Olink Tshawb Nrhiav Sequencing siv NextSeq 550

Taw qhia

Npaj siv
Olink® Explore yog multiplex immunoassay platform rau human protein biomarker discovery. Cov khoom no yog npaj rau Kev Tshawb Fawb Siv nkaus xwb, thiab tsis yog siv rau hauv cov txheej txheem kuaj mob. Lub chaw kuaj mob tsuas yog ua haujlwm los ntawm cov neeg ua haujlwm hauv chaw kuaj mob. Kev ua cov ntaub ntawv tsuas yog ua los ntawm cov neeg ua haujlwm raug cob qhia. Cov txiaj ntsig tau txhais tau tias yog siv los ntawm cov kws tshawb fawb ua ke nrog rau lwm cov kev tshawb fawb soj ntsuam lossis kuaj pom.

Hais txog phau ntawv no
Phau Ntawv Qhia Tus Neeg Siv no muab cov lus qhia uas yuav tsum tau ua raws li Olink® Tshawb Cov Tsev Qiv Ntawv ntawm Illumina® NextSeq™ 550. Cov lus qhia yuav tsum ua raws li nruj me ntsis thiab meej. Ib qho kev sib txawv thoob plaws hauv chav kuaj yuav ua rau cov ntaub ntawv tsis raug. Ua ntej pib ua haujlwm hauv chav kuaj, sab laj Olink® Tshawb Nrhiav Tshajview Cov neeg siv phau ntawv qhia txog kev taw qhia rau lub platform, suav nrog cov ntaub ntawv hais txog cov tshuaj reagents, cov khoom siv thiab cov ntaub ntawv xav tau, dhau musview ntawm kev ua haujlwm, nrog rau cov txheej txheem kuaj. Rau cov lus qhia txog yuav ua li cas khiav Olink® Tshawb Nrhiav Cov Khoom Siv Tshuaj, saib rau Olink® Tshawb Xyuas Cov Neeg Siv Khoom Siv. Rau kev ua cov ntaub ntawv thiab kev tshuaj xyuas ntawm Olink® Tshawb nrhiav cov txiaj ntsig, xa mus rau Olink® MyData Cloud User Guide. Tag nrho cov cim kev lag luam thiab cov ntawv pov thawj muaj nyob hauv cov khoom no yog cov cuab yeej ntawm Olink® Proteomics AB, tshwj tsis yog hais tias lwm yam.

Kev txhawb nqa
Rau kev txhawb nqa kev, hu rau Olink Proteomics ntawm support@olink.com.

Laboratory lus qhia

Tshooj lus no muab cov lus qhia txog yuav ua li cas thiaj li Olink Libraries ntawm NextSeq ™ 550 siv NextSeq™ 500/550 High Output Kit v2.5 (75 Cycles). Cov txheej txheem siv los ua ke yog kev hloov kho ntawm Illumina® tus qauv NGS kev ua haujlwm rau Illumina® NextSeq™ 550. Ua ntej yuav mus ua ntu zus, xyuas kom meej tias qhov zoo ntawm cov tsev qiv ntawv purified tau raug txheeb xyuas. Xa mus rau Olink Explore User Manual rau cov lus qhia txog kev tswj kom zoo.

Npaj qhov kev sib tw khiav
Ib lub tsev qiv ntawv Olink tuaj yeem ua raws li NextSeq ™ 550 High Output flow cell thiab ib qho khiav. Tus naj npawb ntawm High Output flow cells thiab khiav yuav tsum tau ua raws li cov khoom siv sib txawv ntawm Olink Explore Reagent tau piav qhia hauv Table 1. Yog tias xav tau ntau tshaj ib qho khiav, rov ua cov lus qhia hauv phau ntawv no.

