SE300 User Manual – English
Contents
1. e p15 Electrophoresis Troubleshooting problem solution Smile effect on the buffer front To reduce the running temperature Fill the tank to the maximum marked buffer level Prechill the buffer Decrease the current or voltage setting 10 mA per 0 75 mm gel 15 mA per 1 5 mm thick gel Run the gel in the cold room Protein streaks vertically Centrifuge or filter sample before loading to remove particulates Dialyze or desalt the sample Unusually slow or fast run Adjust the solutions Check recipes gel concentrations solutions and dilutions For instance do not use Tris HCI instead of Tris e f the required pH of a solution is exceeded do not back titrate Prepare fresh buffer Dispose of older acrylamide solutions and use only stock of the highest quality Only use freshly deionized urea Adjust the voltage or current settings To increase or decrease the migration rate adjust the voltage or current by 25 50 Bands are skewed or distorted e p16 Check gel preparation and polymerization Degas the stacking gel solution and avoid trapping air bubbles under the comb teeth Overlay the running gel with water saturated n butanol before polymerization begins to avoid forming an uneven gel surface Check sample preparation Dialyze or desalt the sample Centrifuge or filter sample bef2. sti z tohoto n stroje Rozpustidl m zp sob nenapravi teln po kozen jednotka Nejsou provozov na s pufru teplot ch nad maxim ln stanovenou technick mi specifika cemi P eh t zp sob nenapraviteln po kozen jednotka Vigtig Information Danish Hvis dette udstyr bruges i en m de ikke specifice ret ved Hoefer Inc den beskyttelse som er blevet forsynet af udstyret kan m ske sv kkes Dette instrument er designet for indend rs labora toriumbrug bare Bare tilbeh r og del godkendede eller forsynede ved Hoefer Inc kan m ske bruges for drive funk tionsfejl og betjening dette produkt bruger Bare en stramforsyning der er CE markerede eller sikkerhed som er blevet attesteret af en som nationalt er blevet anerkendt prove laboratorium Sikkerhedl get m vaere p plads fer forbinding stramforsyningsblyet til en str mforsyning Drejer alle stremforsyningskontroller af og afbryder kraftblyet for fjerning sikkerhedl get Cirkulerer bare vand eller 50 50 vand ethylene glykol gennem varmeveksleren i s fald udrustet Forbind ikke varmeveksleren til en vandhane eller nogen kelemiddelkilde hvor vandtrykket er unregulated Introducerer Aldrig antifreeze eller noget organisk oplesningsmiddel ind i nogen del af instrumentet Organiske opl sningsmidler vil for rsage uboelig skade til enheden Driver ikke med stedpudetemperaturer over maksimummet specificerede tekniske spe
3. 5 turns in the counterclockwise direction Swing the clamps outward o Remove the gel from the sandwich or cassette Gently loosen and then slide away both spacers Slip an extra spacer or the Hoefer Wonder Wedge into the bottom edge to prevent breaking the ears of the notched plates and separate the plates If using precast gels follow gel manufacturer s instructions a i Carefully lift the gel from the plate and lay it into a tray containing stain fixative or transfer buffer Clean the unit as described in Care and mainte nance below Care and maintenance Do not autoclave or heat any part above 75 C Do not immerse the safety lid in any liquid Do not use organic solvents strong or oxidiz ing cleaning solutions abrasives or strong acids or bases on any part of the instrument 0 Immediately after each use rinse the tank and modules with water and then rinse thoroughly with distilled water Handle the module with care to prevent damage to the banana plugs Allow to air dry o Wipe the lid with a damp cloth If necessary briefly rinse the underside of the lid with water e Clean glass plates and spacers with a dilute solution of a laboratory detergent such as RBS 35 then rinse thoroughly with tap and distilled water Glass plates can be treated with but not stored in acid cleaning solutions o Wipe plates with isopropanol to remove any Gel Seal residue
4. a Fig 11 e Secure the cassette Swing each clamp into position over the sides of the cassette Tighten each screw alternating to apply even pressure until the cassette is secure The gasket around the upper buffer chamber should be fully compressed to provide a seal but the screws should not be tightened to the point that pres sure stresses the cassette Fig 11 Securing the cassette Check that both gel surfaces will contact buffer Check that the bottom gel contact slot is exposed o Move the sealing plate into the half open position to prepare for electrophoresis Apply gentle inward pres sure to both tabs and lock them into the lower notch Final assembly 0 Make sure the sealing plate is in the half open posi tion The arrow in Fig 12 indicates the correct position o Lower each module into the tank seating it in the locating slots The module seats properly in only one orientation with the banana plugs toward the center of the tank and the gel facing outward e Add the appropriate amount of electrophoresis buffer to the tank Add 1 2 1 6 liters of buffer to the tank when only one module is in place and 1 1 1 4 liters when two modules are in place e pll Fig 12 Preparing for electrophoresis Tip As an aid in loading samples mark the well location with a labo ratory marking pen or use locating decal The locating decal only works with miniVE c
5. Jos tata varusteita k ytet n tavassa ei m ritetty Hoefer Inc suojelu ehk isty varusteille saattaa olla avuton T m valine suunnitellaan sis laboratoriok ytolle vain Vain lis varusteet ja osat hyv ksyiv t tai toimitti Hoefer Inc oheen voi k ytt k ytt miselle valvoalle ja servicing t m tuote Vain k ytt k ytt j nnitett joka on CE merkitsi tai turvallisuus joka on todistanut aidoksi ohi joka on kansallisesti tunnustettnut testaaminen labo ratoriota Turvallisuuskansi t ytyy olla paikallaan ennen yhdist minen k yttoj nnitelyijyj k ytt j nnit teeseen Kiert kaikki k yttoj nnitevalvonnat ja irrottaa valtalyijyt ennen poistaminen turvallisuuskantta Kiert vain vesi tai 50 50 vesi ethylene glycol siin tapauksessa varustetun l mm nvaihtimen l pi l yhdist l mm nvaihdinta vesinapautuk seen eik j hdytysnestel hteeseen miss vesi paine on unregulated Pakkasneste eik orgaaninen liuotin v lineen osassa ei esitele Koskaan Orgaaniset liuottimet aiheuttavat korvaamattoman vahingon yksikk n Ei k yt puskuria yll olevia l mp tiloja enint n m ritetyill teknisill t smennyksill Ylikuumen eminen aiheuttaa korvaamattoman vahingon yksikk n Information Importante French Si cet quipement est utilis dans une mani re pas sp cifi par Hoefer Inc la protection fourni par l quipement pourrait t
