CN107881257A - The type of Coxsackie virus A 6 and A10 types combined detection kit and its application - Google Patents
The type of Coxsackie virus A 6 and A10 types combined detection kit and its application Download PDFInfo
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Abstract
The invention belongs to field of biomedicine technology, and in particular to a kind of type of Coxsackie virus A 6 and A10 type combined detection kits, including：The type reactant of Coxsackie virus A 6, the type reactant of Coxsackie virus A 10；Wherein, the type reactant of Coxsackie virus A 6 includes：The type sense primer of Coxsackie virus A 6, the type anti-sense primer of Coxsackie virus A 6, the type probe of Coxsackie virus A 6, the structure of the type probe of Coxsackie virus A 6 is 5 ' first the first base sequence of fluorescent reporter group fluorescent quenching groups 3 '；The type reactant of Coxsackie virus A 10 includes：The type sense primer of Coxsackie virus A 10, the type anti-sense primer of Coxsackie virus A 10, the type probe of Coxsackie virus A 10, the structure of the type probe of Coxsackie virus A 10 is 5 ' second the second base sequence of fluorescent reporter group fluorescent quenching groups 3 '；Second fluorescent reporter group is different from first fluorescent reporter group.
Description
Technical field
The invention belongs to field of biomedicine technology, and in particular to a kind of type of Coxsackie virus A 6 and A10 type joint-detections
Kit and its application.
Background technology
Hand-foot-and-mouth disease, it is a kind of children's infectious disease as caused by a variety of enteroviruses.Children are vulnerable crowd within less than 5 years old,
Be mainly shown as oral cavity, hand, foot position fash, a small number of cases can incidence of meningitis, encephalitis, or even pulmonary edema, circulatory failure etc.
Cause death.Cause the pathogen of hand-foot-and-mouth disease to include Coxsackie virus, echovirus and new human enterovirus etc., with EV71
Type and CA16 types are most common.However, in recent years, the case of other enterovirus infections has the trend gradually increased.
The type of Coxsackie virus A 6 and A10 types are in recent years in hand-foot-and-mouth disease cause of disease spectrum in the cause of disease for the trend that is gradually increasing
Body, belong to Coxsackie virus A group membership in enterovirus genus.In recent years year, Spain, Finland, Japan, the U.S., Thailand, Malaysia
There are the report that the type of Coxsackie virus A 6 causes hand-foot-and-mouth disease in West Asia and China, and non-biography occurs in the patient of the infection type of Coxsackie virus A 6
Brothers' mouth symptom of dye, it is mainly shown as mouth week, crissum, the brothers even extensive eruption at multiple positions such as trunk, head, part
Patient there is also decortication, piptonychia etc. in the disease hair later stage.The type of Coxsackie virus A 10 can often cause the disease of some mild, such as hand
Sufficient stomatosis, herpangina etc., but also have the report of the severe cases such as viral meningitis, death both at home and abroad.
Detection product in the market mostly only detects whether it is enterovirus, but cannot distinguish between and which kind of enteron aisle disease belonged to
Poison, only enterovirns type 71 and coxsackie virus A 16-type have an individually detection product, and rarely found have that to distinguish other enteron aisles sick
The product of malicious type.Therefore establish quick, accurate Coxsackie virus A 6 and A10 type products are just particularly important.
The content of the invention
The defects of in order to overcome in the prior art, it is an object of the invention to provide a kind of type of Coxsackie virus A 6 and A10
Type combined detection kit and its application.To realize the type of single tube joint-detection Coxsackie virus A 6 and A10 types.
In order to achieve the above objects and other related objects, in a first aspect, the invention provides a kind of type of Coxsackie virus A 6
With A10 type combined detection kits, including：The type reactant of Coxsackie virus A 6, the type reactant of Coxsackie virus A 10；Wherein, institute
Stating the type reactant of Coxsackie virus A 6 includes：The type sense primer of Coxsackie virus A 6, the type anti-sense primer of Coxsackie virus A 6, Ke Sa
Strange viral A6 types probe, the structure of the type probe of Coxsackie virus A 6 is 5 '-the first fluorescent reporter group the-the first base sequences
Row-fluorescent quenching group -3 '；The type reactant of Coxsackie virus A 10 includes：The type sense primer of Coxsackie virus A 10, Ke Sa
Strange viral A10 types anti-sense primer, the type probe of Coxsackie virus A 10, the structure of the type probe of Coxsackie virus A 10 is 5 '-the
Two the-the second base sequences of fluorescent reporter group-fluorescent quenching group -3 '；Second fluorescent reporter group and described first glimmering
Light reporter group is different.
