Olink NextSeq 2000 Onani Buku Logwiritsa Ntchito Njira Yotsatirira

Olink® Explore ndi nsanja ya multiplex immunoassay pakutulukira kwa protein biomarker ya anthu. Mankhwalawa amapangidwa kuti agwiritsidwe ntchito pa Research Use Only, osati kuti agwiritsidwe ntchito pofufuza. Ntchito ya labotale idzayendetsedwa ndi ogwira ntchito ophunzitsidwa bwino a labotale. Kukonza deta kudzachitidwa ndi ogwira ntchito ophunzitsidwa bwino. Zotsatirazi zimayenera kugwiritsidwa ntchito ndi ochita kafukufuku mogwirizana ndi zochitika zina zachipatala kapena ma laboratory.

Za bukuli

Buku la Wogwiritsa Ntchitoli limapereka malangizo ofunikira kuti atsatire Olink® Explore Libraries pa Illumina® NextSeq™ 2000. Malangizowo ayenera kutsatiridwa mosamalitsa komanso momveka bwino. Kupatuka kulikonse pamasitepe a labotale kumatha kubweretsa kuwonongeka kwa data. Musanayambe kayendedwe ka labotale, funsani Olink® Explore Overview Buku lachidziwitso pa nsanja, kuphatikiza zambiri za ma reagents, zida ndi zolemba zofunika, kupitiliraview mayendedwe a ntchito, komanso malangizo a labotale. Kuti mumve malangizo amomwe mungayendetsere Olink® Explore Reagent Kits, onetsani ku Olink® Explore User Manual. Kuti mudziwe zambiri komanso kusanthula zotsatira za Olink® Explore, onani Olink® MyData Cloud User Guide. Zizindikiro zonse ndi kukopera zomwe zili m'nkhaniyi ndi za Olink® Proteomics AB, pokhapokha zitanenedwa.

Othandizira ukadaulo

Kuti mupeze chithandizo chaukadaulo, lemberani Olink Proteomics pa: support@olink.com.

Malangizo a Laboratory

Mutuwu umapereka malangizo amomwe mungatsatire Olink Libraries pa NextSeq™ 2000 pogwiritsa ntchito NextSeq™ 1000/2000 P2 Reagents (100 Cycles) v3. Protocol yomwe imagwiritsidwa ntchito pakutsatizana ndikusinthidwa kwa Illumina® standard NGS workflow kwa Illumina® NextSeq™ 2000. Musanayambe kutsatizana, onetsetsani kuti ubwino wa Library ya Olink yoyeretsedwa yatsimikiziridwa. Onani ku Olink Explore User Manual kuti mupeze malangizo okhudza kuwongolera khalidwe.

Konzani ndondomeko yotsatizana

Laibulale imodzi ya Olink imatha kutsatiridwa pa NextSeq™ 2000 P2 cell cell ndikuthamanga. Chiwerengero cha maselo othamanga a P2 ndi kuthamanga kofunikira kuti atsatire zosiyana za Olink Explore Reagent Kits zikufotokozedwa mu Table 1. Ngati kupitirira kumodzi kukufunika, bwerezani malangizo omwe akufotokozedwa m'bukuli.

Table 1. Kutsata ndondomeko yothamanga

Olink® Explore Reagent Kit	Chiwerengero cha Olink Library	Chiwerengero cha ma cell oyenda ndi kuthamanga (ma)
Olink® Explore 384 Reagent Kit	1	1
Olink® Onani 4 x 384 Reagent Kit	4	4
Olink® Explore 1536 Reagent Kit	4	4
Olink® Explore Expansion Reagent Kit	4	4
Olink® Explore 3072 Reagent Kit	8	8

Ikani Chinsinsi cha Olink®

Sungani Chinsinsi cha Olink xml-file Olink_NSQ2K_P2_V1 mu chikwatu choyenera.