6. One gel can be cast in place on each elec trophoresis module A wide range of accessories ordered separately see page 27 lends the miniVE a high degree of versatility These include a large selection of combs and spacers a blot module to convert the miniVE into a mini blotting unit See page 18 for instructions Fig 1 Main components of the Hoefer SE300 miniVE color coded leads connect electrodes to power supply lid electrophoresis module banana plug connectors Unpacking Unwrap all packages carefully and compare contents with the packing list making sure all items arrived f any part is missing contact your local sales office Inspect all components for damage that may have occurred while the unit was in tran sit If any part appears damaged contact the carrier immediately Be sure to keep all packing material for damage claims or for repacking should it become necessary to return the unit Prior to use wash the tank and module with a dilute solution of non abrasive laboratory detergent Thoroughly rinse first with water and then with distilled water sm Specifications Electrophoresis Gel sandwich size 10 0 cm wide x 8 to 10 5 cm long ax tank volume 1 6 liters with one module in place 1 4 liters with two modules in place ax voltage 600 V ax wattage 25 W per electrophoresis module Electrotransfer ax volume blot module 35
7. and three packing sponges The gel determines the size of the membrane and filter paper 0 For each gel cut the membrane and two pieces of filter paper the same size as the gel but no larger than 8 5 x 10 5 cm o Equilibrate the gel in transfer buffer for 10 minutes Equilibration allows the gel to swell or shrink before it contacts the transfer membrane and removes excess buffer salts and detergents from the gel Longer equil ibration may result in diffuse bands e Pre wet nitrocellulose or nylon membranes in distilled water taking care not to trap air bubbles Dip one end of the membrane into the buffer and slowly submerge it allowing it to wet by capillary action Pre wet PVDF or other hydrophobic membranes in methanol After pre wetting soak all membrane types in transfer buffer for 2 5 minutes o Wet the two pieces of filter paper in transfer buffer Q Fig 15 Assembling the transfer stack Assemble the transfer stack so that molecules will migrate to the membrane Fig 15 For negatively charged macromolecules such as proteins run in an SDS gel and nucleic acids assemble the transfer stack on the black cathode side Proteins will transfer towards the red anode side red anode black cathode Note For best results avoid trapping air bubbles as each layer is applied Always establish full contact along one side and maintain
8. 8999 Fax 1 508 893 0176 E mail support hoeferinc com Web www hoeferinc com Hoefer is a registered trademark of Hoefer Inc Coomassie is a trademark of ICI plc Dacron is a trademark of E I duPont Nemours amp Co RBS 35 is a trademark of Pierce Chemical Company 2012 Hoefer Inc All rights reserved Printed in the USA Hoefer
9. contact as the layer is lowered into position a Center a packing sponge on the black cathode side b Lay one piece of wet filter paper on the sponge C Position the equilibrated gel on the filter paper Wet the gel surface with a few drops of transfer buffer d Lay the membrane on the gel Do not reposition the membrane once it contacts the gel Use a glass rod to roll out any air bubbles e Lay one piece of wet filter paper on the membrane f Lay two packing sponges on the filter paper A second transfer stack if added is placed between these two sponges Repeat steps b e e p21 Fig 16 Final assembly cathode side faces the center red anode side faces the outside tank wall e p22 o Check the position of the transfer stack The transfer stack should be centered on the elec trode plate No layer should be pinched when the module is closed o Fold the empty half of the cup over the stack and press the halves together to snap the module closed The transfer stock should be held firmly in place when the cup is closed Replace old and compressed sponges if needed to fill the cup Final assembly 0 Slowly pour 300 350 ml of transfer buffer into the top of the module to allow air to be displaced by the buffer as it fills the cup Tap the blotting cup lightly to dislodge any air bubbles in the packing sponges o Position the module s in the tank with the banana plug
10. membranes parameters Store membranes properly Protect them from temperature extremes and direct sunlight If proteins pass through the selected membrane try a different type or one with a smaller pore size 0 10 0 20 um If different proteins may be migrating in opposite directions place a membrane on both sides of the gel If the sample load may be exceeding the capacity of the binding surface area apply two membranes If blow through occurs reduce the sample load e p25 E Bibliography Polyacrylamide gel electrophoresis Adams L D and Gallagher S R 1992 Two Dimensional Gel Electrophoresis Using the O Farrell System Current Protocols in Molecular Biology 10 4 1 10 4 13 Gallagher S R and Smith J A 1991 Electrophoretic separation of proteins Current Protocols in Molecular Biology EA Ausubel R Brent R E Kingston D D Moore J G Seidman J A Smith and K Struhl eds 10 2 1 10 2 21 Laemmli U K 1970 Cleavage of structural proteins during the assembly of the head of bacteriophage T4 Nature 227 680 685 Matsudaira P T and Burgess D R 1978 SDS microslab linear gradient polyacrylamide gel electrophoresis Anal Biochem 87 386 396 Reisfeld R A et al 1962 Acidic buffer system for resolu tion of cationic proteins Nature 195 281 Sasse J and Gallagher S R 1991 Staining proteins in gels Current Protocols in Molecular Biology F A Ausubel R Brent R E Kingston D