In a possible implementation, the type of Coxsackie virus A 6 and A10 type combined detection kits also include
Internal contrast product and/or positive reference substance.
In a possible implementation, the base sequence such as SEQ ID of the type sense primer of Coxsackie virus A 6
Shown in NO.1, it is specially：5’-TGTCGGTGAATAGRTTCTA-3’；The base sequence of the type anti-sense primer of Coxsackie virus A 6
Row are as shown in SEQ ID NO.2, specially：5’-CTGTGTCATTTAGYACRTCA-3’；The probe of first base sequence
Sequence is as shown in SEQ ID NO.3, specially：5’-AGCCAGTAGATTCTGTCTTCCARCTC-3’.
In a possible implementation, the base sequence such as SEQ ID of the type sense primer of Coxsackie virus A 10
Shown in NO.4, it is specially：5’-GAAGCAACAGGATTRACA-3’；The base sequence of the type anti-sense primer of Coxsackie virus A 10
Row are as shown in SEQ ID NO.5, specially：5’-GTCCAACTAAGGTRGATTG-3’；The probe sequence of second base sequence
Row are as shown in SEQ ID NO.6, specially：5’-TRATACCAGTGTCCTCGCAGAA-3’.
In a possible implementation, first fluorescent reporter group is FAM, or HEX/VIC/JOE, or Cal
Red 610/ROX/TEXAS RED。
In a possible implementation, second fluorescent reporter group is FAM, or HEX/VIC/JOE, or Cal
Red 610/ROX/TEXAS RED。
In a possible implementation, the fluorescent quenching group is BHQ1.
In a possible implementation, first fluorescent reporter group is FAM, second fluorescent reporter group
For HEX.
In a possible implementation, stating the type of Coxsackie virus A 6 and A10 types combined detection kit also includes Taq
Enzyme and reverse transcriptase.
Second aspect, the invention provides the type of Coxsackie virus A 6 described in first aspect and A10 type combined detection reagents
Purposes of the box in hand-foot-and-mouth disease pathogen detection reagent is prepared.
The third aspect, the invention provides the type of Coxsackie virus A 6 described in first aspect and A10 type combined detection reagents
Purposes of the box in the type of Coxsackie virus A 6 and/or the type detection reagent of Coxsackie virus A 10 is prepared.
Compared with prior art, the beneficial effects of the present invention are：
The type of Coxsackie virus A 6 and A10 types can be detected simultaneously in same reaction tube, improve detection efficiency, and have
Help save sample dosage, less detection of drawing materials can be applied.
Brief description of the drawings
Fig. 1 is the sensitivity technique curve map that the type of Coxsackie virus A 6 detects sample；
Fig. 2 is the sensitivity technique curve that the type of Coxsackie virus A 10 detects sample.
Embodiment
Illustrate embodiments of the present invention below by way of specific instantiation, those skilled in the art can be by this specification
Disclosed content understands the further advantage and effect of the present invention easily.The present invention can also pass through specific realities different in addition
The mode of applying is embodied or practiced, the various details in this specification can also be based on different viewpoints with application, without departing from
Various modifications or alterations are carried out under the spirit of the present invention.
Before the specific embodiment of the invention is further described, it should be appreciated that protection scope of the present invention is not limited to down
State specific specific embodiment；It is also understood that the term used in the embodiment of the present invention is specific specific in order to describe
Embodiment, the protection domain being not intended to be limiting of the invention；In description of the invention and claims, unless in text
Explicitly point out in addition, singulative "one", " one " and " this " include plural form.
When embodiment provides number range, it should be appreciated that except non-invention is otherwise noted, two ends of each number range
Any one numerical value can be selected between point and two end points.Unless otherwise defined, in the present invention all technologies for using and
Scientific terminology is identical with the meaning that those skilled in the art of the present technique are generally understood that.Except used in embodiment specific method, equipment,
Outside material, according to grasp of the those skilled in the art to prior art and the record of the present invention, it can also use and this
Any method, equipment and the material of the similar or equivalent prior art of method, equipment described in inventive embodiments, material come real
The existing present invention.
