CN108663523A - It can be used for diagnosing the kit of human enterovirus 71 infection - Google Patents
It can be used for diagnosing the kit of human enterovirus 71 infection Download PDFInfo
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- G—PHYSICS
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- 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
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- 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
- G01N33/569—Immunoassay; Biospecific binding assay; Materials therefor for microorganisms, e.g. protozoa, bacteria, viruses
- G01N33/56983—Viruses
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- G01N2333/085—Picornaviridae, e.g. coxsackie virus, echovirus, enterovirus
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- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
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- G01N2333/085—Picornaviridae, e.g. coxsackie virus, echovirus, enterovirus
- G01N2333/09—Foot-and-mouth disease virus
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Abstract
The present invention provides the kit that can be used for diagnosing human enterovirus 71 infection.The detection kit of the present invention includes one or more solid carriers and the SEQ ID NO being independently connected on the solid carrier：Polypeptide shown in 1 ~ 6.When with whether SEQ ID NO：Whether polypeptide shown in 1 ~ 6 all has response as judgement subject in the case of index infect by human enterovirus 71 the IgM in the blood sample in subject source, can diagnose human enterovirus 71 with 92% or more sensitivity and 5% false positive rate below and infect.
Description
Technical field
The invention mainly relates to detection kits.Specifically, the present invention relates to can be used for detecting human enterovirus 71
The kit of infection.
Background technology
The infection of enterovirus EV 71 type is the disease caused by human enterovirus 71 (EV71), is clinically mainly shown as
Hand-foot-and-mouth disease, hand-foot-and-mouth disease eruption and prevalence mostly in infant, part EV71 infection infants show as herpangina, severe
There is viral encephalitis, viral cerebrospinal meningitis, pulmonary edema, empsyxis etc. in patient.Cause the cause of disease of hand-foot-and-mouth disease, currently,
Most commonly Coxsackie virus A 6 and enterovirns type 71;Severe infection occurs for disease caused by being infected by enterovirns type 71
Large percentage, case fatality rate is also higher, and severe cases case fatality rate is up to 10% -25%, therefore etiological diagnosis is significant.
Three aspects of clinical diagnosis Main Basiss of enterovirus EV 71:When clinical manifestation, second is that serological evidence, three
It is Etiological Evidence.Due to enterovirus EV 71 type infection clinical manifestation it is more, the Serologic detection of enterovirus EV 71 and
Pathogeny detection is current most important foundation.EV71 infection generates immunoprotection, and IgM and IgG antibody can be detected after infection.
Invention content
The purpose of the present invention is to provide the new kit that can be used for diagnosing human enterovirus 71 infection and diagnosis sides
Method.
The present inventor it has surprisingly been found that when with whether SEQ ID NO：Polypeptide shown in 1 ~ 6 all carrys out subject
IgM in the blood sample in source has the case where index whether response has been infected by human enterovirus 71 as judgement subject
Under, it can be with 92% or more sensitivity and 5% false positive rate diagnosis human enterovirus 71 infection below.
Specifically, the present invention provides：
1. a kind of detection kit comprising one or more solid carriers, and be independently connected on the solid carrier
SEQ ID NO：Polypeptide shown in 1 ~ 6.
2. according to the detection kit described in item 1, wherein the SEQ ID NO：Polypeptide shown in 1 ~ 6 is independently connected
In on same solid carrier.
3. according to the detection kit described in item 1, wherein the solid carrier is SJ modified silica-gels.
4. according to the detection kit described in item 1, it is used to diagnose human enterovirus 71 infection.
5. a kind of diagnosis subject whether be human enterovirus 71 infection method, this method includes：
The SEQ ID NO are detected using the detection kit described in any one of item 1 ~ 4：Polypeptide is to subject shown in 1 ~ 6
Whether the IgM in the blood sample in source has response；Wherein,
As the SEQ ID NO：When polypeptide shown in 1 ~ 6 all has response to the IgM in the blood sample in subject source, sentence
It calmly should the infection of subject is a human enterovirns type 71；Otherwise, it is determined that the subject infects for non-human enterovirus 71.
