Opentrons OT-2 Workstations for Efficient Lab Automation Imiyalelo

Ishidi Lokulungiselela
Izilinganiso kanye Nokuhlaziywa Kwezibalo

Ibhalwe nguKennedy Bae, Ph.D. kanye noKinnari Watson, Ph.D.

Ukuqalisa

Ngaphambi kokufundisa uhlelo lwesifundo, qedela izinyathelo ezilandelayo ngaphambi kwekilasi.

Susa ibhokisi le-OT-2
Setha uhlelo lokusebenza lwe-Opentrons
Namathisela ama-pipettes
Linganisa idekhi
Linganisa ubude bethiphu & ne-pipette offset
Ngenisa noma imaphi amaphrothokholi ahlobene kuhlelo lokusebenza
Hlola sebenzisa umthetho olandelwayo ku-OT-2

Udinga Usekelo Olwengeziwe?

Ukuze uthole ukwesekwa kobuchwepheshe, sicela uhlole yethu Isikhungo Sosizo se-Opentrons ngezindatshana ezifanele. Uma udinga ukwesekwa okwengeziwe, sicela uxhumane support@opentrons.com.

Uma unemibuzo ehlobene nohlelo lwesifundo, sicela uthinte ababhali, uKennedy Bae, kokuthi kennedy@opentrons.com, noma uKinnari Watson, ku kinnari@opentrons.com.

Umhlahlandlela kathisha
Izilinganiso kanye Nokuhlaziywa Kwezibalo

Ibhalwe nguKennedy Bae, Ph.D. kanye noKinnari Watson, Ph.D.

Inhloso

Le lebhu ihlose ukuthuthukisa ukuqonda kwabafundi ngokusebenzisa ukuhlanjululwa kwe-serial ukuze kukhiqizwe ijika elijwayelekile, kanye nokusetshenziswa kwamajika ajwayelekile wokuhlanganisaample amanani. Lokhu kubandakanya ukubonga:

• Amakhono ayisisekelo wepayipi
• Ama-serial dilutions
• Ukulinganisa ukuphindaphinda
• Ububanzi bejika elijwayelekile
• Ukusetshenziswa kwezibalo ukuhlola ukubaluleka

Abafundi benza ipayipi yokufaka ngesandla ngokuhambisana nokubamba uketshezi okuzenzakalelayo, okuvumela abafundi ukuthi babone amandla namathuba azo zombili lezi zindlela.

Izilaleli zabafundi

Le lebhu yakhelwe ukusetshenziswa ekungeneni ezifundweni zebhayoloji zezinga elimaphakathi. Kuyavumelana nezimo ukwamukela noma iyiphi inombolo yabafundi ababhalise ekilasini.

Ulwazi Olungemuva

Abafundi kufanele babe nokuqonda komqondo wepayipi, i-spectrophotometry, amajika ajwayelekile, nokuhlaziywa kwezibalo.

Amakhono Abalulekile

Amakhono Elabhorethri
Ukufakwa kwamapayipi, sampukulungiswa kwe-le, kanye nokusetshenziswa kwemishini ye-automation yaselabhorethri

Ukuhlaziywa Kwedatha
Ukuthuthukisa amajika ajwayelekile, ukuhumusha, nokuhlaziywa kwezibalo

Ukucabanga okubucayi
Ukuhunyushwa kwedatha yokuhlola, ukuxazulula inkinga, nokudweba iziphetho eziphusile

Izimpahla

I-Opentrons Protocol

Landa umthetho olandelwayo kusuka
https://protocols.opentrons.com/protocol/customizable_s erial_dilution_ot2

• Khetha amapharamitha alandelayo ku- webikhasi ngokuya ngokusetha kwakho kanye nempahla:
  • Uhlobo lwe-Pipette
  • Mount Side
  • Uhlobo Lwethiphu
  • Uhlobo Lomkhombe
  • Uhlobo Lwepuleti
• Gcina izilungiselelo ezizenzakalelayo ze:
  • I-Dilution factor (3)
  • Inani lokuhlanjululwa (10)
  • Ivolumu yokuxuba isiyonke (150)
  • Akunalutho epuletini lomthombo (yebo)
  • Isu lokusebenzisa isu (sebenzisa ithiphu elilodwa)
  • Umthamo wegebe lomoya (10)

Izinsiza ze-Opentrons

I-Opentrons OT-2 Automated Liquid Handling Robot
I-Opentrons p300 8-channel pipette