Unless otherwise indicated, disclosed in this invention experimental method, detection method, preparation method using this technology lead
Domain conventional molecular biology, biochemistry, chromatin Structure and analysis, analytical chemistry, cell culture, recombinant DNA technology and
The routine techniques of association area.These technologies existing perfect explanation in the prior art, for details, reference can be made to the MOLEC such as Sambrook
μLAR CLONING：A LABORATORY MANUAL, Second edition, Cold Spring Harbor Laboratory
Press, 1989and Third edition, 2001；Ausubel etc., CURRENT PROTOCOLS IN MOLEC μ LAR
BIOLOGY, John Wiley＆Sons, New York, 1987and periodic updates；the series METHODS
IN ENZYMOLOGY, Academic Press, San Diego；Wolffe, CHROMATIN STRUCTURE AND
FUNCTION, Third edition, Academic Press, San Diego, 1998；METHODS IN ENZYMOLOGY,
Vol.304, Chromatin (P.M.Wassarman and A.P.Wolffe, eds.), Academic Press, San
Diego, 1999；With METHODS IN MOLEC μ LAR BIOLOGY, Vol.119, Chromatin Protocols
(P.B.Becker, ed.) Humana Press, Totowa, 1999 etc..
Fluorescent RT-PCR technology is the new technology just to grow up late nineteen nineties in last century, it have quick, specificity,
The advantages that high sensitivity, relatively low cost, it is widely used in the various aspects of clinical detection.Develop at present and utilized fluorescence
Quantitative PCR technique is directed to the total type of enterovirus or the quick detection kit of enterovirns type 71 and coxsackie virus A 16-type.
Therefore the present invention establishes the side of the type of single tube joint-detection Coxsackie virus A 6 and A10 types on the basis of fluorescent RT-PCR technology
Method has very important significance.
In embodiments of the present invention, using Taqman quantitative fluorescent PCR principles, respectively for the type of Coxsackie virus A 6 and
A10 types design specific primer, specific amplification nucleotide sequence, while separately design Taqman probes, and mark different glimmering
Light reporter group, between upstream and downstream primer.Its 5 ' end mark fluorescent reporter group of probe, 3 ' end marks are non-fluorescence to be quenched base
Group.When probe is complete, the fluorescent energy that reporter group is launched is quenched group absorptions, and instrument can't detect signal.
With PCR progress, Taq enzyme runs into the probe combined with template during chain extension, its 5 ' → 3 ' exonuclease activity
Probe will be cut off, reporter group can not be absorbed away from quenching group, its energy, that is, produce fluorescence signal.Therefore, this hair
The fluorescent quantitative PCR technique of bright use has the characteristics that detection in real time, the detection of quantitative and high flux, and easy to operate, sensitivity
The advantages that high, specific good.
The Coxsackie virus A 6 of embodiment 1. and A10 types combined detection kit are to the type of Coxsackie virus A 6 and A10 in mark product
Type detects
11st, nucleic acid fluorescent PCR detection mixed liquors are prepared：
Take the μ l/test of 6 type sense primer of Coxsackie virus A 0.5, the μ l/test of anti-sense primer 0.5, the μ l/test of probe 0.1；
The μ l/test of 10 type sense primer of Coxsackie virus A 0.5；The μ l/test of anti-sense primer 0.5；The μ l/test of probe 0.1；Internal contrast
The μ l/test of product probe 0.1；The μ l/test of real-time quantitative PCR (Realtime-PCR, RT-PCR) MIX 12.5；The μ of process water 3.2
l/test；Mix is the type of Coxsackie virus A 6 and A10 type nucleic acid fluorescents PCR detection mixed liquors.