6. according to the method described in item 5, wherein the blood sample is whole blood, blood plasma or serum.
7. according to the method described in item 5, wherein the subject is the subject with hand-foot-and-mouth disease clinical symptoms.
Polypeptide of the present invention is：
SEQ ID NO：Polypeptide sequence shown in 1 is GKGELCAVFRADPGRNGPWQ；
SEQ ID NO：Polypeptide sequence shown in 2 is EDTHPPYKQTQPGADGFELQ；
SEQ ID NO：Polypeptide sequence shown in 3 is SLDFALSLLRRNIRQVQTDQ；
SEQ ID NO：Polypeptide sequence shown in 4 is NKEPAVLHSRDPRLEVDFEQ；
SEQ ID NO：Polypeptide sequence shown in 5 is GHFTMLGVRDPLAVLPRHSQ；
SEQ ID NO：Polypeptide sequence shown in 6 is LIREYSNRSAIGNTIEALFQ.
The polypeptide suitable can be obtained using the known methods such as (1) chemical synthesis process or (2) enzyme reaction synthetic method
, wherein chemical synthesis is more easy.In the polypeptide of the chemical synthesis present invention, by using peptide synthesizer synthesis or
The semi-synthetic polypeptide carries out.As chemical synthesis process, can enumerate such as peptide solid-phase synthesis.The peptide synthesized in this way
Conventional means may be used to be purified such as ion-exchange chromatography, reverse phase high performance liquid chromatography, affinity chromatography.Such peptide
Solid phase synthesis process with and subsequent peptide purification be all well-known in the art.
In addition, by the polypeptide of the enzyme reaction production present invention, such as International Publication pamphlet may be used
Method described in No. WO2004/011653.I.e., it is possible to produce in this way：By the amino acid of a side or the carboxyl terminal quilt of dipeptides
Amino acid or dipeptides obtained from esterification or amidation, amino acid (such as the carboxy protective for being in amino acid free state
Amino acid) it is reacted in the presence of peptide synthetase, the dipeptides or tripeptides of generation.As peptide synthetase, can enumerate：Tool
There are the culture of the microorganism for the ability for generating peptide, the thalline of the microbial cells or the microorganism that are detached by the culture
Manage object or the microbe-derived peptide synthetase.
The chemical synthesis of especially polypeptide has been realized in commercialization, can by entrust profession Peptide systhesis company come
Synthesize the polypeptide.
Kit of the present invention is：
A kind of detection kit comprising one or more solid carriers, and be independently connected on the solid carrier
SEQ ID NO：Polypeptide shown in 1 ~ 6.
It is verified by experiments, kit of the invention can be diagnosed with 92% or more sensitivity and 5% false positive rate below
Human enterovirus 71 infects.In this specification, sensitivity refers to：In positive sample with the confirmation of " goldstandard " method, by it
His method is measured as the ratio of positive sample.False positive rate refers to：In negative sample with the confirmation of " goldstandard " method, by other
Method is measured as the ratio of positive sample." goldstandard " method that the art detects human enterovirus 71 infection is nucleic acid
Detection method, i.e. PCR methods.In this specification, " diagnosis " can be made a definite diagnosis, and can also be to provide reference information to make a definite diagnosis.
In the present invention, solid carrier is not particularly limited, as long as (being, for example, can as solid or insoluble material
Material to be detached from reaction mixture by filtering, precipitation, Magnetic Isolation etc.) carrier.
Constitute solid carrier material include but not limited to：Silica gel（Dimethyl silicone polymer, PDMS）, cellulose, Teflon
It is grandTM, NC Nitroncellulose, agarose, glucan, chitosan, polystyrene, polyacrylamide, polyester, makrolon, polyamide,
Polypropylene, nylon, polyvinylidene fluoride, latex, silica, glass, glass fibre, gold, platinum, silver, copper, iron, stainless steel, iron
Oxysome, silicon wafer, polyethylene, polyethyleneimine, polylactic acid, resin, polysaccharide, albumen (albumin etc.), carbon or their group
Close etc..