11. 0 3 6 4 8 72 12 4 75 15 2 2 2 9 44 18 2 9 Electrical connections 0 Position the safety lid over the unit and seat the lid so the banana plugs engage the jacks in the lid The lid is symmetrical and fits in either orientation Fig 13 o Plug the color coded leads into the jacks of an approved power supply red to red black to black The minimum power supply rating is 250 V 50 mA constant current or constant voltage Recommended power supply PS300B e p13 e pl4 Electrophoresis For optimal resolution start electrophoresis immediately after sample loading Gels may be run at either constant current or constant voltage For Laemmli SDS separations the recommended voltage range is 100 250 V and should not exceed 300 V If running gels at constant current the current should be 10 20 mA per gel depending on gel thickness 10 mA for 0 75 mm 15 mA for 1 5 mm Check progress after 5 minutes and again after half an hour monitoring the position of the tracking dye The run is complete when the tracking dye reaches the bottom of the gel After electrophoresis 0 Turn off the power supply and disconnect the leads a Remove the safety lid and lift out the module s Release each gel sandwich or cassette from the module Move the sealing plate to the fully open position by pressing inward on both tabs and guiding the plate to open out Then loosen all four screws 4
12. 0 ml per module ax tank volume 1 7 liters with one module in place for passive cooling 1 2 liters with two modules in place ax wattage 15 W per blot module ax current 400 mA SE300 miniVE specifications ax operating temp 75 C Chemical compatibility For use only with dilute aqueous solutions between pH 2 and pH 12 Not compatible with organic solvents or concentrated alcohols acids bases and oxidizing agents Environmental operating Indoor use 4 40 C conditions Humidity up to 80 Altitude up to 2000 m Installation category II Pollution degree 2 Dimensions wx dx h 19 2 x 17 2 x 18 8 cm 7 6 x 6 8 x 7 4 in Weight tank lid 1 2 kg 2 65 Ibs and two gel modules Product certifications EN61010 1 UL61010A 1 CSA C22 2 1010 1 CE Certified This declaration of conformity and the warranty are only valid when the instrument is used in laboratory loca tions within the conditions specified in the user manual as delivered from Hoefer Inc except for alterations described in the user manual and connected to other CE labeled instru ments or products recommended or approved by Hoefer Inc Note The sealing plate has three positions closed or sealed for casting half open for electropho resis and fully open for placing the gel sandwich Fig 2 Module in the closed position ep The Electrophoresis module This section describes the use of the electropho resis module For instr
13. D Moore J G Seidman J A Smith and K Struhl eds 10 6 1 10 6 8 Weber K and Osborn M 1969 The reliability of molecu lar weight determinators by dodecyl sulfate polyacryl amide gel electrophoresis J Biol Chem 224 4406 4412 Blotting Gallagher S Winston S E Fuller S A and Hurrell J G R 1993 Immunoblotting and Immunodetection in Current Protocols in Molecular Biology 10 8 1 10 8 17 Greene Publishing and Wiley Interscience NY Gershoni J M and G E Palade 1983 Protein Blotting Principles and Applications Anal Biochem 131 1 15 Harlow E and Lane D 1988 Antibodies A Laboratory Manual Cold Spring Harbor Press Sambrook J et al 1989 Molecular Cloning A Labora tory Manual Cold Spring Harbor Laboratory Press B 23 Towbin H Staehelin T and Gordon J 1979 Electro phoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets procedure and some applications Proc Natl Acad Sci USA 76 4350 4354 e p26 Ordering information product quantity code no Hoefer SE300 miniVE complete 1 SE300 10A 1 0 Includes Basic unit 3 rectangular glass plates 3 notched plates 2 each 1 0 mm thick 10 well combs and 1 0 mm thick spacer sets Blot Module 1 SE302 Includes 3 Dacron packing sponges D i thick 25 sheets of blotter paper Accessories Glass plates 10 x 10 5 cm 5 pk
14. Qus Hoefer SE300 miniVE Electrophoresis and Electrotransfer Unit um SE300 IM Rev C0 06 12 H O e fe r Contents Important Information 2 cintia ii Waste Electrical and Electronic Equipment WEE vii eet He de en 1 UNPACKING ege egeg csse s tesa aso eege o voie cnp rr ase n doba 2 ee e EE 3 The Electrophoresis Module 4 Electropli r68iS u inienn ssasaptsiserstacianepamespatandiae 14 Care and maintenance eren 15 Electrophoresis troubleshooting 16 The blot lune UE 18 Blotter care and maintenance r 23 Blotter troubleshootiNE lt lt 24 Biblicas US 26 Ordering Information 27 Important Information English If this equipment is used in a manner not speci fied by Hoefer Inc the protection provided by the equipment may be impaired This instrument is designed for indoor laboratory use only Only accessories and parts approved or supplied by Hoefer Inc may be used for operating main taining and servicing this product Only use a power supply that is CE marked or safety certified by a nationally recognized testing laboratory The safety lid must be in place before connecting the power supply leads to a power supply Turn all power supply controls off and disconnect the power leads before removing the safety lid Circulate only water or 50 50 water ethylene glycol through the heat exchanger if so equipped Do not connect the heat exchanger t
15. SE262P 5 Notched glass plates 10 x 10 5 cm 5 pk SE262GN 5 Spacers 0 75 mm thick pair SE2619T 2 75 Spacers 1 0 mm thick pair SE2619T 2 1 0 Spacers 1 5 mm thick pair SE2619T 2 1 5 Comb 5 well 0 75 mm thick 1 SE211A 5 75 Comb 5 well 1 0 mm thick SE211A 5 1 0 Comb 5 well 1 5 mm thick SE211A 5 1 5 Comb 9 well 1 0 mm thick microtiter SE211A 9 1 0 Comb 10 well 0 75 mm thick SE211A 10 75 Comb 10 well 1 0 mm thick SE211A 10 1 0 Comb 10 well 1 5 mm thick SE211A 10 1 5 Comb 12 well 1 0 mm thick SE211A 12 1 0 Comb 15 well 0 75 mm thick SE211A 15 75 Comb 15 well 1 0 mm thick SE211A 15 1 0 Comb 15 well 1 5 mm thick SE211A 15 1 5 Comb 18 well 1 0 mm thick microtiter SE211A 18 1 0 Comb prep ref 0 75 mm thick SE211A R 75 Comb prep ref 1 0 mm thick SE211A R 1 0 Comb prep ref 1 5 mm thick SE211A R 1 5 Gasket cord 100 cm FH2208 e p27 product quantity code no Blotting accessories Sponge Dacron packing 3 pk SE3005 Blotting paper 9 10 5 cm 50 pk TE26 Gel casters Hoefer SE235 4 Gel Caster 2 to 4 gels 10 x 10 5 cm SE235 Power supplies PS200HC Power Supply 200 V 2000 mA 200 W PS200HC PS300B Power Supply 300 V 500 mA 90 W PS300B Gel drying system Hoefer Easy Breeze Air Gel Dryer 115 V SE1200 115V Hoefer Easy Breeze Air Gel Dryer 230 V SE1200 230V e p28 Hoefer Inc 84 October Hill Road Holliston MA 01746 Toll Free 1 800 227 4750 Phone 1 508 893