Wherein, the base sequence of the type sense primer of Coxsackie virus A 6 is specially as shown in SEQ ID NO.1：5’-
TGTCGGTGAATAGRTTCTA-3’；The base sequence of the type anti-sense primer of Coxsackie virus A 6 as shown in SEQ ID NO.2,
Specially：5’-CTGTGTCATTTAGYACRTCA-3’；The structure of the type probe of Coxsackie virus A 6 is 5 '-the first fluorescence reports
Accuse the base sequence of group-the first-fluorescent quenching group -3 '；Wherein, the first fluorescent reporter group is FAM, and fluorescent quenching group is
BHQ1, the first base probe sequence are specially as shown in SEQ ID NO.3：5’-AGCCAGTAGATTCTGTCTTCCARCTC-
3’。
The base sequence of the type sense primer of Coxsackie virus A 10 is as shown in SEQ ID NO.4, specially：5’-
GAAGCAACAGGATTRACA-3’；The base sequence of the type anti-sense primer of Coxsackie virus A 10 as shown in SEQ ID NO.5,
Specially：5’-GTCCAACTAAGGTRGATTG-3’；The structure of the type probe of Coxsackie virus A 10 is 5 '-the second fluorescence reports
Accuse group-the second base sequence-fluorescent quenching group -3 ', second fluorescent reporter group is HEX, the fluorescent quenching base
Group is BHQ1, and the probe sequence of second base sequence is specially as shown in SEQ ID NO.6：5’-
TRATACCAGTGTCCTCGCAGAA-3’.In an alternative, the second fluorescent reporter group can also be VIC/JOE.
The fluorescent reporter group of internal contrast product probe is ROX, and internal contrast product probe is glimmering described in alternative solution
Light reporter group can also be Cal Red 610/TEXAS RED.
Contain dNTP, buffer solution etc. in RT-PCR MIX.
12nd, the nucleic acid extraction of sample：
The total serum IgE in sample is extracted with commercialized viral RNA extraction agent box；Wherein, sample can have with clinical sample
Body can be throat swab, or excrement.
13rd, PCR reaction systems are prepared：
131st, preparation of reagents：
Take the type of Coxsackie virus A 6 and A10 type nucleic acid fluorescents PCR detection mixed liquors and n × 1 prepared by the μ l steps 11 of n × 18
μ l internal contrast product, the μ lRT-PCR enzymes (n is reaction tube number) of and n × 1, vibration mix several seconds, 3000rpm centrifugation several seconds.RT-
PCR enzymes include hot start Taq polymerase and reverse transcriptase.
132nd, it is loaded：
The μ l of mixed liquor 20 for taking step 131 to prepare are placed in thin-walled PCR pipe or PCR reaction plates, then by the sample of extraction
Total serum IgE, positive reference substance, DEPC-H2Each 5 μ l of O are separately added into thin-walled PCR reaction tubes or PCR reaction plates, cover thin-walled
PCR reaction lids or PCR reaction plate films, pcr amplification reaction is carried out immediately after centrifuging the several seconds.
133rd, PCR is expanded：
Thin-walled PCR pipe in step 132 or PCR reaction plates are placed on quantitative fluorescence PCR instrument, loop parameter can be：
45℃×10min；95℃×15min；95 DEG C × 15sec → 60 DEG C × 60sec is pressed again, is circulated 40 times；Single-point is glimmering
Light is detected at 60 DEG C.Reaction system is 25 μ l.
134th, baseline and threshold value setting：
Baseline adjustment takes the fluorescence signal of 6-15 circulation, and threshold value setting principle is just above DEPC-H with threshold line2O is examined
Survey the peak of fluorescence curve.
135th, quality control：
Positive reference substance and internal contrast product must reach following requirement as shown in table 1, and it is invalid otherwise to test.
Table 1
136th, experimental result judges：
Experimental result is as shown in table 2.
Table 2
In embodiment 1, relate to 2 pairs of PCR primers and probe respectively for the type of specific amplification Coxsackie virus A 6 and
A10 types, 2 pairs of PCR primers can expand simultaneously in same PCR reaction tubes, Multiple detection be realized, when greatly shortening detection
Between, it is easy to operate.Specific primer and the highly conserved and specific of primer and probe is ensure that with the design of probe, avoided
Situation without complementary pairing or intersection amplification between two pairs of primer and probes.The fluorescence for three fluorophors that embodiment 1 is selected
Wavelength differs that larger and signal intensity is close, avoids interfering between signal.
The sensitivity analysis of the Coxsackie virus A 6 of embodiment 2. and A10 type combined detection kits.
21st, detection sample is prepared：
Take 1 × 107The copies/ml type plasmid (CA6-S1) of Coxsackie virus A 6 presses 1：10、1：100、1：1000、1：
10000 dilutions, obtain CA6-S2 (1 × 106copies/ml)、CA6-S3(1×105copies/ml)、CA6-S4(1×
104copies/ml)、CA6-S5(1×103Copies/ml) totally 5 parts of conduct Coxsackie virus A 6 types detect sample.