ZINDIKIRANI: Chinsinsi chachizolowezi cha Olink chidzangogwira ntchito ndi NextSeq™ 1000/2000 P2 Reagents (100 Cycles) v3 kit ndi NextSeq™ 1000/2000 control software v1.2 kapena v1.4.

Konzani kutsata ma reagents

Pa sitepe iyi, katiriji reagent munali clustering ndi sequencing reagents ndi thawed ndi otaya selo ndi kukonzekera.

CHENJEZO: Katiriji ya reagent ili ndi mankhwala omwe angakhale owopsa. Valani zida zodzitchinjiriza zokwanira ndikutaya ma reagents ogwiritsidwa ntchito molingana ndi miyezo yoyenera. Kuti mudziwe zambiri, onani Illumina NextSeq 1000 ndi 2000 System Guide (chikalata #1000000109376).

Konzani katiriji reagent

Kukhetsa katiriji yosatsegulidwa kungatheke pogwiritsa ntchito njira zitatu: kutentha kwa chipinda, m'madzi osambira, kapena mufiriji.

Konzani benchi

1x NextSeq™ 1000/2000 P2 Reagent Cartridge (mizungu 100)

Malangizo

Thaw the reagent cartridge monga tafotokozera mu Table 2.

Table 2. Reagent katiriji thawing njira

Njira yothirira	Malangizo
Pa kutentha kwa chipinda	Popanda kutsegula thumba lazojambula zasiliva, ikani katiriji yowundana pa benchi ndikuyisungunula kutentha kwa maola 9. Musapitirire maola 16. Onetsetsani kuti cholembera chachikwama chayang'ana m'mwamba ndipo mpweya ukhoza kuzungulira katiriji.
M'madzi osamba	Popanda kutsegula thumba lazojambula zasiliva, ikani katiriji yowundana yomizidwa ndi theka m'madzi osambira a 25 °C ndikusiya kuti isungunuke kwa maola 6. Musapitirire maola 8. Onetsetsani kuti cholembera chachikwama chayang'ana m'mwamba komanso kuti katiriji simatembenuzika panthawi yosungunuka. Yanikani katiriji bwinobwino ndi thaulo la pepala.
Mufiriji	Popanda kutsegula thumba lazojambula zasiliva, ikani katiriji pa benchi ndikusungunula kutentha kwa maola 6. Onetsetsani kuti cholembera chachikwama chayang'ana m'mwamba ndipo mpweya ukhoza kuzungulira katiriji. Malizitsani kusungunula katiriji mufiriji kwa maola 12. Musapitirire maola 72. Musanayambe kuthamanga, chotsani katiriji yosatsegulidwa mufiriji ndikuyilola kuti ifike kutentha kwa mphindi 15, koma osapitirira ola limodzi.

ZINDIKIRANI: Makatiriji osungunuka sangaumitsidwenso ndipo amayenera kusungidwa pa 4 ° C, kwa nthawi yayitali ya maola 72.

Konzani otaya selo

Konzani benchi

1x NextSeq™ 1000/2000 P2 Flow Cell

Malangizo

Bweretsani firiji otaya selo kutentha firiji kwa mphindi 10-15.

Konzani Library ya Olink® kuti musanthule

Panthawi imeneyi, Olink Library yoyeretsedwa komanso yoyendetsedwa bwino imatsitsidwa mpaka kumapeto komaliza. Dziwani kuti kutanthauzira kwa Library kumangochitika pachombocho.

Konzani benchi

Lib Tube, yokonzedwa molingana ndi Olink Explore User Manual

1x RSB yokhala ndi Tween 20
MilliQ madzi
2x machubu a Microcentrifuge (1.5 mL)
Pipette pamanja (10, 100 ndi 1000 μL)
Zosefera za pipette

Musanayambe

Thaw Lib Tube ngati yachisanu.
Thirani RSB yozizira ndi Tween 20 kutentha kwa mphindi 10. Sungani pa +4 ° C mpaka mutagwiritsidwa ntchito.
Chongani machubu awiri atsopano a 1.5 mL microcentrifuge motere:
- Lembani chubu limodzi "Dil" (kwa Library yochepetsedwa ya 1:100)
- Lembani chubu limodzi "Seq" (kuti mukonzekere kutsitsa Library)

Malangizo

Onjezani 495 μL ya madzi a MilliQ ku Dil Tube.