16. all apparecchiatura potrebbe essere indebolita Questo strumento disegnato per l uso di labora torio interno solo Solo gli accessori e le parti hanno approvato o hanno fornito da Hoefer Inc potrebbe essere usato per operare per mantenere e per revisionare questo prodotto usa Solo un alimentatore che CE ha marcato o la sicurezza certificato da un nazionalmente ricon osciuto testando il laboratorio Il coperchio di sicurezza deve essere nel luogo prima di collegare i piombi di alimentatore a un alimentatore Spegne tutto i controlli di alimentatore e disin serisce i piombi di potere prima di togliere il coper chio di sicurezza Circola solo l acqua o 50 50 glicole di acqua etilene attraverso lo scambiatore di calore se cosi equipag giato Non collegare lo scambiatore di calore a un rubinetto di acqua o qualunque fonte di refriger ante dove la pressione di acqua sregolata Non introduce mai l antigelo o qualunque solvente organico in qualunque parte dello strumento solventi organici causeranno il danno irreparabile all unit Non opera con le temperature di tampone al di sopra del massimo ha specificato le descrizioni tecniche II surriscaldamento causera il danno irreparabile all unit Viktig Informasjon Norwegian Hvis dette utstyret blir brukt i en m te ikke spesi fisert ved Hoefer Inc beskyttelsen som ha blitt git av utstyret kan bli svekket Dette instrumentet er utform
17. along the glass plate edges on the sides of the sandwich Fig 4 Important Proper alignment is essential to prevent leakage Note Once the sandwich is care fully aligned hold the flat sides firmly between your thumb and fingers near the notch Fig 5 Placing the sandwich in the module Fig 6 Positioning the clamps e p6 Place and seal the sandwich on the module 0 Take care to square the three sealing sides of the sandwich Hold the sandwich like a deck of cards and gently tap the bottom against a flat surface o Notched plate side down lay the sandwich on the module Fig 5 Fit the gel sandwich within the guides at both sides a and against the guide feet at the bottom b notch at top While gently holding the sandwich against the module swing one clamp into position over the spacer taking care not to bump the sandwich out of alignment Fig 6 Important Check the alignment of the bottom edge of the sandwich against the guide feet Fig 7 Fig 7 Check the bottom edges for proper alignment guide foot Fig 8 Misalignments cause leaks o Turn each screw alternating to keep the pressure even until the clamps are loosely secured and will allow the spacers to be adjusted if necessary Repeat on the other side If the spacers and glass plates are not perfectly aligned against the stops use the stiff end of the Hoefer Wonder Wedge to press against the ed
18. cifica tions Overheding vil for rsage uboelig skade til enheden e pii Belangrijke Informatie Dutch Indien deze uitrusting in een manier wordt gebruikt die niet door Hoefer Inc is gespecificeerd de bescherming die door de uitrusting is verzorgd kan worden geschaad Dit instrument is voor binnenlaboratoriumgebruik enkel ontworpen Enkel onderdelen en delen keurden goed of leverden door Hoefer Inc kan voor het bedienen worden gebruikt handhavend en onderhouden van dit product gebruik Enkel een netvoeding die CE is markeerde of veiligheid die door een is gecertificeerd die nationaal is herkend testene laboratorium Het veiligheidsdeksel moet in plaats voor het verbinden van de netvoeding leidt tot een netvoeding zijn Doe alle netvoedingscontroles Uit en koppel los de machtleiding voor het verwijderen van het veiligheidsdeksel Circuleer enkel water of 50 50 water ethyleen glycol door de hitte exchanger zo ja uitrust Verbind de hitte exchanger naar een waterkraan of koelmiddelbron niet waar de waterdruk niet geregulariseerd is Stel Nooit antivriesmiddel of organische oplosmid delen in deel van het instrument voor Organische oplosmiddelen zullen onherstelbare schade aan de eenheid veroorzaken Bedien niet met buffertemperaturen boven het maximum specificeerde technische specificaties Oververhittend zal onherstelbare schade aan de eenheid veroorzaken T rke Tietoa Finnish
19. ctar la alimentaci n lleva a una alimentaci n Apaga todos controles de alimentaci n y desco necta los plomos del poder antes de quitar la tapa de la seguridad Circula s lo agua o 50 50 glicol de agua etileno por el intercambiador de calor si se es el caso equiparon No conecte el intercambiador de calor a un toque de la agua ni cualquier fuente del l quido refrigerante donde la presi n del agua est libre Nunca introduce anticongelante ni alg n solvente org nico en cualquier parte del instrumento Los solventes org nicos causar n da o irreparable a la unidad No opera con temperaturas de b fer encima del m ximo especific especificaciones t cnicas Reca lentar causar da o irreparable a la unidad Viktig Information Swedish om denna utrustning anv nds i ett s tt som inte har specificeras av Hoefer Inc skyddet tillhan dah ll vid utrustningen kan skadas Detta instrument formges f r inomhuslaborato rium anv ndning bara Bara medhj lpare och delar godk nde eller lever erade vid Hoefer Inc kan anv ndas f r fungera underh lla och servicing denna produkt anv nder bara en kraft tillg ng som r CE markerade eller s kerhet intygade vid en nationellt erk nd testande laboratorium S kerheten locket m ste vara p platsen f re koppla kraften tillg ngen blyen till en kraft tillg ng Vander sig alla kraft tillg ng kontroller av och kopplar bort kraften bly
20. en f re flytta s kerheten locket Cirkulerar bara vatten eller 50 50 vatten ethylene glycol genom v rmen exchanger i s utrustad fall Inte kopplar v rmen exchanger till en vatten kran eller n got kylmedel k lla d r vattnet trycket r unregulated Inf r aldrig kylv tska eller n got organiska l sningsmedel in i n gon del av instrumentet Organiskt l sningsmedel ska orsaka irreparable skada till enheten Anv nd inte med buffert temperaturer over det h gsta angivna tekniska specifikationerna verhettning skulle orsaka irreparabla skador p enheten e pvi English EXE French X German X pon Italian M zm Spanish La Swedish X Waste Electrical and Electronic Eguipment WEEE This symbol indicates that the waste of electrical and electronic eguipment must not be disposed as unsorted municipal waste and must be collected separately Please contact an authorized representative of the manufacturer for information concerning the decommissioning of your eguipment Ce symbole indique que les d chets relatifs l quipement lectrique et lectronique ne doivent pas tre jet s comme les ordures m nag res non tri es et doivent tre collect s s par ment Contactez un repr sentant agr du fabricant pour obtenir des informations sur la mise au rebut de votre quipement Dieses Symbol kennzeichnet elektrische und elektronische Ger te die nicht mit dem gew hnlichen unsor