Table 1.Sequencing run Planning:

Olink® Tshawb Nrhiav Cov Khoom Siv Tshuaj	Tus naj npawb ntawm Olink Libraries	Number of flow cell(s) thiab khiav(s)
Olink® Tshawb 384 Reagent Kit	1	1
Olink® Tshawb 4 x 384 Reagent Kit	4	4
Olink® Tshawb 1536 Reagent Kit	4	4
Olink® Tshawb Expansion Reagent Kit	4	4
Olink® Tshawb 3072 Reagent Kit	8	8

Nruab Olink® daim ntawv qhia kev cai
Thaum lub sijhawm no, Olink® daim ntawv qhia kev cai raug teeb tsa rau ntawm NextSeq ™ 550. Cov kauj ruam no tsuas yog yuav tsum tau ua ib zaug, ua ntej Olink sequencing khiav yog thawj zaug.
CEEB TOOM: Daim ntawv qhia kev cai Olink tsuas yog ua haujlwm nrog NextSeq ™ 500/550 Cov Khoom Siv Siab Tshaj Tawm thiab NextSeq™ Control Software 4.0

1. Unzip thiab tso Olink daim ntawv qhia kev cai Olink_NSQ550_HighOutput_V1 hauv daim nplaub tshev nram qab no ntawm NextSeq™ 550 ntsuas: C:\Program Files\Illumina\NextSeq Control Software\Recipe\Custom\High\.
2. Nyob rau hauv System Customization> Tswj Cov Cuab Yeej, qhib cov ntawv qhia kev cai. Yog tias tsis xaiv, qhov kev xaiv daim ntawv qhia kev cai yuav tsis tshwm sim thaum lub teeb tsa.

CEEB TOOM: Hauv NCS 4.0 software version, qhov kev xaiv xaiv ib daim ntawv qhia kev cai tsuas yog tshwm sim tom qab lub reagent cartridge tau thauj khoom, tsis yog nyob rau hauv nplooj ntawv teeb tsa ua ntej.
CEEB TOOM: Kev khiav yuav tsum tau teem rau hauv phau ntawv hom kom tso cai rau cov zaub mov txawv.

Npaj sequencing reagents

Nyob rau hauv cov kauj ruam no, lub reagent cartridge uas muaj pawg thiab sequencing reagents yog thawed thiab txaus cell npaj.

Npaj reagent cartridge
CEEB TOOM: Lub reagent cartridge muaj cov tshuaj uas tsim kev puas tsuaj. Hnav cov cuab yeej tiv thaiv tsim nyog thiab muab pov tseg cov tshuaj reagents raws li cov qauv siv tau. Yog xav paub ntxiv, xa mus rau Illumina NextSeq 550 Txoj Kev Qhia (document #15069765).

Npaj lub rooj zaum

• 1x NextSeq™ 500/550 High Output Reagent Cartridge v2 (75 cycles).

Cov lus qhia

1. Muab cov reagent frozen cartridge ib nrab-submerged rau hauv chav dej tempered thiab cia nws thaw rau 1 teev. Xyuas kom tseeb tias tag nrho cov reagent reservoirs ntawm cov cartridges yog thawed kiag li.
   CEEB TOOM: Txhawm rau kom yooj yim, thaw lub cartridge hnub ua ntej thiab khaws cia rau hmo ntuj ntawm 4 ° C. Ntawm qhov ntsuas kub no, cov tshuaj reagents ruaj khov mus txog ib lub lis piam.
2. Qhuav lub hauv paus cartridge kom huv si nrog ib daim ntaub so ntswg thiab blot lub foil seals kom qhuav nrog cov ntaub so ntswg tsis muaj lint yog xav tau.
3. Hloov lub cartridge kaum zaug kom sib tov cov thawed reagents hauv.
4. Coj mus rhaub lub cartridge maj mam rau ntawm lub rooj zaum kom tshem tawm cov npuas. Khaws lub cartridge ntawm chav tsev kub yog tias nws yuav siv li ntawm 4 teev.

Npaj cov xovtooj ntawm tes

Npaj lub rooj zaum

• 1x NextSeq ™ 500/550 High Output Flow Cell v2.5.