The shape of solid carrier includes but not be limited to：Pearl, magnetic bead, film, microcapillary, filter membrane, plate, micro plate, carbon
Nanotube, sensor chip etc..Just as known in the art, it can be set on the flat solid carrier such as film or plate
Set pit, groove, filter membrane bottom etc..
Magnetic bead can be with the sphere diameter of about 25nm ~ about 1mm ranges.In a preferred embodiment, magnetic bead has about
The diameter of the μ ms of 50nm ~ about 10.The size of magnetic bead can be selected according to specific purposes.It is contour by Sepharose
Pearl made of crosslinked spherical agarose has the diameter of about 24 μm ~ about 165 μ ms.Preferably, high crosslinked spherical agarose
Pearl has the diameter of about 24 μm ~ about 44 μ ms.The size of high crosslinked spherical sepharose 4B can according to specific purposes come into
Row selection.
The example of solid carrier with hydrophobic surface include can from Polysciences, Warrington, PA or
The polystyrene latex beads such as the product of Spherotech, Liberville, IL purchase.
Silica (SiO2)-processing or silica (SiO2) base solid carrier example include can be from
The extraordinary magnetic silica pearl etc. of Polysciences, Warrington, PA purchase, can be used for capturing nucleic acid (such as
DNA).It can be from Dynal Biotech M-280 bought etc. alternatively, can also use.
Magnetic bead with hydrophilic surface can be used for capturing bacterial cell, nucleic acid and the other ingredients of proliferation period.As
The example of the magnetic bead can enumerate Polysciences, the pearl (title of Warrington, PA sale：Biomag (registrations
Trade mark) carboxyl) or Bangs Laboratory, Inc., Fishers, IN entitled MC02N/2928 pearl.Or
Person, the M-270 etc. that Dynal Biotech can be used to sell.
In a preferred embodiment of the present invention, the solid carrier is SJ modified silica-gels, is Suzhou Yvonne consor
A kind of microarray solid support material (the iPDMS films, referring to Chinese patent of silicone rubber material of object Materials Co., Ltd exploitation
CN101265329A).This material be based on the common PDMS of biological study, be added wherein specific initiator at
Point（Make the material that can pass through surface initiated polymerization（SIP）Realize surface-functionalized modification）, using polyethylene glycol methyl
What acrylate (poly (oligo (ethylene glycol) methacrylate), pOEGMA) surface modification obtained.SJ
Modified silica-gel has outstanding anti-protein non-specific adsorption (Nonspecific protein adsorption, NPA) energy
Power, the non-specific protein absorption control in can detecting complicated protein immunization is to close to " absolute 0 " is horizontal（It is close or low
In the detectable limit of instrument）, can not only exempt the trouble closed and be cleaned multiple times, can also be expanded by using stronger signal
Increasing means improve the sensitivity of protein microarray.And the essence of its silicon rubber imparts the stronger mechanical performance of the material
With good operability.SJ modified silica-gels are successfully applied to what 11 tumor markers formed by the poly- companies of Suzhou Yvonne
Combination assay microarray ELISA kit realizes high-throughput and highly sensitive detection, it was demonstrated that this material is a kind of excellent
Elegant protein microarray solid support material.Meanwhile this material also has the adjustable characteristic of surface nature, can pass through control
The modification reaction time processed adjusts its surface topography in a certain range.
