3B SCIENTIFIC 1012852 [W19925] Owner's manual

Brand: 3B SCIENTIFIC

Model: 1012852 [W19925]

Category: Mounting kits

Type: Owner's manual

3B SCIENTIFIC 1012852 [W19925] is an electrophoresis chamber designed for horizontal gel electrophoresis. It is equipped with corrosion-resistant platinum electrodes and high-quality connections for the positive and negative poles. The chamber comes with a safety lid with attached power cords, two combs with 12 teeth each, and an instruction manual.

The electrophoresis system can be used with CE-marked power supplies. It is important to disconnect the chamber from the power supply before removing the safety lid. The chamber should not be placed on wet surfaces and should be disconnected from the power supply when not in use. The maximum current and voltage for the chamber are 150 mA and 120 V, respectively.

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Scope of Supply

The model ’Mini’ contains the following items:

• Electrophoresischamberwith corrosionresistantplatinumelectrodes andhigh-quality connectionsforplusand

minuspole

• Safetylidwithattachedpowercords

• 2combs,1.5mmthick,12teetheach

• Instructionmanual

Safety Instructions

• Pleasereadtheinstructionmanualcarefullybeforeusingtheelectrophoresissystem.

• UseonlyCEmarkedpowersupplies.

• Alwaysdisconnecttheelectrophoresischamberfromthepowersupplybeforeremovingthesafetylid.

• Donotputthesafetyliddownonwetsurfaces.

• Alwaysdisconnecttheelectrophoresischamberfromthepowersupplywhennotinuse.

• Thechamberisprovidedforamax.currentandvoltage.Donotexceed150mA,120V.

• Pleasemindthemaximumfillinglevelfortheelectrophoresisbuffer.

Technical Properties

Thehorizontal electrophoresissystem‘Mini’ hasbeen designedin sucha waythatallows theuser tocast andrunthegel

directlyinasinglechamber.Noadditionalcastingequipment,suchasgreaseorsealisrequiredtopreparethegeltrayfor

castingthegel.

Thechamber’sbottomisUV-permeable,thusthegelruncaneasilybedocumentedincaseofusingDNA-fluorescentdyes.Self

evidently,thegelcanalsoberemovedfromthechamberforstaininganddocumentationafterthegelrun.

Please use only UV light with a wave length of 300nm or higher for all applications, as hard UV light (<300 nm) damages

the gel chamber and the nucleic acids.

Ifyouneedonlyshortrunsareneeded,youcanusebothcombswhichthengivestwiceasmanyslotsforsamples.

Warranty

Themanufacturerguarantees thattheelectrophoresischamberwas completelytested beforedeliveryandthatit complies

withtheapplicablesafetyregulations.

Pleasecheckthedeliveryimmediatelyuponreceiptforcompletenessandpossibledamageintransit.Ifanythingisfaultyor

damaged,pleasecontact3BScientificimmediately.

Themanufacturergivesa24-monthwarrantyontheproductconditionalonthechamberbeingusedaccordingtotheinstruc-

tionmanual.Claimsforreplacementorrepairarenotvalidifphysicalabuseisthereasonforthedamage.Anyliabilityfor

conseqentialdamagesresultingfromtheuseoftheelectrophoresischamberisherebyexcluded.Themanufacturer‘sliability

forintentandgrossnegligenceorfordamagesresultingfrominjurytolife,bodyorhealthisunaffectedbythat.

Ourexperienceshows thetwoplatinumelectrodeslast formanyyearsif theproductisused properly. AnyDamageoccur-

ringis usually dueto mechanicalerrors (e.g. bywashing-up brushes)or contactwith so-called“platinum poisons”such

asphosphorus,boronandheavymetals(lead,zinc, etc.). For thisreason,the platinumelectrodesareexcluded fromthe

manufacturer’swarranty.

Inordertointroducedevelopmentspromptly,themanufacturerreservestherighttochangespecificationsandsmallvisual

detailswithoutpriornotice.