Izisetshenziswa ezingezona ze-Opentrons

I-spectrophotometer esekwe epuletini
Isofthiwe yokuhlaziya idatha (isb. I-Excel noma isofthiwe yezibalo ezizinikele)
P1000 ivolumu eguquguqukayo yama-pipettes (eyodwa ngomfundi ngamunye)
I-pipette yevolumu eguquguqukayo ye-P100 (edingekayo eyodwa kuphela, ukuze kuboniswe Ingxenye A)

Ilebhu

I-Opentrons 300 μL I-Tip Rack
12 - umthombo
Ipuleti le-Flat Bottom engu-96
Amashubhu okuhlola (6 umfundi ngamunye)
I-Dropper (1 umfundi ngamunye)

Ama-reagents

1000 mL diH20 umfundi ngamunye
I-FD&C Blue No. 1 (McCormick® yasetshenziswa ekuthuthukisweni kwale lebhu)

Ubude besikhathi sokulinga

Amaseshini Ekilasi Adingekayo
1

Isikhathi Sokugijima Kwelebhu
Isikhathi esiphelele esilinganiselwe: amahora angu-2.5-3
Isingeniso kanye ne-Calibration Curve Generation: imizuzu engama-35
Sample Ukulungiselela nokufaka amapayipi: ihora elingu-1
Ukuqoqwa kwedatha: imizuzu engama-30

Ukuxazulula inkinga Okuyisisekelo

1. Yenza isilingo ngaphambi kwekilasi; ngale ndlela noma yiziphi izehlakalo ezingalindelekile zingaxazululwa ngaphambi kokuba abafundi bafike.
2. Izinkinga ezinamapuleti ahlabayo amathiphu cishe zibangelwa ukusebenzisa enye i-labware noma ukulinganisa irobhothi. Uma uhlangabezana nalokhu futhi uqinisekise ilebula elungile, zama ukulinganisa kabusha irobhothi.
3. Uma udinga ukufinyelela ku-Opentrons Support, sicela ubazise ukuthi uyingxenye yohlelo lwethu lwe-Opentrons for Education kanye nosuku lwekilasi lakho lelebhu elilandelayo.

Imisebenzi Edingekayo Yangaphambi Kwelebhu

Ngaphambi kokuqala le lebhu, abafundi kufanele babe namakhono alandelayo ezobuchwepheshe nolwazi lwethiyori:

• Layisha ithiphu le-pipette ku-pipette yezandla
• Ukulungiswa kwevolumu yamapayipi okwenziwa ngesandla
• Ulwazi lwethiyori lwe-spectrophotometry -Uthisha uzobe esebenzisa i-spectrophotometer yale lebhu
• Ukuqonda amajika ajwayelekile, okuhlanganisa ulwazi lwesibalo oluchaza ijika, nekhono lokuhlanganisa amanani
• Ukuqonda okuqinile kokuhlaziywa kwezibalo okuyisisekelo, okuhlanganisa izindlela zokubala nezokuhumusha, ukuchezuka okujwayelekile, kanye no-R²

Umhlahlandlela Wenqubo

1. Isingeniso Selebhu ~ imizuzu eyi-15

Ngokufanelekile, abafundi kufanele bafunde umhlahlandlela walebhu ngaphambi kokuza ekilasini, nokho, bahlele ukuchitha imizuzu eyi-10 bexoxa ngamasu okufaka amapayipi, kabusha.view kwamajika ajwayelekile, kanye nokuhlanganisa sample amanani.

2. I-Calibration Curve Generation ~ imizuzu engama-20

Okulandelayo, bonisa ukukhiqizwa kwejika elijwayelekile (Ingxenye A) yekilasi (le lebhu ithatha ngokuthi uthisha kuzoba yedwa u-opharetha werobhothi le-OT-2 ne-spectrophotometer):

1. Lungiselela ikhambi likadayi oluhlaza ngo-50 μL wodayi wokudla oluhlaza okwesibhakabhaka no-10 mL we-diH2O.
2. Engeza u-200 μL wesisombululo sikadayi oluhlaza emithonjeni A1-H1 yepuleti eliphansi eliyisicaba elinemithombo engu-96.
3. Faka u-20 ml wamanzi emthonjeni ongu-A1 wechibi lomkhombe.
   (Qaphela: Ikholomu ngemva kokuhlanjululwa kokugcina indawo ezenzakalelayo yezikhala. Umthombo wokugcina womkhombe/indawo yokugcina amanzi udoti owuketshezi ozenzakalelayo.)
4. Hlela i-labware kanje:
   a. Isikhala 1 = Opentrons 300 μL ithiphu yokubeka
   b. Isikhala 2 = 12-channel ichibi
   c. Isikhala 3 = ipuleti eliphansi eliyisicaba elingu-96
5. Qhubekela kuhlelo lokusebenza olusebenzayo lwe-OT-2 ukuze uqalise umthetho olandelwayo.
6. Linganisa ukumuncwa kwepuleti lemithombo engu-96 ku-450 nm.