In the present invention, the connection of polypeptide and solid carrier may be used that well known to a person skilled in the art polypeptides and solid to carry
The connection method of body carries out.For example, for the connection of protein/polypeptide and modified silica-gel surface, 1- second can be passed through
Base -3- (3- dimethyl aminopropyls)-carbodiimides [1-ethyl-3- (3-dimethyl ami-nopropyl)
Carbodiimide, EDC] and n-hydroxysuccinimide (N-hydroxysuccinimide
, NHS) reaction by the carboxyl on the macromolecular chain of modified silica-gel surface（-COOH）Group is changed to activated group, the activation base
Group can be with the amino of institute's band on protein/polypeptide（-NH2）Protein/polypeptide is fixed on solid carrier surface by reaction to realize.
The concentration of polypeptide in the sampling liquid that is used when point sample is not particularly limited, those skilled in the art can be according to normal
Rule selection, preferably 1 μ of μ g ~ 1000 g/mL, the more preferable 10 μ μ of g ~ 500 g/mL.In addition, being distributed on a solid carrier for polypeptide
Density be not particularly limited, those skilled in the art can be according to conventional selection, preferably 1 ~ 100 point/10mm2, more preferable 5 ~
50 points/10mm2。
In the present specification, " independent " refers to：The polypeptide forms a point on the solid carrier, is not wrapped in the point
Preferably only include the polypeptide containing on other polypeptides of the influential amount of detection.
Solid carrier in the kit of the present invention can be one, can also be multiple, for example, more than two but excellent
It is selected as 1.
The kit can also include：
1. the serum dilution or serum dilution component solution that prepare：Serum dilution, such as there is Beijing to match biotechnology of speeding
The Sample dilution of Co., Ltd（Product identification 070021-S2）, the Zhengzhou bio tech ltd Bo Weijia sample-adding discoloration
Sample dilution（Product identification bwj010103）Deng.The serum dilution is used for dilute serum, and the serum of kit detection wants dilute
Release suitable multiple, such as 2 ~ 200 times, preferably 10 ~ 100 times.
The kit can also include：
2. concentrating washing lotion：After solid carrier surface is incubated serum and ELIAS secondary antibody, solid carrier surface need to be washed with washing lotion not
In conjunction with antibody and ELIAS secondary antibody.Concentration washing lotion is, for example, 1% polysorbas20 aqueous solution, when use need to dilute 2 ~ 40 times, preferably 5 ~
20 times.
The kit can also include：
3. ELIAS secondary antibody solution：IgM in human enterovirus 71 infected person anteserum and solid carrier（Such as SJ modified silica-gels）
On polypeptide combine, ELIAS secondary antibody can be combined with IgM, and the marker on ELIAS secondary antibody can be reacted with assay chromogenic substrate solution, thus
Generate macroscopic colored precipitate.ELIAS secondary antibody can be the goat-anti people IgM of such as horseradish peroxidase-labeled.To enzyme mark
Concentration of the secondary antibody in ELIAS secondary antibody solution is not particularly limited, and can be such as 1ng ~ 1000ng/mL.
The kit can also include：
4. chemical colour reaction substrate solution（TMB, Tetramethylbenzidine, tetramethyl benzidine）：Chemical colour reaction substrate TMB can
It is reacted with the horseradish peroxidase marked on ELIAS secondary antibody, precipitation generates macroscopic bluish violet speckle signal.
5. one or more reaction cavity.
Such as can be biological incubation reactor more than one or two, it may be used according to demand for example two-sided
Biological incubation reactor, referring to Chinese patent ZL 201120177686.3 or ZL201110142518.5；Or the life of single side
Object incubation reaction device, referring to Chinese patent ZL201220430886.x.
The kit can also include：
6. other are used to detect the detection molecules of human enterovirus 71 infection（Such as polypeptide, protein, nucleic acid etc.）.
The kit can also include：
7. operation instructions.
The kit can also include：
8. being connected to the positive quality control point on one or more of solid carriers or other independent solid carriers（It can be used
E.g. people IgM is as positive quality control point）And/or negative Quality Control point（Such as phosphate buffer, abbreviation PB）.