General Instructions

Casting of agarose gels

1. Pleasewearprotectiveclothing,gogglesandglovesforheatingtheagarosesolution.

2. Removethesafetylidoftheelectrophoresischambercarefully.

3. Forthepreparationofthegelsolution,onlysuitableagarosesandelectrophoresisbuffersshouldbeused.Forpreparing

theagarosesolution,weighasuitableamountofagaroseinasmallErlenmeyerflask,addanappropriateamountofelec-

trophoresisbufferandastirbar(ifheatingwithheatingstirrers).Notetheweightofthefilledflasksothatanylossduring

heatingcanbecompensatedforbytheadditionofdistilledH2O.Thus,thegelsolutionwillhavetheagaroseconcentration

Electrophoresis Chamber ‘Mini’

English

thatyouneed.Todissolvetheagarose,heattheErlenmeyerflaskeitherinamicrowaveorwithaheatingstirrer.Forthe

latter,usemediumheatingpowerandstirconstantly.Heatinginamicrowaveshouldonlybeforshortperiodsoftime,on

mediumheatinglevel,repeatedmultipletimes.Whenheatinginamicrowave,removetheErlenmeyerflaskfromtimeto

time(remembertoweargloves,gogglesandbecarefultoavoidboiling!)andgentlyswirlthegelsolutioninacircle.Putit

backinthemicrowaveafterwardsandrepeattheentireprocess3-4times,untiltheagaroseiscompletelydissolved.

4. Beforethecastingofthegel,allowthegelsolutiontocooldownto60°C.Castthegelsolutionwithoutanyairbubblesin

thegapbetweenthetwowhiteplasticstripesonthechamber’sground(itscapacityisabout40ml).Theninsertthecomb

(orbothcombsrespectively).Makesurethattherearenoairbubblesontheedgesoftheslots.Gelsmadeofstandardaga-

rosesolidifywithin20minutesatroomtemperature.

5. Afterthesolidificationoftheagarosegel,addsomeoftheelectrophoresisbufferin3-mmlayersinthechamberoverthe

geltopreventifromdryingout.Aftercarefullyremovingthecomb(s),thegelcannowbeloaded,Thecoatedgelcanbe

storedforafewdaysat4°C.Forbestresultsleavethecombinsertedandcoverthegelwithwrappingfilmtoprotectit

fromdryingout.

Loading of gels and electrophoresis

1. Afterthesolidificationoftheagarosegel,coverthegelwiththeelectrophoresisbuffer.Pleasenotethemaximumfilling

levelfortheelectrophoresisbuffer.

2. Removethecomb(s)fromtheagarosegelbygentlymovingbackandforthandthencarefullypullingoutupright.

3. Inthenextstep,theslotsinthegelcanbeloadedwiththesamples.Preparingthesampleswithanappropriategelloading

bufferwhichincreasesthespecificweightofthesamplesmakesiteasiertopourthemintotheslotswithamicropipette

(seesection“gelloadingbuffer”).

4. Whenloadingthegelslots,dipthetipofthemicropipettecarefullyintotheslotandthenslowlypressoutthesample.

Makesurenottodamagethebottomoftheslot.

 Beginnersshouldpractisethiswith„practicesamples“consistingofH2Oandgelloadingbufferonly.

5. AtleastonesizestandardshouldbeappliedoneachgelsothatthesizeofthevariousDNAfragmentscanbedetermined.

6. Nowplacethesafetylidontheelectrophoresischamberandconnectthechambertoasuitablepowersupply.

 Pleasenotethecorrectpolarity.Inanalkalineuptoaneutralmedium,nucleicacidsarechargednegativelyandmigrateto

theanode(redpole).

7. Turnonthepowersupplyandruntheelectrophoresisatavoltageintherange80-120V.Theprocessoftheelectrophore-

siscanbeobservedwiththehelpofthedye,whichisinsidethegelloadingbuffer.

8. Bromophenolblueandxylenecyanolarefrequentlyuseddyesforthegelloadingbuffer.Thesedyesarealsonegatively

chargedandmigrateto theanode.This process,the so-calledco-migrationwiththedouble-stranded DNAfragments,

dependsondifferentfactors,suchasagarosetype,electrophoresisbufferandgelstrength.

 Foraroughestimation:bromophenolbluemigratesin1xTAEelectrophoresisbufferand1%-standardagarosegelinthe

samewayasaDNAfragmentof650basepairs.Underthesameconditions,xylenecyanolmigrateslikeafragmentwith

5000basepairs.

Staining of nucleic acids

Sincethere aredifferentmethods ofstaining nucleicacids, wereferat thispoint tothe correspondingtechnicalliterature

(Sambrooket.al.,1989).

