Ikhithi ye-Magnetic Universal Viral DNA/RNA HYD521
I-Magnetic Universal Viral DNA/RNA Kit ifanele ukukhipha i-viral nucleic acid egazini eliphelele noma emanzini omzimba angenamaseli (njenge-serum, i-plasma, uketshezi lwe-cerebrospinal, ama-nasal/pharyngeal swabs, uketshezi lokuhlanza i-alveolar, njll.). Le ndlela isebenzisa ubuchwepheshe bokuhlanza ama-magnetic bead, futhi akukho ukukhishwa kwe-phenol chloroform okunobuthi okudingekayo ngesikhathi senqubo yokukhipha. Iphephile, ayinabo ubuthi futhi iyashesha. Umkhiqizo otholiwe ungasetshenziswa ngqo ku-PCR, qPCR, ukulandelana kwesizukulwane sesibili kanye nezinye izivivinyo. Uma isetshenziswa nezinsimbi zokukhipha ezizenzakalelayo zama-magnetic bead, ukukhishwa okuphezulu kwama-nucleic acid kungatholakala.
Ulwazi lomkhiqizo
| Izingxenye | 48T |
| Isixazululo se-Lysis kanye ne-Binding | 33 mL |
| Ukumiswa kobuhlalu obunobuthi | 1.1 mL |
| I-buffer yokuwasha A | 40 ml |
| I-buffer yokuwasha B | 80 ml |
| Isixazululo se-Elution | 10 ml |
| I-Proteinase K | 1.1 mL |
Izimo Zokugcina
I-Proteinase K kufanele igcinwe ku-2 ~ 8℃ kanti ezinye izithako kufanele zigcinwe ekushiseni kwegumbi. Isikhathi sokuphila seshelufu siyizinyanga ezingu-18.
Incazelo Yesampula
1. Izinhlobo zesampula ezisebenzayo: igazi eliphelele, i-serum, i-plasma, uketshezi lwe-cerebrospinal, ama-nasal/pharyngeal swabs, uketshezi lwe-alveolar lavage kanye nezinye izibonelo.
2. Ukugcinwa kanye nokuthuthwa kwesampula: Isampula ingasetshenziswa ukuhlolwa ngokushesha noma igcinwe ku--70℃ noma ngaphansi ukuze ihlolwe. Isikhathi sokugcina siyizinyanga eziyi-6. Ukuqandisa okuphindaphindiwe kanye nokuncibilika kufanele kugwenywe. Isampula ithuthwa ngeketanga elibandayo.
3. Izidingo zokuqandisa nokuncibilikisa: Ukuqandisa nokuncibilikisa ngokushesha, gwema ukuqandisa nokuncibilikisa okuphindaphindiwe.
Ukusebenza
Ngaphambi kokuhlola, hlola ukuthi ikhambi linemvula yini nokuthi ama-magnetic beads angaphinde axhunywe yini.
1. Dlulisa isampula engu-200~300 μL kushubhu le-centrifuge elingu-1.5 mL, engeza i-600 μL ye-Lysis and Binding Solution, i-20 μL ye-Proteinase K, kanye ne-20 μL ye-magnetic bead suspension ngokulandelana, kanye ne-vortex ngesivinini esiphezulu imizuzwana engu-30. Faka ku-50℃ imizuzu engu-5~7. Uma i-thermostat ingenawo umsebenzi wokunyakaza, faka i-vortex izikhathi ezintathu ngesikhathi sokufukamela, isikhathi ngasinye imizuzwana engu-15.
2. Dlulisela esitendini sikamazibuthe esingu-1.5 mL ukuze kuhlukaniswe umazibuthe kuze kube yilapho ikhambi selicacile futhi licacile, bese ulahla ikhambi.
3. Engeza i-700 μL ye-wash buffer A, i-vortex imizuzwana eyi-10 ukuze uhlukanise ubuhlalu obunamandla, bese udlulisela esitendini esinamandla ukuze kuhlukaniswe amandla kuze kube yilapho ikhambi selicacile, bese ulahla ikhambi.
4. Engeza i-700 μL ye-wash buffer B, i-vortex imizuzwana eyi-10 ukuze uhlukanise ubuhlalu obunamandla, udlulisele esitendini esinomagnetic ukuze kuhlukaniswe amandla kagesi kuze kube yilapho ikhambi selicacile, bese ulahla ikhambi.
5. Engeza u-700 μL we-wash buffer B futhi, uvale i-vortex imizuzwana eyi-10 ukuze uhlukanise ubuhlalu obunamandla, udlulisele esitendini esinamandla ukuze kuhlukaniswe amandla kuze kube yilapho ikhambi selicacile, bese ulahla ikhambi.
6. Faka i-centrifuge isikhashana ukuze uqoqe amaconsi odongeni lwepayipi, uwadlulisele endaweni yokuma kagesi ukuze acwebezele, bese umunca uketshezi olusele. Yomisa emoyeni imizuzu emi-3-5.
Qaphela: Izinsalela ze-ethanol zizovimbela ukusabela okulandelayo kwe-enzyme, ngakho-ke qiniseka ukuthi i-ethanol iyahwamuka ngokuphelele lapho yoma. Ungomi isikhathi eside ukuze ugweme ukuthinta umphumela olandelayo we-elution.
7. Engeza isixazululo se-Elution esingu-40~100 μL, i-vortex ngesivinini esiphezulu imizuzu emi-2~3 ukuze uhlukanise ubuhlalu obunamandla. Faka ku-60℃ imizuzu emi-5, bese ufaka i-vortex ngesivinini esiphezulu imizuzwana engama-60.
8. Faka i-centrifuge esheshayo ukuze uqoqe uketshezi esivalweni sethubhu bese uludlulisela endaweni yokushaja kagesi kuze kube yilapho ubuhlalu be-magnetic bumuncwa ngokuphelele. Bese udlulisela uketshezi ngokucophelela kuthubhu entsha ye-centrifuge ukuze uthole isixazululo se-nucleic acid.
9. Isixazululo se-nucleic acid singagcinwa ku--20°C ukuze sigcinwe isikhathi esifushane kanye no--80°C ukuze sigcinwe isikhathi eside.
Amanothi
1. Ama-buffer ahlukahlukene akule khithi aqukethe usawoti we-guanidine. Ukuze uphephe futhi uphile kahle, sicela ugqoke ijazi lelebhu kanye namagilavu angasetshenziswa. Futhi uphathe ngokwezinyathelo zokuphepha ezijwayelekile. Ungavumeli i-buffer ithintane nesikhumba, amehlo kanye nolwelwesi lwamafinyila. Uma kwenzeka, sicela ugeze ngokushesha ngamanzi amaningi bese ucela usizo lwezokwelapha.
2. Uma isixazululo sivuthuluka, sidinga ukugezwa emanzini angu-30°C kuze kube yilapho i-precipitate ihlakazeka ngokuphelele ngaphambi kokusetshenziswa.
3. Uma ukumiswa kobuhlalu obunobuthi kuqandisiwe, ungakusebenzisi.
4. Ama-buffer ahlukahlukene akule khithi aqukethe usawoti we-guanidine. Ungasebenzisi izibulali-magciwane ezikhipha i-oxidizing njenge-sodium hypochlorite ukuze uziphathe, kungenjalo kuzokhishwa amagesi anobuthi futhi kumele aphathwe njengemfucuza yezokwelapha.
5. Kungase kube khona ama-magnetic bead asele ngesikhathi sokuhlanza. Zama ukugwema ukuphefumula ama-magnetic beads lapho uphefumula amasampula.