Products
Lifecosm SARS-Cov-2-RT-PCR Detection kit ye2019-nCoV
Kushandiswa kunotarisirwa
Iyi kit inoshandiswa pakuongorora kwemhando yekoronavirus itsva (2019-nCoV) uchishandisa huro swabs, nasopharyngeal swabs, bronchoalveolar lavage fluid, sputum. humbowo hwekuongororwa kwekiriniki uye kurapwa.Kuongorora kwakadzama kwemamiriro ezvinhu kunokurudzirwa pamwe chete nezviratidzo zvechipatara zvemurwere uye mamwe ma laboratory bvunzo.
Inspection musimboti
Iyo kit yakavakirwa pane imwe-nhanho RT-PCR tekinoroji.Muchokwadi, iyo 2019 nyowani coronavirus (2019-nCoV) ORF1ab uye N genes dzakasarudzwa senzvimbo dzakanangwa dzekusimudzira.MaSpecific primers uye fluorescent probes (N gene probes akanyorwa neFAM uye ORF1ab probes akanyorwa neHEX) akagadzirirwa kuona 2019 nyowani-mhando coronavirus RNA mumasample.Iyo kit inosanganisirawo endogenous yemukati yekudzora yekuongorora system (yemukati yekudzora gene probe yakanyorwa neCY5) yekutarisa maitiro ekuunganidza sampuli, RNA uye PCR amplification, nekudaro ichidzikisa nhema dzisina kunaka mhedzisiro.
Main components
| Zvikamu | Volume(48T/Kit） |
| RT-PCR reaction solution | 96µl |
| nCOV primer TaqMan probemixture (ORF1ab,N Gene,RnaseP Gene) | 864µl |
| Negative control | 1500µl |
| nCOV Positive contro (l ORF1ab N Gene) | 1500µl |
Own reagents: RNA kudhirowa kana kuchenesa reagents.Negative/positive control: Iyo yakanaka control iRNA ine chidimbu chakatarwa, nepo iyo yakaipa kutonga iri nucleic acid-isina mvura.Panguva yekushandiswa, ivo vanofanirwa kutora chikamu mukuchera uye vanofanirwa kunzi vane hutachiona.Dzinofanirwa kubatwa uye kuraswa maererano nemitemo yakakodzera.
Iyo yemukati referensi gene ndeyemunhu RnaseP gene.
Mamiriro ekuchengetedza uye zuva rekupera
-20 ± 5 ℃, dzivisa kuomesa nechando uye kunyunguduka kanopfuura ka5, inoshanda kwemwedzi mitanhatu.
Chimbo chinoshanda
Iine FAM / HEX / CY5 uye mamwe akawanda-channel fluorescent PCR chiridzwa.
Zvido zvemuenzaniso
1.Anoshanda emhando dzemuenzaniso: swabs pahuro, nasopharyngeal swabs, bronchoalveolar lavage fluid, sputum.
2.Specimen collection (aseptic technique)
Pharyngeal swab: Pukuta matonsi uye posterior pharyngeal madziro nemaswabhi maviri panguva imwe chete, wobva wanyudza musoro we swab muchubhu yekuyedza ine sampling solution.
Sputum: Mushure mekunge murwere ane chikosoro chakadzika, tora sputum yakakosora mune screw cap test tube ine sampling solution;bronchoalveolar lavage fluid: Sampling nenyanzvi dzekurapa.3.Kuchengetedza uye kutakura kwemasampuli
Mienzaniso yekusarudzika kwehutachiona uye kuongororwa kweRNA inofanirwa kuongororwa nekukurumidza sezvinobvira.Mienzaniso inogona kuonekwa mukati maawa makumi maviri nemana inogona kuchengetwa pa 4 ℃;izvo zvisingagoni kuonekwa mukati me24
maawa anofanirwa kuchengetwa pa -70 ℃ kana pazasi (kana pasina mamiriro ekuchengetedza -70 ℃, anofanirwa kunge ari
yakachengetwa kwenguva pfupi pa -20 ℃firiji).Mienzaniso inofanira kudzivisa kuomesa nechando uye kunyunguduka panguva yekufambisa.Mienzaniso inofanira kutumirwa ku laboratori nokukurumidza sezvinobvira mushure mekuunganidza.Kana sampuli dzichida kutakurwa madaro marefu, kuchengetwa kweaizi yakaoma kunokurudzirwa.
Miedzo Nzira
1 Sample process uye RNA kudhirowa (sample processing area)
Zvinokurudzirwa kutora 200μl yemvura sampu yeRNA kutorwa.Nematanho ekubvisa ane hukama, tarisa kune mirairo yekutengeserana RNA yekubvisa kits.Zvose zvakaipa nezvakaipa
zvinodzora mukiti iyi zvakabatanidzwa mukubvisa.
2 PCR reagent kugadzirira (reagent nzvimbo yekugadzirira)
2.1 Bvisa zvese zvikamu kubva mukiti uye nyungudutsa uye sanganisa pakamuri tembiricha.Centrifuge pa8,000 rpm kwemasekonzi mashoma isati yashandiswa;verenga huwandu hunodiwa hwema reagents, uye iyo reaction system inogadzirirwa sezvinoratidzwa mune inotevera tafura:
| Zvikamu | N kushanda (25µl system) |
| nCOV primer TaqMan probemixture | 18µl × N |
| RT-PCR reaction solution | 2 µl × N |
| *N = nhamba yemasampuli akaedzwa + 1 (negative control) + 1 (nCOVkudzora kwakanaka) | |
2.2 Mushure mekunyatsosanganisa izvo zvinoumba, centrifuge kwenguva pfupi kurega mvura yese iri pachubhu madziro ichiwira pasi pechubhu, uyezve aliquot iyo 20 µl amplification system muPCR chubhu.