Take 1 × 107The copies/ml type plasmid (CA10-S1) of Coxsackie virus A 10 presses 1：10、1：100、1：1000、1：
10000 dilutions, obtain CA10-S2 (1 × 106copies/ml)、CA10-S3(1×105copies/ml)、CA10-S4(1×
104copies/ml)、CA10-S5(1×103Copies/ml) totally 5 parts of conduct Coxsackie virus A 10 types detect sample.
22nd, reagent preparation：
The μ l of mixed liquor 20 prepared by the step 131 in Example 1 are placed in thin-walled PCR pipe or PCR reaction plates, then will
Each detection sample, positive reference substance, the DEPC-H of step 12 preparation2Each 5 μ l of O are separately added into thin-walled PCR reaction tubes or PCR reactions
In plate, thin-walled PCR reaction lids or PCR reaction plate films are covered, pcr amplification reaction is carried out immediately after centrifuging the several seconds.
23rd, PCR is expanded：
Thin-walled PCR pipe in step 132 or PCR reaction plates are placed on quantitative fluorescence PCR instrument, loop parameter can be：
45℃×10min；95℃×15min；95 DEG C × 15sec → 60 DEG C × 60sec is pressed again, is circulated 40 times；Single-point is glimmering
Light is detected at 60 DEG C.Reaction system is 25 μ l.
24th, testing result：
The testing result of each inspection sample is as shown in table 3.The type of Coxsackie virus A 6 detects the sensitivity technique curve of sample
As shown in Figure 1；The type of Coxsackie virus A 10 detection sample is as shown in Figure 2.
Table 3
Remarks：+ represent positive findings, that is, detect the type of Coxsackie virus A 6 or the type of Coxsackie virus A 10
Table 3 shows, for the type plasmid of Coxsackie virus A 6, CA6-S1 (1 × 107copies/ml)—CA6-S5(1×
103Copies/ml) equal test positive, show that the detection sensitivity of the kit detection type of Coxsackie virus A 6 can reach 1 ×
103copies/ml.For the type plasmid of Coxsackie virus A 10, CA10-S1 (1 × 107copies/ml)——CA10-S5(1×
103Copies/ml) equal test positive, 1 × 10 can be reached by showing the detection sensitivity of kit detection A10 types3copies/
ml。
In summary, the type of Coxsackie virus A 6 and A10 types can be detected simultaneously in same reaction tube, improves detection effect
Rate, and help to save sample dosage, less detection of drawing materials can be applied.
Embodiment above is to illustrate embodiment disclosed by the invention, can not be interpreted as the limit to the present invention
System.In addition, in various modifications and invention listed herein method, composition change, do not departing from the scope of the present invention
Be obvious for those skilled in the art on the premise of spirit.Although a variety of specific of the present invention has been combined
Preferred embodiment has carried out specific description to the present invention, it is to be understood that, the present invention should not be limited only to these specific embodiments.
In fact, various, obvious modification should all include to obtain invention for those skilled in the art as described above
Within the scope of the invention.
Sequence table
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<120>Coxsackie virus A 6 and A10 types joint combined detection kit and its application
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Claims (10)
1. a kind of type of Coxsackie virus A 6 and A10 type combined detection kits, it is characterised in that including：The type of Coxsackie virus A 6
Reactant, the type reactant of Coxsackie virus A 10；Wherein,
The type reactant of Coxsackie virus A 6 includes：The type sense primer of Coxsackie virus A 6, the type downstream of Coxsackie virus A 6 are drawn
Thing, the type probe of Coxsackie virus A 6, the structure of the type probe of Coxsackie virus A 6 is 5 '-the first fluorescent reporter groups-the first
Base sequence-fluorescent quenching group -3 '；
The type reactant of Coxsackie virus A 10 includes：The type sense primer of Coxsackie virus A 10, the type downstream of Coxsackie virus A 10
Primer, the type probe of Coxsackie virus A 10, the structure of the type probe of Coxsackie virus A 10 for 5 '-the second fluorescent reporter groups-
Second base sequence-fluorescent quenching group -3 '；Second fluorescent reporter group is different from first fluorescent reporter group.