Cov lus qhia

1.  Nqa lub tub yees txias mus rau chav sov li 30 feeb.
2.  Muab cov hnab looj tes tshiab tshiab tsis pub dawb (kom tsis txhob muaj kab mob hauv lub khob ntawm cov dej ntws).
3.  Thaum npaj txhij los thauj cov ndlwg ntawm tes mus rau hauv lub ntsuas, tshem tawm cov dej ntws ntawm lub pob thiab cov yas clamshell.
4.  Tshawb xyuas lub flow cell. Yog hais tias muaj feem los yog plua plav pom ntawm ib qho ntawm cov iav qhov chaw, ntxuav qhov chaw siv nrog lint-dawb isopropyl cawv so thiab qhuav nws nrog cov ntaub so ntswg qis.

Npaj Olink® Cov Tsev Qiv Ntawv rau kev ua ntu zus

Thaum lub sij hawm cov kauj ruam no, NaOH thiab Tris-HCl dilutions yog npaj, thiab cov purified thiab zoo tswj Olink Library yog diluted thiab denatured nyob rau hauv sequential cov kauj ruam.

Npaj NaOH dilution
Lub NaOH dilution yog siv los qhia cov Tsev Qiv Ntawv.

Npaj lub rooj zaum

• 1 NPO NPO
• MilliQ dej
• 1 x Microcentrifuge tube (1.5 mL)
• Phau ntawv pipette (10-100 μL)
• Lim pipette tswv yim

Ua ntej koj pib
Kos lub microcentrifuge raj "NaOH".

Cov lus qhia

1. Npaj 0.2 N NaOH dilution hauv NaOH Tube raws li Table 2.
2. Vortex lub NaOH Tube kom huv si thiab tig mus. Siv hauv 12 teev.

Table 2. 0.2 N NaOH dilution

Reagent	Ntim (μL)
MilliQ dej	80
1 NPO NPO	20

2.4.2 Npaj Tris-HCl dilution
Tris-HCl dilution yog siv los nruab nrab lub tsev qiv ntawv denaturated.

Npaj lub rooj zaum

• 1 M Tris-HCl pH 7.0 Tshuag (Trizma® hydrochloride tov)
• MilliQ dej
• 1 x Microcentrifuge tube (1.5 mL)
• Phau ntawv pipette (10-100 μL)
• Lim pipette tswv yim

Ua ntej koj pib

• Kos lub microcentrifuge raj "Tris-HCl"

Cov lus qhia

1. Npaj 200 mM Tris-HCl dilution hauv Tris-HCl Tube raws li Table 3.
2. Vortex lub Tris-HCl Tube kom huv si thiab tig nws.

Table 3. 200 mM Tris-HCl dilution:

Reagent	Ntim (μL)
MilliQ dej	80
1M Tris-HCl pH 7.0 Tshuag (Trizma® hydrochloride tov)	20

Dilute Olink® Cov Tsev Qiv Ntawv
Thaum lub sij hawm no, lub purified thiab zoo tswj Olink Library yog diluted 1:33.

Npaj lub rooj zaum

• Lib Tube, npaj raws li siv tau Olink Tshawb Xyuas Cov Neeg Siv Khoom
• MilliQ dej
• 1 x Microcentrifuge tube (1.5 mL)
• Los ntawm txhais tes pipettes (0.5-10 thiab 100-1000 μL)
• Lim pipette tswv yim

Ua ntej koj pib

• Thaw lub Lib Tube yog khov.
• Kos lub microcentrifuge raj tshiab: "Dil".

Cov lus qhia

1. Ntxiv 96 μL ntawm MilliQ dej rau Dil Tube.
2. Vortex lub Lib Tube thiab tig nws luv luv.
3. Hloov 3 μL los ntawm Lib Tube mus rau Dil Tube.
4. Vortex lub Dil Tube thiab tig nws luv luv

Denature thiab dilute Olink® Library mus rau qhov kawg loading concentration
Thaum lub sij hawm no, lub diluted Olink Library yog denatured thiab ntxiv diluted mus rau qhov kawg loading concentration.