Evaluation / documentation

Sizedeterminationsofnucleicacidfragmentscanbecarriedoutbycomparisonwiththefragmentsofalengthstandard.A

detaileddescriptionofthisprocedurecanbefound,forexample,inMolecularGeneticsbyR.Knippers(2008).

Cleaning and Maintenance

Attention: Disconnecttheelectrophoresischamberfrompowersupplybeforecleaning.

Thegelchamberandthecombsshouldbecleanedwithwarmwaterimmediatelyaftereachuse.Ifnecessary,afewdropsof

awashing-upliquidcanbeadded.Pleasedonotusedishwashingbrushesorthelike,asthiscandamagetheplatinumelec-

trodes.Rinse theelectrophoresischamberaftercleaningwith distilledor demineralisedwatertoprevent theformationof

limescale.

Neverleavethechamberuncleanedforhoursorevenovernightasresiduesfromgelorbuffercancausestainswhicharediffi-

culttoremovecompletely.

Attention:Pleaseavoidthe electricalconnectionsbeing incontactwith water.Ifhoweverthis doesoccur, pleasecarefully

wipedrywithasoftclothandallowtoair-dry.Pleasedonotusepaperbecauseitisoftentooroughandcausessmallscrat-

ches.Donotuseahairdryereither,asthehotaircandamageconnectors,fittingsandtheacrylicmaterial.

Attention:Donotuseethanolnorotherorganicsolventsforcleaning.Itcancausecracks,scratchesorotherunwantedmate-

rialchanges(blindnessetc.).

Required Material and Processes

Electrophoresis buffer

Theelectrophoresisbufferprovides thenecessary ionsforthe electrophoresisprocess. It’sadditionprovidesaconstantpH

valuesothatnucleicacidshavethedesirednetcharge.Nucleicacidsarenegativelychargedinanalkalineuptoneutralmedi-

um.Normally,electrophoresisbuffercontainscomponentsthatprotectnucleicacidsfromdegradation,e.g.EDTA,whichcom-

plexesdivalentcationsandthereforeinhibitsDNases.

Atthispoint,thefrequentlyusedTAEelectrophoresisbufferforelectrophoresisofDNAisdescribedfornon-denaturingcon-

ditions.TAEstandsforTris-acetate-EDTA.Youcaneitherproducethebufferyourselfororderitfrom3BScientificasabuffer

concentrate

Agarose, gel volumes and gel concentrations

Eventhoughvariousagarosesareoffered,thesocalledstandardagaroseiscertainlyofprimeimportance.Forthecastingof

anagarosegelinthischamberabout40mlofgelsolutionisneeded.Pleasepreparealittleextragelsolutiontoallowforthe

factthatsmallresiduesalwaysremaininthevessel.

Optimalseparationofmoleculesofdifferentsizeswillbeachievedbyvaryingtheconcentrationofthegelaccordingtothe

informationinthechartbelow.

Optimal separation ranges of the following DNA length (double-stranded DNA) corresponding to

various concentrations of agarose (standard agarose)

concentration of agarose (%) agarose (g) buffer (ml) optimal separation range (kbp)

 0,5 0,25 50 1–15

 0,7 0,35 50 0,8–10

 1,0 0,5 50 0,5–7

 1,2 0,6 50 0,3–6

 1,5 0,75 50 0,2–4

 2,0 1,0 50 0,1–3

Gel loading buffer

Thesampleswhicharetobeanalyzedneedtobemixedwithasuitablegelloadingbufferbeforeapplyingthemtothegel.Gel

loadingbuffercontainsdye(s)fortrackingtherunoftheelectrophoresisaswellasglycerin,saccharoseandthelike.Therefore,

thepreparedsamplesareheavierthantheelectrophoresisbufferandwilldecreaseslightlyintothegelslotsduringthesam-

pleapplication.Bromophenolblueandxylenecyanolarefrequentlyuseddyesforgelloadingbuffers.Thepatternsoftheir

run(orthesocalledco-migrationtodouble-strandedDNAfragments)dependonthetypeofagarose,gelstrengthandthe

typeofelectrophoresisbuffer.