3. Ingxoxo ~ imizuzu emi-5

Ngemva kokusebenzisa i-serial dilution nokukhiqiza ijika elijwayelekile, ungase ufise ukubuya ndawonye njengekilasi futhi nixoxe ngamandla namathuba okuzenzakalela.

4. Ingxenye B: Imanuwali Sample Ukulungiselela ~ 1 ihora

Ngale ngxenye yelebhu, abafundi bazocelwa ukuthi bagcwalise ngokuzimela Ingxenye B yelebhu:

1. Sebenzisa ishubhu elilodwa lokuhlola ukuze uqoqe i-aliquot engu-200 μL kadayi ogxilile.
2. Sebenzisa ishubhu lokuhlola lesibili ukuqoqa u-4 mL we-diluent (diH2O).
3. I-Pipette okungenani i-800 μL ye-diH20 kushubhu ngalinye kwamane asele. Lezi zizoba ezakhoampLes.
4. Sebenzisa i-dropper ukuze ukhiphe amanani ahlukahlukene odayi ogxilile esigabeni ngasinyeampithubhu.
5. I-Pipette 200 μL yemizuzwana yakho yokuqalaampungene emthonjeni wepuleti elinemithombo engama-96. Phinda lokhu izikhathi ezi-3 ngaphezulu ukuze ube nezimpinda ezi-4 zaleso sigabaample emithonjeni emi-4 yepuleti. Bhala phansi ukuthi yimiphi imithombo oyibekile leample into (cishe uzokwabelana ngepuleti lakho lemithombo engama-96 nabanye abafundi - ufuna ukwazi ukuthi isiphi isitshaamplezi ezakho!).
6. Phinda isinyathelo esingenhla kumasekhondi angu-3 aseleampLes. Kufanele ube nemithombo emi-4 yesekhondi ngalinyeample, ngesamba semithombo engu-16, egcwaliswe epuleti elinemithombo engama-96.
7. Linganisa ukumuncwa kwepuleti lemithombo engu-96 ku-450 nm.

Ngenkathi abafundi beqedela Ingxenye B, hlela ukuzungeza ilebhu ukuze baphendule imibuzo futhi babheke indlela yomfundi yokufaka amapayipi, bakhumbuze abafundi ukuthi barekhode ukuthi yimiphi imithombo abayisebenzisayo, futhi banikeze usizo lomuntu ngamunye njengoba kudingeka.

5. Ukuqoqwa Kwedatha ~ imizuzu engama-30

Vumela imizuzu engu-30 yokuqoqa idatha.

6. Ukuhlukana ~ 10 imizuzu

Shiya isikhathi ekupheleni kwelebhu ukuze wethule kafushane umbiko walebhu futhi unikeze abafundi isikhathi esanele sokuhlanza iziteshi zabo futhi babuze imibuzo esele.

Umbiko Welebhu

Iziyalezo

Yabela abafundi ukuthi balungiselele umbiko welebhu ophelele ohlanganisa idatha yekhevu yokulinganisa, i-interpolated sample amanani, nokuhlaziywa kwezibalo. Nikeza imihlahlandlela yesakhiwo sombiko kanye nokwethulwa kwedatha. Eminye imibono yokuhlola ingezansi.

• Misa ijika elijwayelekile
  • Hlanganisa amanani angawodwana ngezindinganiso zakho.
  • Thola ukuchezuka okumaphakathi nokujwayelekile kokunye.
  • Sebenzisa amanani amaphakathi ukuze uchaze ijika elijwayelekile.
  • Nquma ukuthi ijika elibaliwe liwafanela kahle kangakanani amaphuzu edatha ngokubala inani elingu-R2. Khumbula ukuthi umugqa olingana kahle namaphoyinti edatha unevelu engu-R2 engu-1. Kubalulekile ukwakha ijika elijwayelekile elisha elinevelu engu-R2 ebaliwe yokuhlolwa ngakunye.
• Ukuhunyushwa kuka-Sample Amagugu
  • Sebenzisa ijika lakho elijwayelekile elibaliwe ukuze uhlanganise u-sample amanani mathupha.
  • Kuyini ukuchezuka okujwayelekile phakathi kwezimpinda zesigaba sakho ngasinyeampLes?
  • Yiziphi ezinye zezizathu ezingaba khona zalokhu kwehluka?
  • Yiziphi izinzuzo zokulinganisa izimpinda?
• Imaphi amathrendi ongawabona phakathi kwamasethi ahlukene wama-sampLes? Ingabe noma yimuphi umehluko ophawuliwe obalulekile ngokwezibalo?
  • Ingabe akhona ama-outliers? Ukusetshenziswa kwezibalo kuvumela kanjani ukuhlolwa kwedatha ye-outliers vs yokukha ama-cherry?

