7/27/2019 1 Bioanalyzer Instructions Edge Lab

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Edge lab, updated June 2011

Bioanalyzer InstructionsAdapted from Agilent RNA 6000 Nano Kit Guide G29389003!

Notes

"f t#ere$% a problem, u%e diagno%ti&% tab to get info to %end to Agilent'

Repla&e t#e %(ringe at t#e &#ip priming %tation ea&# time (ou %tart a ne) bo* of2+ &#ip%' Agilent RNA 6000 Nano Kit: 5067-1511

#en RNA ladder fir%t arri-e% .&ome% )it# 30/ per -ial, #eat denature tube for

2 min at 04, &ool on i&e, and ali5uot into dail( u%e -ial%' Agilent rep

re&ommend% + /, but onl( 1/ i% needed per &#ip' tore -ial% at 804 and t#a)on i&e )#en read( to u%e'

Allo) reagent% to e5uilibrate to room temperature for 30 minute% before u%ing'

At least 30 minutes beore beginning

7ull out %ample% and ladder .ali5uoted into indi-idual u%e tube% from 804

freeer :eep on i&e until u%e 7ull out t#e follo)ing reagent% from !4 fridge ; allo) to &ome to room temp

o 6000 Nano d(e .blue &ap

o 6000 Nano mar:er .green &ap

o 6000 Nano gel matri* .red &ap < only if needed; see below

!re"aring gel matri#$ =ou &an get eig#t6+/ ali5uot% of gel matri* from one prep'

1' +00/ Agilent RNA 6000 Nano gel matri* .red into %pin &olumn'2' pin at !000 rpm for 10 min'

3' 6+ / elutant into eig#t 0'+ / tube%' tore at !4 for 1 mont#'

B%&'R% ()% 'N*+ , Aing .ye

!' >orte* 6000 Nano d(e .blue for 10 %e& and 5ui&: %pin+' Add 1 / d(e to t#e 6+/ ali5uot of filtered gel

6' >orte* and %pin at 1!000 rpm for 10 min'?Note@ "f (ou are running multiple &#ip% in 1 da(, (ou &an u%e remaining geld(e

matri* %olution' B#ere %#ould be enoug# geld(e matri* %olution in 1 ali5uot for at

lea%t 2 &#ip%'

)am"le "re" / loaing gel matri#

1' Cioanal(er &an anal(e a ma*imum of +00 ng/' "f ne&e%%ar(, dilute %ample

)it# RNa%efree )ater for a final -olume of 1 /'

2' Deat denature %ample% at 04 for 2 minute% t#en &ool on i&e'

3' #en loading t#e &#ip, al)a(% put t#e pipette tip to t#e bottom of t#e )ell todi%pen%e li5uid'

!' 7ipette 9 / gel d(e mi* at t#e bottom of t#e )ell )it# t#e bold $ 7ut &#ip in

 priming %tation, %et timer to 30 %e&, ma:e %ure plunger i% at 1 m, and depre%% t#e%(ringe until it lo&:% beneat# t#e lat&#' Bime )ait for 30 %e&' Relea%e plunger and

it )ill relea%e it%elf at lea%t until t#e 0'3 m mar:' ait for + %e& t#en pull plunger 

up to t#e 1 m mar:'

7/27/2019 1 Bioanalyzer Instructions Edge Lab

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Edge lab, updated June 2011

+' pen priming %tation and pipette 9 µ gel d(e mi* into ea&# remaining G )ell'

*oaing marer2 laer2 an sam"les

1' 7ipette + / RNA 6000 Nano mar:er .green into )ell mar:ed )it# ladder%(mbol and ea&# of t#e 12 %ample )ell' .All )ell% NB mar:ed G

2' 7ipette 1 / ladder into )ell mar:ed )it# a ladder %(mbol'

3' 7ipette 1 / %ample% into 12 %ample )ell%'!' F NB run empt( )ell% on a &#ip' Al)a(% fill e-er( )ell )it# a %ample'

+' >orte* on 2!00 rpm for 60 %e& .it %#ould %top b( it%elf'

6' 7la&e &#ip in bioanal(er, &lo%e lid, load %oft)are'' ele&t appropriate a%%a( from Assay menu . Electrophoresis, RNA, %ele&t

 Eukaryotic Total RNA Nano, t#en &li&: t#e Start  button' ill out %ample name%'

Run )ill ta:e H30 min'

?Note < if (ou are running multiple &#ip%, a-oid allo)ing &#ip% to %it around for

more t#an + minute% before running on t#e bioanal(er' ait until t#e &omputer

nearl( fini%#e% one &#ip before %tarting t#e prep on t#e ne*t &#ip'

.ata )aing

• Cioanal(er automati&all( %a-e% file% under t#e 4 dri-e a% an '*ad file, )#i&# i%

onl( readable b( bioanal(er %oft)are' ile dire&tor(@ I( 4omputer → 4 dri-e

→ 7rogram ile% → Agilent → 2100 Cioanal(er → 2100 e*pert → data' nder

t#e Fata folder, file% are organied b( data

• Bo print data a% 7F, &li&: 7rint'L B#i% )ill allo) (ou to %a-e t#e data a% a 7F

file in t#e folder Cioanal(er 7F%L .file dire&tor(@ I( 4omputer → Fata .F

dri-e → Cioanal(er 7F%, )#ere t#e( are organied b( date' B#ere i% a

%#ort&ut on t#e de%:top to t#e Cioanal(er 7F% folder a% )ell'

• n&e (ou open t#e 7F, (ou &an print' B#e 7F i% 18 page% long, %o be %ure to print onl( t#e 1%t %ummar( page, unle%% (ou need more' Ce %ure to %a-e a &op( of

t#e 7F under t#e appropriate I dri-e folder'

4leanu"

1' 4lean tip of %(ringe after e-er( u%e )it# i%opropanol, )ipe dr( )it# Kim )ipe

to pre-ent &logging' Repla&e if gel doe% not run e-enl('2' After e-er( run, u%e &leaner &#ip% .1 )it# RNa%e A)a(, 1 RNa%e free )ater to

&lean ele&trode pin %et' Ju%t allo) bioanal(er to be on for 1 min ea&#'

oft)are doe% not #a-e to be running' 4#ip% %#ould be filled )it# 3+0/RNa%e A)a( or )ater' ait 10 %e&ond% for )ater to e-aporate from ele&trode%

 before &lo%ing lid'

3' n&e a )ee:, &lean probe%' 7our RNa%efree )ater onto toot#bru%# and gentl()ipe' ea-e t#e lid of t#e bioanal(er open for 10 %e&ond% to allo) )ater to

e-aporate from probe%'