Vortex Lib Tube ndikuyizungulira mwachidule.
Sinthani pamanja 5 μL kuchokera ku Lib Tube kupita ku Dil Tube.
Vortex the Dil Tube ndikuyizungulira pang'ono.
Onjezani 20 μL ya RBS yokhala ndi Tween 20 ku Seq Tube.

Sinthani pamanja 20 μL kuchokera ku Dil Tube kupita ku Seq Tube.
Vortex Seq Tube ndikuyizungulira mwachidule.
Nthawi yomweyo pitilizani ku 2.5 Load flow cell ndi Olink® Library mu cartridge ya reagent.

ZINDIKIRANI: Sungani ma Lib Tube pa -20 °C ngati atha kubwerezanso.

Kwezani cell flow and Library ya Olink® mu cartridge ya reagent

Pa sitepe iyi, selo yothamanga ndi Library ya Olink yochepetsedwa imayikidwa mu cartridge ya reagent thawed.

Konzani benchi

1x thawed NextSeq™ 1000/2000 P2 Reagent Cartridge (100 cycle), yokonzedwa mu sitepe yapita
1x NextSeq™ 1000/2000 P2 Flow Cell, yokonzedwa kale

Seq Tube (yokonzeka kutsitsa Library ya Olink), yokonzedwa kale
Pipette yapamanja (100 μL)
Pipette nsonga (1 mL)

Konzani katiriji

Chotsani katiriji ku thumba lazojambula zasiliva.
Pindutsani katiriji kakhumi kuti musakanize bwino ma reagents osungunuka mkati.

ZINDIKIRANI: Si zachilendo kumva zigawo zamkati zikugwedezeka.

Kwezani cell cell mu katiriji

Mukakonzeka kukweza selo lotaya mu katiriji, chotsani selo lotaya kuchokera phukusi. Gwirani cell yotaya ndi tabu yotuwa, ndikulemba pa tabu yoyang'ana m'mwamba. Gwiritsani ntchito magolovesi opanda ufa atsopano kuti mupewe kuwononga galasi pamwamba pa cell yotaya.
Ikani otaya selo mu otaya selo kagawo kutsogolo kwa katiriji. Kudina komveka kukuwonetsa kuti cell yotaya imayikidwa bwino.
Chotsani tabu imvi poyikoka.

Kwezani Library ya Olink® mu katiriji

kuboola mosungiramo Library ndi nsonga yoyera ya 1 mL ya pipette.
Kwezani 20 μL ya Library ya Olink kuchokera ku Seq Tube mpaka pansi pankhokwe ya library.

Pangani sequencing run ya Olink®

Pa sitepe iyi, Buffer cartridge yokhala ndi cell flow flow and Olink Library imakwezedwa mu NextSeq™ 2000, ndipo kutsatizana kumayambika pogwiritsa ntchito Chinsinsi cha Olink.

Konzani benchi

1x NextSeq™ 1000/2000 P2 Reagent Cartridge (mizungu 100) yodzaza ndi NextSeq™ 1000/2000 P2 Flow Cell ndi Library ya Olink yochepetsedwa, yokonzedwa mu gawo lapitalo.