21. entrated zone in the stacking gel Pour a taller stacking gel For best results allow a stacking gel height of 2 5 times the height of the sample in the well Dispose of outdated acrylamide solutions and use only the highest grade of acrylamide When preparing samples avoid using solutions with a high sodium or potassium concentration e p17 Fig 14 Install a gasket into each cup half e p18 The blot module The Hoefer miniVE Blot Module which can be ordered separately performs electrotransfers on mini format gels Each module holds up to two gels 8 2 cm wide and up to 10 4 cm long One or two modules can be run at the same time Assembly 0 Prior to use wash the tank and blot module with a dilute solution of non abrasive laboratory detergent Thoroughly rinse with water and distilled water o Separate out two of the four strands of gaskets included with each module e Open the module by releasing both tabs o Lay a gasket along the entire groove around three sides of each cup half Avoid stretching or twisting the gasket the length should just fit Gently press into place Each gasket fits into the groove around three sides of each cup half Preparation Optional Passive cooling Chill approximately 2 liters of deionized water to 4 C Filling the tank with chilled water serves as a heat sink during electrotransfer Prepare transfer buffer Stac
22. equired for good binding to nitrocellulose membranes Increase the length of time DNA blots are depurinated Check the buffer pH Most buffers should not be titrated make fresh buffer For native gels increase the net charge on the protein by chang ing to a transfer buffer with a different pH Lower pH 6 7 increases the positive charge on proteins higher pH 26 7 increases the negative charge on proteins Diffuse band Conduct the electrotransfer immediately after electrophoretic patterns separation Shorten or eliminate the equilibration step before electrotransfer or conduct equilibration in the cold room If the transfer buffer contains methanol 210 equilibrate the gel for 30 minutes to allow it to shrink fully Note Gel shrinkage may slow the migration of large molecules out of the gel Take care that the gel does not shift once it contacts the membrane Check that any preferred binding surface of the membrane faces the gel e p24 problem possible cause solution Inefficient Chemical Fix or crosslink the mole to the requirements of the nucleic acid binding parameters protein or membrane type Prepare protein transfer buffer without SDS SDS can improve transfer efficiency but reduces binding Verify the optimal amount of methanol required for the membrane type and check the buffer solution Add 10 20 methanol to the transfer buffer to enhance binding to nitrocellulose Membrane Wear gloves when handling
23. et for innenders labo ratoriumbruk bare Bare tilbehor og deler godkjente eller forsynte ved Hoefer Inc kan bli brukt for drive vedlikeholde og betjene dette produktet bruker Bare en kraftforsyning som er CE merket eller sikkerhet som ha blitt sertifisert av et som nasjonalt ha blitt anerkjent prover laboratorium e piv Sikkerheten lokket ma v re p plass for forbinding kraftforsyningene blyene til en kraftforsyning Vender all kraftforsyningsstyring av og frakopler kreftene blyene f r fjerning sikkerheten lokket Sirkulerer bare vann eller 50 50 vann ethylene glykol gjennom oppvarmingen veksleren i s fall utstyrer Ikke forbind oppvarmingen veksleren til en vanntapp eller noe kj lemiddelkilde hvor vannet trykket er unregulated Introduserer Aldri antifreeze eller noe organisk l semiddel inn i noe del av instrumentet Organi ske l semiddler vil for rsake irreparabel skade p enheten Driver med buffertemperaturer over maksimum ikke spesifiserte teknisk spesifikasjoner overop pheting vil for rsake irreparabel skade p enheten Wazne Informacje Polish Je eli ten sprz t jest wykorzystywany w spos b nie okre lone przez Hoefer Inc do ochrony przewid zianej przez urz dzenie mo e zosta obni ony Instrument ten jest przeznaczony do u ytku w laboratoriach kryty tylko Tylko akcesori w i cz ci zatwierdzone lub dostarc zone przez Hoefer Inc mog by wykorzystane do eksploatacj
24. eurotoxin Always wear gloves and observe all laboratory safety procedures Note Approximately 10 ml of monomer solution is required to cast one 1 mm thick gel A 1 cm stacking gel below the wells Fill to 3 cm below the top of the rectangular glass plate Overlay each gel with a thin layer of water saturated n butanol water or diluted gel buffer to prevent exposing the monomer solu tion to oxygen Use a glass syringe fitted with a 22 gauge needle to apply 100 pl of the overlay solution slowly to one side of the sandwich near the spacer Allow the solution to flow across the surface unaided After polymerization Allow a minimum of one hour for the gel to polymerize o If a comb is in place remove it by carefully pulling on the comb while gently rocking it back and forth to break the vacuum Rinse the wells with electrophore sis buffer to remove any unpolymerized acrylamide If an overlay was applied rinse the sandwich several times with double distilled water to remove it Invert the module to drain To ensure seamless contact between the resolving and stacking gels remove residual liquid by blotting one corner of the gel with a lint free tissue Casting the stacking gel Prepare the stacking gel monomer solution o Deaerate the stacking gel monomer solution add catalyst and initiator and then pour Use a pipette to deliver the solution into one corner of the plate taking care not to trap any bubble