In another aspect, the present invention provide it is a kind of diagnosis subject whether be human enterovirus 71 infection method
（The diagnostic method of the present invention）, this method includes：
The SEQ ID NO are detected using the detection kit of aforementioned present invention：Blood of the polypeptide shown in 1 ~ 6 to subject source
Whether the IgM in liquid sample has response；Wherein,
As the SEQ ID NO：Polypeptide shown in 1 ~ 6 (is sentenced when all having response to the IgM in the blood sample in subject source
According to 1), judgement should subject is a human enterovirns type 71 infection；Otherwise, it is determined that the subject is non-human enterovirus 71 sense
Dye.
In the present specification, " response " refer to：Polypeptide site shows the royal purple chrominance signal stronger than negative control point.
In this specification, the positive reaction to the kit of the present invention refers to：On the blood sample in subject source meets
State criterion 1；Otherwise it is negative reaction.
The blood sample can be whole blood, blood plasma or serum.
In one preferred embodiment, the subject can be the subject with hand-foot-and-mouth disease clinical symptoms.
In the present invention, hand-foot-and-mouth disease clinical symptoms, which are embodied in the positions such as fever and hand, foot, oral cavity, fash or bleb.
In the complete process of the present invention, cooperation unit Shenzhen Children's Hospital has been obtained, Yvonne poly- biomaterials in Suzhou have
The support energetically of limit company indicates sincere and thanks herein.
Specific implementation mode
Embodiment 1
1）The preparation and confirmation of polypeptide
The polypeptide used in embodiment is by gill biochemistry（Shanghai）Co., Ltd synthesizes, and is confirmed to it by mass spectrum.
Wherein,
SEQ ID NO：1 GKGELCAVFRADPGRNGPWQ.
SEQ ID NO：2 EDTHPPYKQTQPGADGFELQ.
SEQ ID NO：3 SLDFALSLLRRNIRQVQTDQ.
SEQ ID NO：4 NKEPAVLHSRDPRLEVDFEQ.
SEQ ID NO：5 GHFTMLGVRDPLAVLPRHSQ.
SEQ ID NO：6 LIREYSNRSAIGNTIEALFQ.
2）The preparation of kit
Detection chip is with SJ modified silica-gels（IPDMS films）It is molten by point sample immobilized polypeptide on it for solid support material
Liquid is prepared.SJ modified silica-gels are that with olefin-terminal, surface is added in traditional polydimethyl siloxane material to cause
The initiator of polymerisation, and pass through heat cross-linking（Si-h bond is bonded）In three-dimensional structure fixed to dimethyl silicone polymer, obtain
To a kind of new material, that is, SJ modified silica-gels.Its manufacturing process is referring to International Patent Publication WO2014/044184.
The SJ modified silica-gel films made can save in 4 DEG C of refrigerators.
Polypeptide microarrays are prepared on modified silica-gel (i.e. using 16 people's point sample instruments of brilliant core PersonalArrayerTM
Detection chip), process is：
1）Pretreatment
By SJ modified silica-gel thin slices（15×15mm2）It is immersed in activating solution, taking-up deionized water elution 3 times after 30min,
It is dried up with nitrogen, is immediately used to point sample.
2）Point sample
Sampling liquid is diluted good and is transferred in the corresponding micropore of 384 orifice plates, 384 orifice plates with sample are placed in point sample instrument base
On platform, while pretreated modified silica-gel thin slice being placed on the base station of point sample instrument, carries out point sample at once.Point sample environmental condition is
Room temperature（25℃）, humidity set 50%.By SEQ ID NO：Polypeptide shown in 1 ~ 6 is put on SJ modified silica-gel thin slices into micro- battle array
It arranges, one point of each polypeptide point sample, in addition one people IgM point of point sample is used as negative matter as positive quality control point and a PB point
Control point.The point sample amount each put on manufactured polypeptide microarrays is about 0.6nL, and sampling point radius is 200 μm.