Konzani Run Mode

Kuchokera pa menyu yowongolera mapulogalamu, sankhani Zikhazikiko.
Pansi pa BaseSpace Sequence Hub Services & Proactive Support, sankhani Local Run Setup.
Sankhani Proactive Support Pokhapokha ngati zokonda zina. Izi zimafuna intaneti.
Sankhani Malo Osungira deta yanu. Malo Osungira ayenera kukhala pafupi kapena pafupi ndi dera lanu.
Khazikitsani malo a Output Folder kuti mupeze deta yaposachedwa. Sankhani Sankhani kuyenda ndi kusankha linanena bungwe chikwatu.
Sankhani bokosi loyang'ana la Denature ndi Dilute On Board kuti musinthe ndikusintha Library yomwe ili m'chidacho.
Sankhani bokosi loyang'ana pa Purge Reagent Cartridge kuti muchotseretu ma reagents osagwiritsidwa ntchito kugawo la cartridge lomwe lagwiritsidwa ntchito.
Sankhani bokosi la Autocheck la zosintha zamapulogalamu kuti mungoyang'ana zosintha zamapulogalamu (posankha). Izi zimafuna intaneti.
Sankhani Sungani.

Khazikitsani magawo a run

ZINDIKIRANI: Malangizowa amagwira ntchito ku mtundu 1.4 wa pulogalamu yowongolera ya NextSeq™ 1000/2000. Zina mwazomwe tafotokozazi zitha kukhala zosiyana mukamagwiritsa ntchito v1.2

Kuchokera kuwongolera mapulogalamu menyu, kusankha Start.
Sankhani Pamanja Khazikitsani New Run ndikusindikiza Setup.
Patsamba la Run Setup, khazikitsani magawo othamanga motere:
- Pagawo la Run Name, lowetsani ID yoyesera yapadera.
- Pamndandanda wotsikirapo wa Read Type, sankhani njira ya Single Read.
- Lowetsani kuchuluka kwa mizungulira motere:
  - Werengani 1: 24
  - Mlozera 1: 0
  - Mlozera 2: 0
  - Werengani 2: 0

ZINDIKIRANI: Ndikofunikira kuti Read 1 ikhazikitsidwe ku 24, apo ayi kuthamanga konseko kulephera. Musanyalanyaze mauthenga ochenjeza pamene mukulowetsa chiwerengero cha maulendo.

Pamndandanda wotsikira pansi wa Custom Primer Wells, sankhani Ayi.
M'munda Chinsinsi (mwazosankha), sankhani Sankhani kuti muyende ndikusankha XML yachizolowezi file Olink_NSQ2K_P2_V1. Sankhani Open.
Osatumiza kunja kwa Sampndi Mapepala.
Onetsetsani kuti malo a Output Folder ndi olondola. Apo ayi, kusankha Sankhani kuyenda ndi kusankha ankafuna linanena bungwe chikwatu malo.

M'munda wa Denature ndi Dilute Onboard, sankhani Yathandizidwa kuchokera pamndandanda wotsitsa.
Sankhani Kukonzekera.

Kwezani cartridge yodzaza

Sankhani Katundu. Visor ya chida imatsegulidwa ndipo thireyi imatulutsidwa.
Ikani katiriji yodzaza pa thireyi ndi chizindikiro choyang'ana m'mwamba ndi cell cell mkati mwa chida.

Sankhani Tsekani.
Katiriji ikadzazidwa bwino, tsimikizirani zoyendetsa ndikusankha Sequence. Chidacho chimapanga cheke chisanachitike cha chidacho ndi ma fluidic.
- ZINDIKIRANI: Pakuwunika kwa fluidics, zimayembekezereka kumva phokoso lambiri.
Onetsetsani kuti kuthamanga kuyambika pambuyo poti cheke chodziwikiratu chatha (~ mphindi 15). Nthawi yotsatizana ndi pafupifupi 10h30 min.
- ZINDIKIRANI: Pazolephera zilizonse zomwe zisanachitike, onani malangizo a wopanga. Samalani kuti musakumane kapena kusokoneza NextSeq™ 2000 panthawi yotsatizana. Chidacho chimakhudzidwa ndi kugwedezeka.
Yeretsani malo ogwirira ntchito.