25. ges of the spacer and glass plates and position them flush against the guide foot Complete clamping by tightening each screw firmly hand tight Do not overtighten as the plates may crack Check the spacer alignment The spacer must The glass plate should not not protrude out of be resting on the head of the the sandwich or be spacer T recessed into it Fig 9 Assembled module with tabs engaged in top notch Note To test for alignment pass a corner of the Wonder Wedge across the bottom edge of the spacers and glass plates If an edge catches realign Check both sides Fig 10 Hang the module on the narrow side of the tank to pour the gel e p8 Lock the sealing plate into the closed position by engaging each tab in its topmost notch Fig 9 tabs engaged in top notch Hang the module from the narrow side of the tank or stand it on the benchtop to cast the gel Fig 10 When hanging the module on the tank either fill the tank or hang the second module on the other side as a counterbalance Pouring the resolving gel 0 Prepare the monomer solution o Pipet the solution into the sandwich slowly so that it flows along a spacer taking care not to trap any air pockets No stacking gel Fill the solution to the desired level and then insert a comb at a slight angle into the sand wich taking care not to trap air under the teeth Warning Acrylamide is a n
26. hutz verschlechtert werden Dieses Instrument wird f r den Innenlaborge brauch nur daf r entworfen Nur Zus tze und Teile genehmigten oder lieferten durch Hoefer Inc kann f r das Funktionieren das Aufrechterhalten und die Wartung dieses Produk tes verwendet werden Verwenden Sie nur eine Energieversorgung die CE gekennzeichnet oder durch ein national anerkanntes Probelaboratorium bescheinigte Sicherheit ist DerSicherheitsdeckel muss im Platz vor dem AnschlieBen der Energieversorgung sein f hrt zu einer Energieversorgung Alle Energieversorgungssteuerungen abdrehen und die Macht trennen f hrt vor dem Entfernen des Sicherheitsdeckels Nur Wasser oder 50 50 Glykol des Wassers Athylens durch den W rmeaustauscher wenn so ausgestattet in Umlauf setzen Verbinden Sie den W rmeaustauscher mit einem Wasserklaps oder jeder K hlmittel Quelle nicht wo der Wasserdruck ungeregelt wird F hren Sie nie Frostschutzmittel oder jedes organische L sungsmittel in jeden Teil des Instru mentes ein Organische L sungsmittel werden nicht wiedergutzumachenden Schaden der Einheit verursachen Mit Puffertemperaturen ber angegebenen technischen Spezifizierungen des Maximums nicht funktionieren Die berhitzung wird nicht wiedergutzumachenden Schaden der Einheit verursachen Informazioni Importanti Italian Se quest apparecchiatura usata in un modo specificato da Hoefer Inc la protezione fornito d
27. i utrzymania i obs ugi tego produktu korzysta jedynie zasilacza e jest nosz ce ozna kowanie CE lub bezpiecze stwa uwierzytelnione przez uznane na poziomie krajowym laboratorium badawcze Bezpiecze stwo lid musi by w miejsce przed pod czeniem zasilania prowadzi do zasilania Za wszystkie r d a zasilania urz dzenia steruj ce off i od czy moc prowadzi przed odbiorem bezpiecze stwa lid Kr tylko wody lub wody 50 50 ethylene glycol wymiennik ciep a poprzez je li tak wyposa one Nie nale y po czy wymiennik ciep a woda z kranu lub jakimkolwiek chtodziwo r d a je eli ci nienie wody jest nieuregulowanych Nigdy nie wprowadza rozpuszczalnika organ icznego przeciw zamarzaniu lub jakichkolwiek na dowoln cz dokumentu Rozpuszczalniki organiczne spowoduje nieodwracalne szkody dla jednostki Nie dzia aj w buforze temperatury powy ej maksymalnego okre lone specyfikacje techniczne Przegrzania spowoduje nieodwracalne szkody dla jednostki Informa es Importantes Portuguese Se este equipamento usado numa maneira n o especificada por Hoefer Inc que a protecc o forne cida pelo equipamento pode ser comprometida Este instrumento projectado para uso de interior de laborat rio s S6 acess rios e partes aprovaram ou forneceu por Hoefer Inc pode ser usada para operar manter e servicing este produto S usa um estoque de poder que CE marcou ou seg
28. k assembly requires approximately 250 ml transfer buffer and an additional 300 350 ml buffer is required to fill each module The recipe for Towbin buffer is listed below The Bibliogra phy on page 26 lists sources for other buffers Towbin buffer 25 mM Tris 192 mM glycine 0 20 v v methanol pH 8 3 1 liter Tris FW 121 1 25 mM 3 0g Glycine FW 75 07 192 mM 14 4g SDS FW 288 4 up to 0 1 3 5 mM 1 0g Optional Adding SDS can improve transfer efficiency 0 Dissolve in 750 ml distilled water o Add methanol as required Depending on the membrane type selected adding methanol can improve the transfer results Because buffers containing methanol may deteriorate if stored for long periods add methanol just prior to transfer e Bring to 1 liter with distilled water Do not adjust the pH which should be between 8 2 and 8 4 Optional Chill before use e p19 Important Try to place the gel correctly the first time Proteins may begin to transfer immediately Once transfer begins moving the gel will distort results or cause shadow bands on the blot e p20 Prepare the transfer stack Transfer the sample as soon as possible after electrophoresis to minimize sample diffusion within the gel Electrophoretic transfer can be performed on as many as four mini gels at one time if two gels are placed in each of two modules The transfer stack consists of the gel and membrane filter paper
29. le Remove the gels and membranes Save the packing sponges Discard the blotting paper e Label each membrane and indicate the sample side Lift the membrane s with blunt forceps and allow to air dry o Rinse the unit immediately after use Blotter care and maintenance Do not autoclave or heat any part above 75 C Do not use organic solvents strong or oxidizing cleaning solutions abrasives or strong acids or bases on any part of the instrument mmediately after each use rinse the unit with water and then rinse thoroughly with distilled water Handle the module with care to prevent damage to the electrode plugs Allow to air dry e p23 Blotter troubleshooting problem possible cause solution Incomplete Blank areas on Remove all trapped air bubbles in the transfer stack take espe transfer the membrane cially great care during stack assembly to prevent air bubbles from forming as each layer is placed Check electrode continuity Use a lower ionic strength buffer Molecules do Increase the field strength not migrate out M Increase the transfer period Try doubling it of gel Do not expose the gel to staining or fixing agents before transfer Use a thinner gel Reduce the gel acrylamide concentration For proteins use 3 5 mM SDS 0 1 in the transfer buffer Decrease the methanol in the protein transfer buffer or reduce the amount to a minimum Typically 1096 methanol is r