3 Sampling (nzvimbo yekugadzirira nzvimbo)
Wedzera 5μl yezvisina kunaka uye zvakanaka zvinodzora mushure mekutora.Iyo RNA yemuyedzo ichaedzwa inowedzerwa kune PCR reaction chubhu.
Bata chubhu zvakasimba uye centrifuge pa8,000 rpm kwemasekonzi mashoma usati waiendesa kunzvimbo yekuona amplification.
4 PCR kukwidziridzwa (yakawedzera yekuona nzvimbo)
4.1 Isa iyo reaction chubhu muyemuenzaniso sero yechiridzwa, uye isa paramita sezvizvi:
| stage | Cycle nhamba | Tembiricha(°C) | Nguva | collectionsite |
| Reversetranscription | 1 | 42 | 10min | - |
| Pre-denaturation | 1 | 95 | 1min | - |
| Cycle | 45 | 95 | 15s | - |
| 60 | 30s | data collection |
Kusarudzwa kwechiteshi chezviridzwa: Sarudza iyo FAM, HEX, CY5 chiteshi chechiratidzo chefluorescence.Zvereferensi fluorescent HAPANA, ndapota usasarudza ROX.
5 Mhedzisiro yekuongorora (Ndokumbira utarise kune mirairo yekuyedza yechiridzwa chimwe nechimwe chekumisikidza)
Mushure mekuita, chengetedza mhinduro.Mushure mekuongorora, gadzirisa kukosha kwekutanga, kukosha kwekupedzisira, uye kukosha kwechikumbaridzo chepakutanga maererano nemufananidzo (mushandisi anogona kugadzirisa maererano nemamiriro ezvinhu chaiwo, kukosha kwekutanga kunogona kuiswa ku3 ~ 15, kukosha kwekupedzisira kunogona kuiswa 5 ~ 20, kugadzirisa) mugirafu ye logarithmic Pachikumbaridzo chehwindo, mutsara wepakati uri muchikamu chelogarithmic, uye mutsara wekusimudzira wehutori husina kunaka mutsara wakatwasuka kana pasi pemutsetse).
6 Quauty control (A procedural control inosanganisirwa muyedzo) Negative control: Hapana pachena amplification curve yeFAM, HEX, CY5 yekuona nziran
COV positive control: zviri pachena amplification curve yeFAM neHEX yekuona nzira, Ct value≤32, asi hapana amplification curve yeCY5 chiteshi;
Izvo zvinodiwa pamusoro apa zvinofanirwa kusangana panguva imwe chete mukuedza kumwe chete;kana zvisina kudaro, kuedza hakuna kushanda uye kunoda kudzokororwa.
7 Kusarudza mhedzisiro.
7.1 Kana pasina amplification curve kana Ct value> 40 muFAM neHEX chiteshi che test sample, uye paine amplification curve muCY5 chiteshi, zvinogona kutongwa kuti hapana 2019 new coronavirus (2019-nCoV) RNA mumuenzaniso;
.2 Kana test sample iine amplification curves aripachena muFAM neHEX chiteshi, uye kukosha kweCt kuri ≤40, zvinogona kutongwa kuti sampu iyi yakanaka kune 2019 new coronavirus (2019-nCoV).
7.3 Kana sampu yekuyedza iine yakajeka amplification curve chete mune imwe chiteshi cheFAM kana HEX, uye kukosha kweCt iri ≤40, uye pasina amplification curve mune imwe chiteshi, mhedzisiro inoda kuongororwazve.Kana mhinduro dzekudzokorora dzichienderana, sampuli inogona kunzi yakanaka kune itsva
coronavirus 2019 (2019-nCoV).Kana retest mhedzisiro isiriyo, inogona kutongwa kuti sampu haina kunaka ye2019 nyowani coronavirus (2019-nCoV).
Positive kutonga kukosha
Iyo ROC curve nzira inoshandiswa kuona iyo yereferensi CT kukosha kwekiti uye iyo yemukati control referensi kukosha ndeye 40.
Kuturikira mhinduro dzebvunzo
1.Kuedza kwega kwega kunofanira kuongororwa kune zvakaipa uye zvakanaka kutonga.Mhedzisiro yebvunzo inogona kutariswa chete kana zvidzori zvasangana nemhando yekudzora zvinodiwa
2.Apo FAM neHEX yekuona nzira dzakanaka, chigumisiro chinobva kuCY5 channel (internal control channel) inogona kunge isina kunaka nekuda kwemakwikwi ehurongwa.
3.Kana chigumisiro chemukati chekutonga chisina kunaka, kana tthe test tube FAM neHEX yekuona nzira dzisina kunaka, zvinoreva kuti hurongwa hunovharwa kana kushanda kwakashata, t muedzo hauna kukodzera.Naizvozvo, masampula anofanira kuongororwa zvakare.
