilled water is to 100ml；
Terminate liquid Main Ingredients and Appearance is 2M H2SO4：Distilled water 178.3ml, is added dropwise over 98% concentrated sulphuric acid 21.7ml；
Shrouding film is overlay of the same size with the plate face of ELISA Plate.
Above-described HRP- anti-human IgM antibodies are selected from HRP- goat anti-human igg antibody, HRP- rabbit anti-human igg's antibody, HRP-
One of anti-human chain antibody of Mus, but not limited to this.
Embodiment 3
The detection method of human cytomegalic inclusion disease virus IgG antibody, the method adopts the reagent that embodiment 1 provides.With confining liquid 1:400 is dilute
The serum to be checked released, in ELISA Plate in the hole sample-adding, every hole 100 μ l, after 37 DEG C of incubation 1h, PBST washes plate 3 times, pats dry；Plus confining liquid
1:The HRP- goat anti-human iggs of 4000 dilutions, every hole 100 μ l, 37 DEG C of 1h, PBST wash plate 3 times；Plus the every hole of substrate nitrite ion 100 μ l,
Color development at room temperature 15min；Plus terminate liquid 50 μ l, microplate reader detection OD450Value.
The present embodiment and Italian De Sai company HCMV-IgG ELISA reagent are shown in the testing result of 100 parts of samples respectively
Following table.Detect 89 and 86 positives respectively, positive rate is respectively 89% and 86%.The coincidence rate of two kinds of reagent is 95%.
The present embodiment shows OD to the testing result of HSV- I type, HSV- II type and EBV IgG positive serum450Average is respectively
0.097th, 0.084,0.101, respectively less than marginal value 0.135, it was demonstrated that no cross reaction, eliminates self-control reagent non-specific binding bleb
The possibility of other antiviral antibodies of exanthema virus section.With PP65 McAb to PP65322-561The identification knot of protein fragments Western Blot
Also explanation self-control reagent has good specificity to fruit.
The present embodiment variation within batch coefficient between 2.1%～5.1%, interassay coefficient of variation between 3.4%～6.4%, respectively less than
10%, illustrate that detection system is stable, stability and repeatability are good.
Embodiment 4
The detection method of human cytomegalic inclusion disease virus IgM antibody, the method adopts the reagent that embodiment 2 provides.By IgG-RF adsorbent
With test serum 1:1 mixing, room temperature places 30 min；1:The test serum that 200 dilutions are crossed through sorbent treatment, in ELISA Plate hole
Interior sample-adding, every hole 100 μ l, after 37 DEG C of incubation 1h, PBST washes plate 3 times, pats dry；Plus confining liquid 1:The HRP- goat-anti people of 2000 dilutions
The every hole of IgM 100 μ l, 37 DEG C of incubation 1h, wash plate 3 times；Plus the every hole of substrate nitrite ion 100 μ l, color development at room temperature 15min；Plus terminate liquid
Every hole 50 μ l, microplate reader detects OD450Value.
Before blood serum sample measures, IgG-RF adsorbent must be added to eliminate IgG, RF interference.The present invention experiment verify by
IgG-RF adsorbent and test serum 1:1 mixing, room temperature is placed 30 min and be can remove interference.In addition, utilizing beta -mercaptoethanol solution
The function of poly- IgM multimeric structure, 37 DEG C of process testing sample 1 h of beta -mercaptoethanol of equivalent 0.2 mol/L find 5 parts
HCMV IgM antibody positive serum testing result all switchs to feminine gender, and 1 part of only positive serum of HCVM IgG antibody is unchanged,
Illustrate that the surveyed antibody of the present invention is IgM.
The present embodiment shows OD to the testing result of HSV- I type, EBV IgM positive serum450It is respectively less than marginal value 0.179,
Confirm the present invention and HSV-1 type, the equal no cross reaction of EBV IgM, specificity is good.
Precision Analyze result shows：Variation within batch coefficient is 5.3%～8.3%, the coefficient of variation of replica test between batch
For 2.3%～7.9%, respectively less than 10%, illustrate that detection system is stable, have preferable repeatability.
The present embodiment and Italian De Sai company HCMV-IgM ELISA reagent doubtful HCMV infection to 100 parts of clinics respectively
The testing result of serum see table.Detect 58 and 51 positives respectively, positive rate is respectively 58% and 51%, and the present embodiment is sensitive
Property be higher than company of DESAY reagent.The coincidence rate of two kinds of reagent is 81%.
Embodiment 5
The detection method of human cytomegalic inclusion disease virus IgG antibody, the method adopts the reagent that embodiment 1 provides.With confining liquid 1:400 is dilute
The serum to be checked released, in ELISA Plate in the hole sample-adding, every hole 100 μ l, after 37 DEG C of incubation 1h, PBST washes plate 3 times, pats dry；Plus confining liquid
1:The HRP- rabbit anti-human iggs of 4000 dilutions, every hole 100 μ l, 37 DEG C of 1h, PBST wash plate 3 times；Plus the every hole of substrate nitrite ion 100 μ l,
Color development at room temperature 15min；Plus terminate liquid 50 μ l, microplate reader detection OD450Value.