30. o a water tap or any coolant source where the water pressure is unregulated Never introduce antifreeze or any organic solvent into any part of the instrument Organic solvents will cause irreparable damage to the unit Do not operate with buffer temperatures above the maximum specified technical specifications Overheating will cause irreparable damage to the unit Dule it Informace Czech Pokud by toto za zen je pou ito zp sobem kter nen podle Hoefer Inc ochrana poskytovan na z klad za zen m e b t naru ena Tento n stroj je ur en pro vnit n pou it v laborato i pouze Pouze p slu enstv a sti schv len nebo poskyt nut ch Hoefer Inc mohou b t pou ity pro provoz dr bu a dr b tohoto v robku zdroj nap jen pou vaj jen e je opat en ozna en m CE osv d ena nebo bezpe nost vnitrost tn uznan mi zku ebn mi laborato Bezpe nosti lid mus b t zavedena p ed p ipojen m nap jec zdroj nap jen vede k Turn ve ker nap jen kontroly vypnuto a odpojit p ed odb rem energie vede bezpe nostn v ko Rozeslat pouze voda nebo 50 50 voda ethyleng lykolu prost ednictv m v m n k tepla je li to vybav ena Nemaj p ipojen v m n k tepla s vodn mi set epn nebo jak koli chladic kapaliny zdroje kde tlak vody je neregulo Nikdy zav st prost edek proti zamrznut nebo jak koli organick rozpou t dla do jak koli
31. ombs and not precast gels e p12 the half open postion for electrophoresis upper buffer chamber The minimum and maximum levels are marked Verify that the lower electrode which is approximately 2 cm from the bottom of the module is completely submerged To prevent buffer from entering the upper buffer chamber verify that the buffer level is not above the maximum level o Add the appropriate amount of electrophoresis buffer to the upper buffer chamber Fill the upper buffer chamber to a level 3 5 mm above the notched plate This reguires approximately 100 ml e Prepare and apply the sample Increase liquid sample density with 10 glycerol or sucrose Add a tracking dye such as bromophenol blue Note The amount of protein sample added to each well depends on both the sensitivity of the staining method and the distri bution of protein among separate bands With Coomassie Blue it is possible to detect 1 ug in a single band with the more sensi tive silver stains it is possible to detect as little as 10 ng Fig 13 Fully assembled miniVE with electrophoresis module Underlay the sample into the wells using a micro pipet or fine tipped microsyringe Table 1 shows the volume of sample required for different numbers of wells and comb thicknesses Table 1 Well capacities volume of sample yl per 1mm depth comb thickness mm no of wells 0 75 1 0 1 5 9 5 12 7 19 1 5 8 1
32. ore loading to remove particulates problem solution Stained sample collects Near the buffer front Protein is not sufficiently restricted by the resolving gel increase the T Near the top of the gel when the buffer front has reached the bottom The gel pore size is too small Decrease the T of the resolving gel The protein has precipitated Heat the sample at a lower temperature 70 C or less for 1 2 minutes Poor band resolution Use only the highest quality reagents Conduct the separation at a lower current or voltage setting Dialyze or desalt the sample Reduce the sample volume or concentration Only use freshly deionized urea Improve dissociation of subunits by heating sample in SDS sample buffer 1 2 minutes at 100 C Add more mercaptoethanol or dithiothreitol check sample treatment Only use gels that were recently prepared Check pH values of the separating and stacking gel solutions Do not back titrate buffers Sample preparation e Heat samples for no more than 1 2 minutes at 100 C Store on ice after heating Store sample on ice before it is denatured Add protease inhibitors if necessary to prevent proteolytic degradation of sample Store samples to be frozen in aliquots to prevent repeated freezing and thawing Store at 40 to 80 C Bromophenol blue doesn t sharpen into a conc
33. re diminu e Cet instrument est concu pour l usage de labora toire int rieur seulement Seulement les accessoires et les parties ont approuv ou ont fourni par Hoefer Inc pourrait tre utilis pour fonctionner maintenir et entrete nir ce produit utilise Seulement une alimentation qui est CET a marqu ou la s curit certifi par un nationale ment reconnu essayant le laboratoire Le couvercle de s curit doit tre sa place avant connecter l alimentation mene une alimentation Tourner tous contr les d alimentation de et d brancher les avances de pouvoir avant enlever le couvercle de s curit Circuler seulement de l eau ou 50 50 glycol d eau thyl ne par l exchanger de chaleur si si quip Ne pas connecter l exchanger de chaleur un robinet d eau ou la source d agent de refroidissement o la pression d eau est non r gul e Ne Jamais introduire d antigel ou du dissolvant organique dans n importe quelle partie de e piii l instrument Les dissolvants organiques causeront des dommages irr parables l unit Ne pas fonctionner avec les temp ratures de tampon au dessus du maximum a sp cifi des sp cifications techniques La surchauffe causera des dommages irr parables l unit Wichtige Informationen German Wenn diese Ausr stung gewisserma en nicht angegeben durch Hoefer Inc verwendet wird kann der durch die Ausr stung zur Verf gung gestellte Sc