3）Chemistry is fixed
The polypeptide microarrays just made will be placed on climatic chamber（26 DEG C, 60% humidity）Middle fixation at least 6h.Chemical fixation procedure
Referring to International Patent Publication WO2014/044184.
It will include first to capture the buffer solution point of peptide molecule on modified silica-gel film by point sample instrument, then buffer
Liquid starts to evaporate, and capture peptide molecule and the surface intimate contact of SJ modified silica-gels simultaneously interacts, and passes through and is chemically combined, modified silicon
High molecular end-the COOH of ploy (OEGMA) on glue surface and peptide molecule-- NH2It is formed and stablizes covalent bond, and then will
There is chemically active peptide molecule to be fixed on SJ modified silica-gels surface.
4）Assembly
The polypeptide microarrays of fixed 6h must assemble in two days.SJ modified silica-gel thin slices are attached to specially by gum first
Reaction column on, cover reaction cavity.Biological incubation reactor can use two-sided biological incubation reactor according to demand, that is,
One reactor is made of two reaction columns and a reaction cavity or single side biological incubation reactor, that is, a reactor
It is made of a reaction column and a reaction cavity.
5）It preserves
The polypeptide microarrays assembled are needed to vacuumize sealing, are stored in 4 DEG C of refrigerator, spare.
3）It is detected with kit
Checking procedure：
1) temperature of reagent is balanced：All reagents are balanced to room temperature.
2) preparation of washing lotion：Washing lotion purified water will be concentrated or distilled water presses 1:9 dilutions, such as：450mL purified waters or steaming
50 mL concentration washing lotions are added in distilled water, mix well.The washing lotion being not used is placed on 2 ~ 8 DEG C of preservations, can save 3 months.
3) dilution of sample：Sample to be tested serum dilution is pressed 1:100 dilutions（Such as：2 μ L serum are added to 198 μ L
Serum dilution mixes well）.Sample detection needs the 200 μ L of sample after diluting.
4) wetted chip：Take out the biochip incubation reaction device needed for detection.1mL washing lotions are added, infiltrate chip surface 3
Minute.Discard the washing lotion for impregnating chip.
5) it is loaded：In the present embodiment, it is loaded with single side biological incubation reactor, it is to be measured by what is diluted using liquid-transfering gun
Sample passes through liquid feeding venthole（Any one in two）It is added in single side biological incubation reactor, each single side biological incubation is anti-
It answers device to add 200 μ L, liquid feeding venthole is closed with sealing paste（Two）.
6) sample incubation：The single side biological incubation reactor for adding sample is placed in 150rpm on horizontal shaker, is incubated at room temperature
30 minutes.
7) it cleans：Reaction column is taken out from the reaction cavity of single side biological incubation reactor, discards liquid in reactor,
It rinses inside reaction column top chip surface and reaction cavity 3 times repeatedly（12mL washing lotions are about used every time, are rinsed 1-2 minutes）.
8) enzyme labelled antibody is incubated：Reaction column and reaction cavity are reconfigured, clamping.Throw off sealing paste, each single side life
The 200 μ L of goat-anti people IgM solution of horseradish peroxidase-labeled are added in object incubation reaction device.Again liquid feeding is closed with sealing paste
Venthole.It is placed in 150 rpm on shaking table, is incubated at room temperature 30 minutes.
9) it cleans again：Reaction cavity is discarded, chip surface is carefully cleaned with washing lotion 3 times.
10) assay chromogenic substrate solution is added：Each chip surface is separately added into 100 μ L assay chromogenic substrate solutions, keeps colour developing liquid energy uniform
Be laid on chip surface.
11) result judgement：It for every a serum, observes by the naked eye, whether each polypeptide in statistics kit
There is response（That is the visible royal purple chrominance signal for being better than negative control point of naked eyes）, and according to described in above-mentioned " diagnostic method " part
Criterion 1 judged.