Foraroughestimation:bromophenolbluemigratesin1xTAEelectrophoresisbufferand1%-standardagarosegelinthesame

wayasaDNAfragmentof650basepairs.Underthesameconditions,xylenecyanolmigrateslikeafragmentwith5000base

pairs.

English

DNA length standard

Forthesizingofthesamples,applyoneDNAsizestandardoneachgelatleasttoonelane.DNAsizemarkesaremadeofDNA

fragmentsoffamiliarsizes.

Staining of nucleic acids

Sincethere aredifferentmethods ofstaining nucleicacids, wereferat thispoint tothe correspondingtechnicalliterature

(Sambrooket.al.,1989).

Declaration of Conformity

Theelectrophoresischamber’Mini’complieswiththeapplicablesecurityrequirementsaccordingtoEN:50081-1(1992)and

EN:50082-1(1992).

Literature

Ausubel,FrederikM.etal.(Ed.),(2005).ShortProtocolsinMolecularBiology,

ACompendiumofMethodsfromCurrentProtocolsinMolecularBiology.Wiley-VCH,Hoboken.

Knippers,R.(2008).MolekulareGenetik.ThiemeVerlag,Stuttgart.

SambrookJ,Fritsch E.F. ManiatisT.(1989). MolecularCloning: ALaboratory Manual.Cold SpringHarbor Laboratory Press,

NewYork.

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DNA-Längenstandard

EinDNA-LängenstandardwirdaufjedemGelmindestensineineSpuraufgetragen,umeineGrößenbestimmungderProben

vernehmenzukönnen.DNA-LängenmarkerbestehenausDNA-FragmentenbekannterGröße.

Anfärbung von Nukleinsäuren

Daes verschiedene Möglichkeitender Anfärbung vonNukleinsäurengibt, wirdan dieser Stelleauf dieeinschlägige

Fachliteraturverwiesen(Sambrocket.al.,1989).

Konformitätserklärung

DieElektrophoresekammer „Mini“entspricht dengrundlegendenSicherheitsanforderungengemäß EN:50081-1 (1992)und

EN:50082-1(1992).

Literatur

Ausubel,FrederikM.etal.(Ed.),(2005).ShortProtocolsinMolecularBiology,

ACompendiumofMethodsfromCurrentProtocolsinMolecularBiology.Wiley-VCH,Hoboken.

Knippers,R.(2008).MolekulareGenetik.ThiemeVerlag,Stuttgart.

SambrookJ,Fritsch E.F. ManiatisT.(1989). MolecularCloning: ALaboratory Manual.Cold SpringHarbor Laboratory Press,

NewYork.

Deutsch

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Français

L’échelle de marqueur de taille moléculaire

Uneéchelledemarqueurdetailled’ADNestappliquéesurchaquegelaumoinssurunetrace,pourpouvoirmesurerleséchantil-

lons.Cesmarqueursdetailled’ADNconsistentenfragmentsd’ADNdetailleconnue.

Coloration des acides nucléiques

DaesverschiedeneMöglichkeitenderAnfärbungvonNukleinsäurengibt,wirdandieserStelleaufdieeinschlägigeFachliteratur

verwiesen(Sambrocket.al.,1989).

Konformitätserklärung

Commeilyadesméthodesdifférentesdecolorationdesacidesnucléiques,nousattironsvotreattentionsurlalittératurespécia-

liséecorrespondante(Sambrocket.al.,1989).

Littérature

Ausubel,FrederikM.etal.(Ed.),(2005).ShortProtocolsinMolecularBiology,

ACompendiumofMethodsfromCurrentProtocolsinMolecularBiology.Wiley-VCH.

Knippers,R.(2008).MolekulareGenetik,ThiemeVerlag,Stuttgart.

Sambrook,J,Fritsc,E.F.,Maniatis, T. (1989).MolecularCloning: ALaboratory Manual.ColdSpring HarborLaboratory Press,

NewYork.

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3B SCIENTIFIC 1012852 [W19925] Owner's manual

Brand: 3B SCIENTIFIC

Model: 1012852 [W19925]

Category: Mounting kits

Type: Owner's manual

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français: 3B SCIENTIFIC 1012852 [W19925] Le manuel du propriétaire
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3B SCIENTIFIC 1012852 [W19925] Owner's manual

3B SCIENTIFIC 1012852 [W19925] Owner's manual

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