Npaj lub rooj zaum

• Dil Tube, npaj hauv cov kauj ruam dhau los
• 0.2 N NaOH dilution, freshly npaj nyob rau hauv cov kauj ruam dhau los
• 200 mM Tris-HCl (pH 7.0) dilution, npaj hauv kauj ruam dhau los
• Hybridization buffer 1 (HT1) suav nrog hauv NextSeq™ Accessory Box v2
• 2 x Microcentrifuge tubes (1.5 mL thiab 2 mL)
• Los ntawm txhais tes pipettes (0.5-10 thiab 100-1000 μL)
• Lim pipette tswv yim

Ua ntej koj pib

• Thaw lub khov HT1 tsis nyob ntawm chav tsev kub. Khaws ntawm +4 ° C kom txog thaum siv.
• Kos lub tshiab 1.5 mL microcentrifuge raj: "Den" (rau lub tsev qiv ntawv denaturated).
• Kos tus tshiab 2 mL microcentrifuge raj: "Seq" (rau cov npaj txhij mus thauj cov tsev qiv ntawv).

Cov lus qhia

1. Hloov 5 μL los ntawm Dil Tube mus rau Den Tube.
2. Ntxiv 5 μL ntawm 0.2 N NaOH rau Den Tube.
3. Vortex lub Den Tube thiab tig nws luv luv.
4. Incubate lub Den Tube rau 5 feeb ntawm chav tsev kub kom denature lub Library.
5. Ntxiv 5 μL ntawm 200 mM Tris-HCl (pH 7.0) rau Den Tube kom tsis txhob muaj cov tshuaj tiv thaiv.
6. Vortex lub Den Tube thiab tig nws luv luv.
7. Ntxiv 985 μL ntawm prechilled HT1 rau Den Tube.
8. Vortex lub Den Tube thiab tig nws luv luv. Lub raj tuaj yeem khaws cia ntawm +4 ° C kom txog thaum siv (ib hnub).
9. Hloov 205 μL los ntawm Den Tube mus rau Seq Tube.
10. Ntxiv 1095 μL ntawm prechilled HT1 rau Seq Tube.
11. Hloov lub Seq Tube kom sib tov cov reagents thiab tig nws luv luv. Qhov kawg loading ntim yog 1.3 mL.
12. Tam sim ntawd txuas ntxiv mus rau 2.5 Ua Olink® ua ntu zus.

Ua haujlwm Olink® sequencing

Thaum lub sij hawm cov kauj ruam no, cov ntaub ntawv tsis siv neeg, cov dej ntws ntawm tes thiab cov ntaub ntawv npaj reagent uas muaj Olink Library tau thauj khoom rau hauv NextSeq 550, thiab kev sib tw khiav yog pib siv Olink daim ntawv qhia kev cai.

Npaj lub rooj zaum

• Seq Tube (nrog npaj txhij mus thauj khoom Library), npaj nyob rau hauv cov kauj ruam dhau los
• 1x NextSeq ™ 500/550 High Output Reagent Cartridge v2, npaj hauv kauj ruam dhau los
• 1x NextSeq ™ 500/550 High Output Flow Cell v2.5, npaj hauv kauj ruam dhau los
• 1x NextSeq™ 500/550 Buffer Cartridge v2 (75 cycles), ntawm chav tsev kub

Teeb tsa kev ua haujlwm sib luag
Thaum lub sijhawm cov kauj ruam no, cov kev ua haujlwm sib luag tau raug xaiv ntawm NextSeq ™ 550.