Yang'anirani Kupititsa patsogolo kwa Run

Olink amagwiritsa ntchito NGS monga kuwerenga kuti athe kuwerengera kuchuluka kwa mndandanda wodziwika bwino kuti athe kuyerekeza kuchuluka kwa mapuloteni operekedwa mu s.amples (zogwirizana ndi sampizi). Ubwino wa data kuchokera pamayendedwe aliwonse a Explore amatsatiridwa makamaka ndi magawo a QC apadera kuukadaulo wa Olink. Chifukwa chake, ma metric owongolera omwe amagwiritsidwa ntchito mu NGS wamba, monga Q-score, ndiofunikira kwambiri.

Chotsani ndi kutaya katiriji mukatha kuthamanga

CHENJEZO: Ma reagents awa ali ndi mankhwala omwe angakhale oopsa. Valani zida zodzitchinjiriza zokwanira ndikutaya ma reagents ogwiritsidwa ntchito molingana ndi miyezo yoyenera. Kuti mudziwe zambiri, onani Illumina NextSeq 1000 ndi 2000 System Guide (chikalata #1000000109376).

Kuthamanga kukatha, sankhani Eject Cartridge.
- ZINDIKIRANI: Katiriji yogwiritsidwa ntchito kuphatikiza cell cell imatha kusiyidwa m'malo mpaka kuthamanga kwina, koma osapitilira masiku atatu.
Chotsani katiriji mu thireyi.
Tayani ma reagents molingana ndi miyezo yoyenera.
Sankhani Close Door. Thireyiyo yalowetsedwanso.
Sankhani Kunyumba kuti mubwerere ku Sikirini yakunyumba.
- ZINDIKIRANI: Popeza katiriji ili ndi njira zonse zoyendetsera dongosololi, komanso posungira kuti asonkhanitse ma reagents ogwiritsidwa ntchito, palibe chifukwa chotsuka zida pambuyo pothamanga.

Mbiri yobwereza

Baibulo	Tsiku	Kufotokozera
1.0	2021-12-01	Chatsopano

www.olink.com

Kugwiritsa Ntchito Kafukufuku Pokha. Osagwiritsidwa Ntchito mu Njira Zowunika.

Izi zikuphatikiza chilolezo chogwiritsa ntchito osagulitsa zinthu za Olink. Ogwiritsa ntchito malonda angafunike zilolezo zowonjezera. Chonde lemberani Olink Proteomics AB kuti mumve zambiri. Palibe zitsimikizo, zofotokozedwa kapena kutanthauza, zomwe zimapitilira kufotokozera uku. Olink Proteomics AB siyiyenera kuwononga katundu, kuvulaza munthu, kapena kuwonongeka kwachuma chifukwa cha mankhwalawa. Chizindikiro chotsatirachi ndi cha Olink Proteomics AB: Olink®. Izi zimaphimbidwa ndi ma Patent angapo ndi mapulogalamu a patent omwe amapezeka https://www.olink.com/patents/.

Olink Proteomics, Dag Hammarskjölds väg 52B , SE-752 37 Uppsala, Sweden

1193, v1.0, 2021-12-01

Zolemba / Zothandizira

Olink NextSeq 2000 Onani Njira Yotsatirira [pdf] Buku Logwiritsa Ntchito
NextSeq 2000, Onani Njira Yotsatirira, NextSeq 2000 Onani Njira Yotsatirira

Maumboni

Buku Logwiritsa Ntchito

Olink NextSeq 2000 Onani Njira Yotsatirira

Zolemba

Mawu Oyamba

Othandizira ukadaulo

Malangizo a Laboratory

Konzani kutsata ma reagents

Konzani katiriji reagent

Konzani otaya selo

Konzani Library ya Olink® kuti musanthule

Kwezani cell flow and Library ya Olink® mu cartridge ya reagent

Kwezani cell cell mu katiriji

Pangani sequencing run ya Olink®

Khazikitsani magawo a run

Chotsani ndi kutaya katiriji mukatha kuthamanga

Mbiri yobwereza

Zolemba / Zothandizira

Maumboni

Siyani ndemanga

Letsani yankho