To 588 parts of clinical serum being collected into, using the testing result of mentioned reagent box and using traditional goldstandard method-
Nucleic acid detection method（Using the fluorescent quantificationally PCR detecting kit of Takara）Testing result be compared, as a result such as 1 institute of table
Show.The supplier that 588 parts of above-mentioned clinical serum all has the clinical symptoms of hand-foot-and-mouth disease.
The comparison of the kit of 1 embodiment of table and the fluorescent quantificationally PCR detecting kit of Takara
The above result shows that the present invention is for detecting enterovirns type 71 antibody in human serum（IgM）With good accuracy rate,
Sensitivity and specificity are suitable for the aetology laboratory auxiliary diagnosis of enterovirns type 71, and easy to operate, are suitble to doctors at different levels
It treats and disease control department uses.
More than, more specific description is carried out to the present invention by embodiment, but be not the limit to the technology of the present invention range
It is fixed.By the record of this specification, those skilled in the art readily to the present invention can modify/change, these include
Within the technical scope of the present invention.
SEQUENCE LISTING
<110>Fudan University；Shanghai Public Health Clinical Center
<120>It can be used for diagnosing the kit of human enterovirus 71 infection
<160> 6
<170> PatentIn version 3.2
<210> 1
<211> 20
<212>Amino acid
<213>Artificial sequence
<400> 1
GKGELCAVFRADPGRNGPWQ 20
<210> 2
<211> 20
<212>Amino acid
<213>Artificial sequence
<400> 2
EDTHPPYKQTQPGADGFELQ 20
<210> 3
<211> 20
<212>Amino acid
<213>Artificial sequence
<400> 3
SLDFALSLLRRNIRQVQTDQ 20
<210> 4
<211> 20
<212>Amino acid
<213>Artificial sequence
<400> 4
NKEPAVLHSRDPRLEVDFEQ 20
<210> 5
<211> 20
<212>Amino acid
<213>Artificial sequence
<400> 5
GHFTMLGVRDPLAVLPRHSQ 20
<210> 6
<211> 20
<212>Amino acid
<213>Artificial sequence
<400> 6
LIREYSNRSAIGNTIEALFQ 20
Claims (7)
1. a kind of detection kit comprising one or more solid carriers, and be independently connected on the solid carrier
SEQ ID NO：Polypeptide shown in 1 ~ 6.
2. detection kit according to claim 1, wherein the SEQ ID NO：Polypeptide shown in 1 ~ 6 independently connects
It is connected on same solid carrier.
3. detection kit according to claim 1, wherein the solid carrier is SJ modified silica-gels.
4. detection kit according to claim 1 is used to diagnose human enterovirus 71 infection.
5. a kind of diagnosis subject whether be human enterovirus 71 infection method, this method includes：
The SEQ ID NO are detected using the detection kit described in any one of claim 1 ~ 4：Polypeptide pair shown in 1 ~ 6
Whether the IgM in the blood sample in subject source has response；Wherein,
As the SEQ ID NO：When polypeptide shown in 1 ~ 6 all has response to the IgM in the blood sample in subject source, sentence
It calmly should the infection of subject is a human enterovirns type 71；Otherwise, it is determined that the subject infects for non-human enterovirus 71.
6. according to the method described in claim 5, wherein, the blood sample is whole blood, blood plasma or serum.
7. according to the method described in claim 5, wherein, the subject is the subject with hand-foot-and-mouth disease clinical symptoms.
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| CN201710205811.9A CN108663523A (en) | 2017-03-31 | 2017-03-31 | It can be used for diagnosing the kit of human enterovirus 71 infection |
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| CN201710205811.9A CN108663523A (en) | 2017-03-31 | 2017-03-31 | It can be used for diagnosing the kit of human enterovirus 71 infection |
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| WO2022021902A1 (en) * | 2020-07-28 | 2022-02-03 | 中国科学院武汉病毒研究所 | Broad-spectrum antiviral drug for enterovirus, and application |
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