Umhlahlandlela Womfundi
Izilinganiso kanye Nokuhlaziywa Kwezibalo

Inhloso

Le lebhu izothuthukisa ukuqonda kwakho kokusebenzisa ukuhlanjululwa kwe-serial ukuze ukhiqize ijika elijwayelekile, kanye nokusetshenziswa kwamajika ajwayelekile okufaka phakathiample amanani. Lokhu kubandakanya ukubonga:

• Amakhono ayisisekelo wepayipi
• Ama-serial dilutions
• Ukulinganisa ukuphindaphinda
• Ububanzi bejika elijwayelekile
• Ukusetshenziswa kwezibalo ukuhlola ukubaluleka

Kule lebhu abafundi bazosebenzisa idatha etholwe ngokufaka amapayipi okwenziwa ngesandla kanye nesibambi soketshezi esizenzakalelayo se-OT-2, okuvumela abafundi ukuthi babone amandla namathuba azo zombili lezi zindlela.

Izisetshenziswa ezidingekayo

• I-Opentrons OT-2 Automated Liquid Handling Robot
• I-Opentrons p300 8-channel pipette
• I-Opentrons 300 μL I-Tip Rack
• I-pipette eyenziwe ngesandla (P1000)
• 12 - umthombo
• Ipuleti le-Flat Bottom engu-96
• 1000 mL amanzi enziwe deionized (diH2O)
• I-FD&C Blue No. 1 (McCormick® yasetshenziswa ekuthuthukisweni kwale lebhu)
• 6 Amashubhu okuhlola
• I-Dropper

Inqubo Yokuhlola

Ingxenye A: Qaphela Isizukulwane Esijwayelekile Sejika

I-OT-2 izokwenza i-automated pipetting ye-serial dilution. Uma uthisha wakho ekala la mazinga nge-spectrophotometer uzokwazi ukwenza ijika elijwayelekile.

Uthisha wakho uzolandela le miyalo ukuze alungiselele futhi asebenzise irobhothi le-OT-2. Izinyathelo 1-4 zizoqedelwa mathupha (ngothisha wakho noma amavolontiya ayisitshudeni) ngaphambi kokusebenzisa izinyathelo zamarobhothi e-OT-2:

1. Lungiselela ikhambi likadayi oluhlaza ngo-50 μL wodayi wokudla oluhlaza okwesibhakabhaka no-10 mL we-diH2O.
2. Engeza u-200 μL wesisombululo sikadayi oluhlaza emithonjeni A1-H1 yepuleti eliphansi eliyisicaba elinemithombo engu-96.
3. Faka u-20 ml wamanzi emthonjeni ongu-A1 wechibi lomkhombe.

(Qaphela: Ikholomu ngemva kokuhlanjululwa kokugcina indawo ezenzakalelayo yezikhala. Umthombo wokugcina womkhombe/indawo yokugcina amanzi udoti owuketshezi ozenzakalelayo.)

4. Hlela ilebula kanje:

ο Isikhala 1 = I-Opentrons 300 μL ithiphu yokubeka
ο Isikhala 2 = 12-indawo yokugcina amanzi
ο Isikhala 3 = 96-ipuleti eliphansi eliphansi eliphansi

5. Qhubekela kuhlelo lokusebenza lwe-OT-2 ukuze usebenzise iphrothokholi ezenzakalelayo.

ο Landa umthetho olandelwayo kusuka
https://protocols.opentrons.com/protocol/customiza ble_serial_dilution_ot2