34. s Note If the gel has wells skip to Final assembly on page 11 Tip To calculate the volume measure the distance in centime ters from the top of the resolving gel to the notch in the glass plate This should be at least 2 cm Multiply this distance by the gel width 8 cm and the gel thickness cm for the required volume ml Note To aid in sample loading mark the well locations with a laboratory marking pen e plo Insert a comb at a slight angle to prevent trapping air into the sandwich allowing the comb sides to rest on the spacers o Allow a minimum of one hour for the gel to polymerize Working with precast gels 0 Prepare the electrophoresis module as described in Preparing the module on page 4 Follow the manu facturer s instructions to prepare the gel for electro phoresis This may involve removing tape or breaking off the sealing edge from the bottom of the cassette o Remove the comb and rinse the wells with electropho resis buffer to remove any unpolymerized acrylamide e If the gel is ready for electrophoresis move the sealing plate into the half open position Apply gentle pressure to both tabs and lock them into the lower notch o Position the cassette on the module Orient the cassette so that the notched side is against the gasket and the wells are at the top of the module Center the cassette within the guiderail at both sides of the module
35. s toward the center the red side facing outward e Add deionized water to the tank 1 7 liters for one module and 1 2 liters for two modules To avoid rapid evaporation buffer temperature should not exceed 75 C Passive cooling is recommended if the transfer will be longer than one hour if biological activity must be retained or if transferring nucleic acids Chill deionized water to 4 C before adding to the tank o Place the safety lid on the tank Either orientation fits and is correct Plug the color coded leads into the jacks of an approved power supply such as the PS300B or PS200HC red to red black to black Important Buffer conductivity increases with increasing temperature providing a positive feedback that results in rapid heating We recommend programming the power supply to hold the current setting constant to avoid possible overheating especially if no passive cooling is in place If the only programming option is to hold the voltage setting constant monitor and adjust the voltage to maintain the current at or below 400 mA Electrotransfer Electrophoretic transfer conditions for blotting proteins in Towbin buffer 25 V for 1 2 hours 300 400 mA After electrotransfer Turn off the power supply and disconnect the leads o Remove the safety lid e Lift out each module and drain it by inverting it over a sink Avoid wetting the banana plugs with buffer o Open the modu
36. tierten Hausm ll entsorgt werden d rfen sondern separat behandelt werden m ssen Bitte nehmen Sie Kontakt mit einem autorisierten Beauftragten des Herstellers auf um Informationen hinsichtlich der Entsorgung Ihres Ger tes zu erhalten Questo simbolo indica che i rifiuti derivanti da apparecchiature elettriche ed elettroniche non devono essere smaltiti come rifiuti municipali indifferenziati e devono invece essere raccolti separatamente Per informazioni relative alle modalit di smantellamento delle apparecchiature fuori uso contattare un rappresentante autorizzato del fabbricante Este s mbolo indica que el equipo el ctrico y electr nico no debe tirarse con los desechos dom sticos y debe tratarse por separado Contacte con el representante local del fabricante para obtener m s informaci n sobre la forma de desechar el equipo Denna symbol anger att elektriska och elektroniska utrustningar inte f r avyttras som osorterat hush llsavfall och m ste samlas in separat Var god kontakta en auktoriserad tillverkarrepresentant f r information ang ende avyttring av utrustningen e pvii Introduction Note Minimum power supply The Hoefer SE300 miniVE vertical electropho ratings 50 mA 250 V constant resis system performs vertical gel electrophoresis current or constant voltage on mini format gels The basic unit includes two electrophoresis modules Each module holds one gel sandwich 10 cm wide and up to 10 5 cm long
37. uctions on using the blot module see page 18 The electrophoresis module accepts both self cast and precast gels 8 cm wide from 8 10 5 cm long and 0 75 1 5 mm thick For instructions on using the module with precast gels see page 10 Preparing the module To position the module to accept the gel sand wich each of the three hinged sealing elements must be opened 0 Release the sealing plate by applying gentle inward pressure to both tabs as indicated by the arrows Fig 2 a Holding the tabs move the plate into the fully open position Loosen all four screws 4 5 turns in the counter clockwise direction Do not attempt to remove the screws from the clamps clamp screws 4 apply inward pressure to release sealing plate 0 Fig 3 Module in the open position Fig 4 Gel sandwich assembly To open the module swing the clamps outward clamp 2 Lay the module flat on a work surface Preparing self cast gels One single gel can be cast on the module To cast several gels use a 4 gel caster such as the Hoefer SE235 caster see Ordering information on page 27 Assemble the gel sandwich 0 Prepare the module as described on page 4 o Choose one notched plate one rectangular glass plate and two spacers Use only unchipped plates to prevent leaking e Assemble the gel sandwich with the notch at the top of the sandwich and the spacer ridges align
38. uranca registrada por um nacionalmente recon hecido testando laborat rio A tampa de seguranca deve estar em lugar antes de ligar o estoque de poder leva a um estoque de poder Desliga todos controlos de estoque de poder e desconecta os chumbos de poder antes de retirar a tampa de seguranca Circulam s gua ou 50 50 glicol de gua ethylene pelo exchanger de calor se for assim eguiparam N o ligue o exchanger de calor a uma torneira de gua nem qualquer fonte de refrigerante onde a press o de gua n o regulado Nunca introduz anticongelante nem qualquer org nico solvente em qualquer parte do instru mento Org nico solvente causar agress o irrepar vel unidade N o opera com temperaturas de buffer acima do m ximo especificou especificac es t cnicas Super aguecer causar agress o irrepar vel a unidade Informaci n Importante Spanish Si este equipo es utilizado en una manera no espe cificado por Hoefer Inc la protecci n proporcio nado por el equipo puede ser da ada Este instrumento es dise ado para el uso interior del laboratorio s lo S lo accesorios y partes aprobaron o suministraron por Hoefer Inc puede ser utilizado para operar para mantener y para atender a este producto S lo utiliza una alimentaci n que es CE marc o la seguridad certificada por un nacionalmente reconocido probando el laboratorio Latapa de la seguridad debe estar en el lugar antes de cone
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