1. Ntawm NextSeq™ 550 Home screen, xaiv Kev sim.
2. Nyob rau ntawm Xaiv Kev Ntsuam Xyuas, xaiv Sequence.
3. Hauv Khiav Teeb nplooj ntawv, xaiv Manual khiav hom thiab tom qab ntawd Next.
4. Teeb tsa cov kev khiav haujlwm raws li hauv qab no:
   • Hauv Khiav Lub Npe, nkag mus rau qhov kev sim ID tshwj xeeb.
   • Nyob rau hauv lub tsev qiv ntawv ID teb, sau tus ID ntawm lub tsev qiv ntawv koj tab tom khiav (yeem).
   • Hauv Kev Nyeem Hom teb, xaiv qhov Kev Xaiv Ib Leeg Nyeem.
   • Sau tus lej ntawm lub voj voog raws li hauv qab no:
     • Nyeem 1:24
     • Index 1: 0
     • Index 2: 0
     • Nyeem 2:0
   • Khaws lub checkbox rau kev cai primers unselected.
   • Teem lub tso zis nplaub tshev qhov chaw rau cov ntaub ntawv nyoos khiav tam sim no. Xaiv Browse los hloov cov zis nplaub tshev qhov chaw.
5. Tsis txhob teeb tsa Sample Sheet.
6. Xaiv Purge consumables rau no khiav.
7. Xaiv Tom ntej.

Load flow cell mus rau NextSeq™ 550

1. Tshem tawm cov khoom siv ntws los ntawm kev khiav dhau los.
2. Muab qhov tshiab npaj ntws ntawm tes rau ntawm stage.
3. Xaiv Load. Lub qhov rooj kaw cia li kaw.
4. Thaum lub xov tooj ntawm tes ID tshwm rau ntawm qhov screen thiab cov sensors raug kuaj xyuas ntsuab, xaiv Tom ntej

Tshem lub thawv reagent
CEEB TOOM: Cov txheej txheem reagents no muaj cov tshuaj muaj txiaj ntsig zoo. Hnav cov cuab yeej tiv thaiv tsim nyog thiab muab pov tseg cov tshuaj reagents raws li cov qauv siv tau. Yog xav paub ntxiv, xa mus rau Illumina NextSeq 550 System Guide.

1. Qhib lub qhov rooj ntawm qhov tsis sib xws, tshem tawm cov khoom siv reagents los ntawm qhov chaw qis, thiab pov tseg cov ntsiab lus raws li cov qauv siv.
2. Xaub lub thawv ntim reagent rov qab rau hauv qhov chaw tsis muaj qhov qis. Lub suab nyem qhia tau hais tias lub thawv ntim tau raug tso.

Load buffer cartridge

1. Tshem tawm cov khoom siv tsis siv los ntawm cov khoom siv sab sauv thiab pov tseg cov ntsiab lus raws li cov qauv siv tau.
2. Xaub ib daim cartridge tshiab rau hauv qhov tsis zoo sab saud. Lub suab nyem qhia tau hais tias lub cartridge yog raug muab tso rau. Nco ntsoov tias daim npav ID tsis tshwm nyob rau ntawm qhov screen thiab cov sensors raug kuaj xyuas ntsuab.
3. Kaw lub qhov rooj ntawm qhov tsis sib xws thiab xaiv Next.

Load reagent cartridge

1. Qhib lub qhov rooj ntawm lub reagent compartment, tshem tawm cov ntaub ntawv reagent siv, thiab pov tseg cov ntsiab lus tsis siv raws li cov qauv siv. Lub reservoir ntawm txoj hauj lwm 6 yog tshem tau kom yooj yim pov tseg.
2. Pierce lub foob ntawm lub reservoir #10 sau tias "Load Library Here" nrog ib tug huv 1 mL pipette taub hau.
3. Load 1.3 mL lub Olink Library los ntawm Seq Tube rau hauv reservoir #10 sau tias "Load Library Here".
4. Xaub lub reagent cartridge tshiab rau hauv lub reagent compartment thiab kaw lub qhov rooj reagent compartment.
5. Xaiv Load thiab tos rau ~ 30 vib nas this kom txog thaum lub reagent cartridge ID tshwm ntawm qhov screen thiab cov sensors raug kuaj ntsuab.
6. Los ntawm daim ntawv qhia zaub mov nco-down, xaiv [Kev Cai] "Olink_NSQ550_HighOutput_V1" xaiv daim ntawv qhia.
7. Xaiv Tom ntej.