2. the type of Coxsackie virus A 6 according to claim 1 and A10 type combined detection kits, it is characterised in that also wrap
Include internal contrast product and/or positive reference substance.
3. the type of Coxsackie virus A 6 according to claim 1 and A10 type combined detection kits, it is characterised in that described
The base sequence of the type sense primer of Coxsackie virus A 6 is as shown in SEQ ID NO.1, specially：5’-
TGTCGGTGAATAGRTTCTA-3’；
The base sequence of the type anti-sense primer of Coxsackie virus A 6 is as shown in SEQ ID NO.2, specially：5’-
CTGTGTCATTTAGYACRTCA-3’；
The probe sequence of first base sequence is as shown in SEQ ID NO.3, specially：5’-
AGCCAGTAGATTCTGTCTTCCARCTC-3’。
4. the type of Coxsackie virus A 6 according to claim 1 and A10 type combined detection kits, it is characterised in that described
The base sequence of the type sense primer of Coxsackie virus A 10 is as shown in SEQ ID NO.4, specially：5’-
GAAGCAACAGGATTRACA-3’；
The base sequence of the type anti-sense primer of Coxsackie virus A 10 is as shown in SEQ ID NO.5, specially：5’-
GTCCAACTAAGGTRGATTG-3’；
The probe sequence of second base sequence is as shown in SEQ ID NO.6, specially：5’-
TRATACCAGTGTCCTCGCAGAA-3’。
5. the type of Coxsackie virus A 6 according to claim 1 and A10 type combined detection kits, it is characterised in that described
First fluorescent reporter group is FAM, or HEX/VIC/JOE, or Cal Red 610/ROX/TEXAS RED.
6. the type of Coxsackie virus A 6 according to claim 1 and A10 type combined detection kits, it is characterised in that described
Second fluorescent reporter group is FAM, or HEX/VIC/JOE, or Cal Red 610/ROX/TEXAS RED.
7. the type of Coxsackie virus A 6 according to claim 1 and A10 type combined detection kits, it is characterised in that described
Fluorescent quenching group is BHQ1.
8. the type of Coxsackie virus A 6 according to claim 1 and A10 type combined detection kits, it is characterised in that also wrap
Include Taq enzyme and reverse transcriptase.
9. the type of Coxsackie virus A 6 and A10 types combined detection kit described in claim any one of 1-8 are preparing brothers' mouth
Purposes in encephalapthy agent detection reagent.
10. the type of Coxsackie virus A 6 and A10 types combined detection kit described in claim any one of 1-8 are preparing COxsackie
Purposes in the viral type detection reagent of A6 types and/or Coxsackie virus A 10.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109856408A (en) * | 2019-04-09 | 2019-06-07 | 潍坊市康华生物技术有限公司 | A kind of 6 type of Coxsackie virus A and A10 type IgM antibody combined detection kit and preparation method thereof |
| CN109917140A (en) * | 2019-04-09 | 2019-06-21 | 潍坊市康华生物技术有限公司 | A kind of reagent strip and preparation method thereof of 6 type of Coxsackie virus A and A10 type IgM antibody joint-detection |
| CN113564130A (en) * | 2021-09-23 | 2021-10-29 | 北京民海生物科技有限公司 | Coxsackie virus A10 type strain and application thereof |
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| CN103789451A (en) * | 2014-01-12 | 2014-05-14 | 浙江大学 | Fluorescent quantitative kit for detecting coxsackie viruses A6 and A10 |
| CN104195264A (en) * | 2014-08-13 | 2014-12-10 | 江苏硕世生物科技有限公司 | Nucleic acid test kit for rapidly detecting Coxsackie virus A6 type/A10 type and application thereof |
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| CN109856408A (en) * | 2019-04-09 | 2019-06-07 | 潍坊市康华生物技术有限公司 | A kind of 6 type of Coxsackie virus A and A10 type IgM antibody combined detection kit and preparation method thereof |
| CN109917140A (en) * | 2019-04-09 | 2019-06-21 | 潍坊市康华生物技术有限公司 | A kind of reagent strip and preparation method thereof of 6 type of Coxsackie virus A and A10 type IgM antibody joint-detection |
| CN113564130A (en) * | 2021-09-23 | 2021-10-29 | 北京民海生物科技有限公司 | Coxsackie virus A10 type strain and application thereof |
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