• Khetha amapharamitha alandelayo ku- webikhasi ngokuya ngokusetha kwakho kanye nempahla:
  • Uhlobo lwe-Pipette
  • Mount Side
  • Uhlobo Lwethiphu
  • Uhlobo Lomkhombe
  • Uhlobo Lwepuleti
• Gcina izilungiselelo ezizenzakalelayo ze:
  • I-Dilution factor (3)
  • Inani lokuhlanjululwa (10)
  • Ivolumu yokuxuba isiyonke (150)
  • Akunalutho epuletini lomthombo (yebo)
  • Isu lokusebenzisa isu (sebenzisa ithiphu elilodwa)
  • Umthamo wegebe lomoya (10)

6. Linganisa ukumuncwa kwepuleti le-96-well ku-450 nm.

Qaphela: Ukuhlanjululwa kwe-serial kwenziwa ngokudlulisela ngokulandelana ama-aliquots wezinga emananini akhulayo e-diluent (bona umdwebo). I-OT-2 izofuna i-aliquot yezindinganiso ezigxilisiwe futhi iyikhiphe ku-diluent. Izobe-ke ixuba udayi ogxilile kanye ne-diluent ngokuphindaphinda ngamapayipi uketshezi phezulu naphansi. Bese irobhothi lizofuna i-aliquot yezinga lokuqala elixutshiwe futhi liyikhiphe kukholomu elandelayo ye-diluent.

Ingxenye B: Ukuziphatha Imanuwali Sample Ukulungiselela

1. Sebenzisa ishubhu elilodwa lokuhlola ukuze uqoqe i-aliquot engu-200 μL kadayi ogxilile.
2. Sebenzisa ishubhu lokuhlola lesibili ukuqoqa u-4 mL we-diluent (diH2O).
3. I-Pipette okungenani i-800 μL ye-diH20 kushubhu ngalinye kwamane asele. Lezi zizoba ezakhoampLes.
4. Ukudala ama-s ahlukeneampi-les enamanani ahlukene okugxilisa ingqondo uma kuqhathaniswa ne-diluent: Sebenzisa i-dropper ukuze ukhiphe amanani ahlukahlukene kadayi ogxilisiwe ku-s ngayinye.ampithubhu.
5. I-Pipette 200 μL yemizuzwana yakho yokuqalaampungene emthonjeni wepuleti elinemithombo engama-96. Phinda lokhu izikhathi ezi-3 ngaphezulu ukuze ube nezimpinda ezi-4 zaleso sigabaample emithonjeni emi-4 yepuleti. Bhala phansi ukuthi yimiphi imithombo oyibekile leample into (cishe uzobe wabelana ngepuleti lakho lemithombo engama-96 nabanye abafundi ofuna ukwazi ukuthi isiphi isitshaamplezi ezakho!).
6. Phinda izinyathelo 5 no-6 kumasekhondi angu-3 aseleampLes, ngakho-ke ugcina ngokuphindaphinda okungu-4 kwe-s ngayinyeample emithonjeni emi-4. Kufanele ube nemithombo emi-4 yesekhondi ngalinyeample, ngesamba semithombo engu-16, egcwaliswe epuleti elinemithombo engama-96.
7. Linganisa ukumuncwa kwepuleti lemithombo engu-96 ku-450 nm.

Imibuzo Yabafundi
Izilinganiso kanye Nokuhlaziywa Kwezibalo

Imibuzo Yabafundi

1. Iyini inzuzo yokusebenzisa i-serial dilution ukuze ukhiqize ijika elijwayelekile?
2. Kungani ukukhiqizwa kwamajika okulinganisa kubalulekile ezilinganisweni zebhayoloji?
3. Yisho inselele eyodwa engaba khona ekukhiqizeni amajika okulinganisa bese uphakamisa isisombululo.
4. Chaza indima yezinto ezisetshenziswa kule lebhu.
5. Yimiphi imithombo evamile yokuhlukahluka kusuka kumapayipi okwenziwa ngesandla?
6. Kungani kuwusizo ukukala ukuphindaphinda kwe-s ngayinyeample?
7. Iyini inhloso yokuhumusha isi-sampamavelu kulesi sivivinyo?
8. Ukuhlaziywa kwezibalo kungasiza kanjani ekuchazeni umehluko phakathi kwezigaba ezahlukahlukeneampLes?
9. Okutholwe kule lebhu kungase kuhlobane kanjani ocwaningweni lwebhayoloji?

Amadokhumenti / Izinsiza

Opentrons OT-2 Workstations for Efficient Lab Automation [pdf] Iziyalezo I-OT-2 Workstations for Efficient Lab Automation, OT-2, Workstations for Efficient Lab Automation, Efficient Lab Automation, Lab Automation, Automation

Izithenjwa

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