Pib sequencing khiav

1. Paub meej tias qhov khiav tsis tau tso tawm rau ntawm Review npo. Txhawm rau hloov kho tej yam tsis muaj, nias Rov qab rov qab mus rau Run Setup screen.
2. Xaiv Tom ntej. Kev khiav pib tom qab kev kuaj xyuas ua ntej tsis siv neeg. Lub sij hawm ua ntu zus yog kwv yees li 7h30 min.
3. Ntxuav thaj chaw ua haujlwm.

CEEB TOOM: Thaum qhov kev sib txuas ua tiav tiav, lub software pib ua haujlwm tsis siv neeg tom qab kev ntxuav siv cov kev daws teeb meem uas tau muab rau hauv daim cartridge tsis thiab NaOCl muab rau hauv daim cartridge reagent. Qhov kev ntxuav no yuav siv li 90 feeb. Lub khawm hauv tsev pib ua haujlwm thaum ntxuav tiav. Siv cov cartridges thiab ntws ntawm tes tuaj yeem tso rau hauv qhov chaw kom txog rau thaum lub sijhawm ua haujlwm tom ntej.

Saib xyuas kev khiav haujlwm
Olink siv NGS raws li kev nyeem ntawv txhawm rau txheeb xyuas tus lej ntawm qhov paub ua ntu zus txhawm rau kwv yees qhov concentration ntawm cov protein muab hauv samples (txog rau lwm tus samples). Cov ntaub ntawv zoo los ntawm txhua qhov Kev Tshawb Fawb ua raws li kev khiav haujlwm yog txiav txim siab los ntawm QC qhov tshwj xeeb rau Olink thev naus laus zis. Yog li ntawd, cov qauv kev tswj xyuas kev ntsuas zoo siv hauv cov qauv NGS, xws li Q-score, tsis tshua muaj qhov tseem ceeb.

Kev kho keeb kwm

Version	Hnub tim	Kev piav qhia
1.1	2021-12-13 Ib	Kev hloov pauv hloov
1.0	2021-12-01 Ib	Tshiab

www.olink.com
Rau kev tshawb fawb siv nkaus xwb. Tsis siv rau hauv cov txheej txheem kuaj mob.
Cov khoom no suav nrog daim ntawv tso cai rau kev siv tsis yog lag luam ntawm Olink cov khoom. Cov neeg siv khoom lag luam yuav xav tau daim ntawv tso cai ntxiv. Thov hu rau Olink
Proteomics AB. Tsis muaj kev lav phib xaub, qhia lossis qhia, uas txuas ntxiv dhau qhov kev piav qhia no. Olink Proteomics AB tsis yog lub luag haujlwm rau cov khoom puas tsuaj, kev raug mob ntawm tus kheej, lossis kev poob nyiaj txiag los ntawm cov khoom no.
Cov cim lag luam hauv qab no yog Olink Proteomics AB: Olink®.
Cov khoom no yog them los ntawm ob peb patents thiab daim ntawv thov patent muaj nyob ntawm https://www.olink.com/patents/.
© Copyright 2021 Olink Proteomics AB. Tag nrho cov khoom lag luam thib peb yog cov cuab yeej ntawm lawv cov tswv. Olink Proteomics, Dag Hammarskjölds väg 52B, SE-752 37 Uppsala, Sweden
1192, v1.1, 2021-12-13

Cov ntaub ntawv / Cov ntaub ntawv

Olink Tshawb Nrhiav Sequencing siv NextSeq 550 [ua pdf] Tus neeg siv phau ntawv Tshawb Sequence siv NextSeq 550, Tshawb Sequence, siv NextSeq 550, NextSeq 550

Cov ntaub ntawv

• Cov neeg siv phau ntawv