Embodiment 6
The detection method of human cytomegalic inclusion disease virus IgM antibody, the method adopts the reagent that embodiment 2 provides.By IgG-RF adsorbent
With test serum 1:1 mixing, room temperature places 30 min；1:The test serum that 200 dilutions are crossed through sorbent treatment, in ELISA Plate hole
Interior sample-adding, every hole 100 μ l, after 37 DEG C of incubation 1h, PBST washes plate 3 times, pats dry；Plus confining liquid 1:The HRP- rabbit of 2000 dilutions is anti-human
The every hole of IgM 100 μ l, 37 DEG C of incubation 1h, wash plate 3 times；Plus the every hole of substrate nitrite ion 100 μ l, color development at room temperature 15min；Plus terminate liquid
Every hole 50 μ l, microplate reader detects OD450Value.
Ibid detection method, plus HRP- Mus anti-human γ chain antibody detection human cytomegalic inclusion disease virus IgG antibody, plus HRP- Mus are anti-human
Chain antibody detects human cytomegalic inclusion disease virus IgM antibody.The scope of protection of present invention is not limited to above specific embodiment, right
For those skilled in the art, the present invention can have institute within various deformation and change, all designs in the present invention and principle
Any modification, improvement and the equivalent made all should be included within protection scope of the present invention.
Claims (8)
1. a kind of ELISA kit based on PP65 Protein Detection human cytomegalic inclusion disease virus IgG and IgM antibody is it is characterised in that wrap
Include：
It is coated with restructuring HCMV PP65322-561The ELISA Plate of proteantigen；
ELIAS secondary antibody, including for detecting the HRP- anti-human IgG antibodies of IgG and being used for detecting that the anti-human IgM of HRP- of IgM resists
Body.
2. test kit according to claim 1 is it is characterised in that described is coated with restructuring HCMV PP65322-561Albumen resists
The preparation method of former ELISA Plate includes step：Prokaryotic expression purification HCMV is by phospholamban PP65322-561；It is coated liquid dilution
Restructuring PP65322-561Proteantigen is 10 g/ml, and every hole 100 μ l adds in ELISA Plate, 37 DEG C 1 hour, then 4 DEG C be coated 12 ~
24 hours；Discard in the hole liquid, PBST pats dry after washing, using the every hole of the PBS confining liquid 100 μ l containing 1%BSA, 37 DEG C of closing 2h,
PBST pats dry after washing.
3. test kit according to claim 1 and 2 it is characterised in that：Described HRP- anti-human IgG antibodies are selected from HRP- sheep
One of anti-human IgG antibodies, HRP- rabbit anti-human igg's antibody, HRP- Mus anti-human γ chain antibody.
4. test kit according to claim 1 and 2 it is characterised in that：Described HRP- anti-human IgM antibodies are selected from HRP- sheep
Anti-human IgG antibodies, HRP- rabbit anti-human igg's antibody, one kind of HRP- Mus anti-human chain antibody kind.
5. a kind of detection human cytomegalic inclusion disease virus IgG antibody method it is characterised in that：Using the test kit described in claim 3.
6. method according to claim 5 is it is characterised in that include step：The serum to be checked of dilution, in ELISA Plate in the hole
37 DEG C of incubations of sample-adding, wash plate；Plus 37 DEG C of incubations of ELIAS secondary antibody HRP- anti-human IgG antibodies, wash plate；Plus substrate nitrite ion, room temperature show
Color；Plus terminate liquid, microplate reader detection OD450Value.
7. a kind of detection human cytomegalic inclusion disease virus IgM antibody method it is characterised in that：Using the test kit described in claim 4.
8. method according to claim 7 is it is characterised in that include step：IgG-RF adsorbent and test serum are mixed
Even, room temperature is placed；Dilute the test serum crossed through sorbent treatment, PBST washes plate after ELISA Plate in the hole sample-adding, 37 DEG C of incubations,
Pat dry；Plus 37 DEG C of incubations of ELIAS secondary antibody HRP- anti-human IgM antibodies, wash plate；Plus substrate nitrite ion, color development at room temperature；Plus terminate liquid,
Microplate reader detects OD450Value.
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| CN201610722838.0A CN106405074A (en) | 2016-08-25 | 2016-08-25 | ELISA kit for detection of human cytomegalovirus IgG and IgM antibodies based on PP65 protein |
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| CN201610722838.0A CN106405074A (en) | 2016-08-25 | 2016-08-25 | ELISA kit for detection of human cytomegalovirus IgG and IgM antibodies based on PP65 protein |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111337689A (en) * | 2020-04-03 | 2020-06-26 | 山西医科大学 | Novel coronavirus detection kit |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1651918A (en) * | 2005-01-08 | 2005-08-10 | 王明丽 | Method for making recombinant antigen pp 65 envelope enzyme-linked reaction plate and ELISA test kit |
-
2016
- 2016-08-25 CN CN201610722838.0A patent/CN106405074A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1651918A (en) * | 2005-01-08 | 2005-08-10 | 王明丽 | Method for making recombinant antigen pp 65 envelope enzyme-linked reaction plate and ELISA test kit |
Non-Patent Citations (4)
| Title |
|---|
| 李娜等: "人巨细胞病毒PP65322-561片段的原核表达及功能鉴定", 《国际病毒学杂志》 * |
| 王厚照等: "《临床检验手册》", 30 November 2008 * |
| 钱春艳等: "人巨细胞病毒pp65抗体检测方法的建立", 《检验医学》 * |
| 闻玉梅等: "《现代微生物学》", 31 January 1991 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111337689A (en) * | 2020-04-03 | 2020-06-26 | 山西医科大学 | Novel coronavirus detection kit |
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