Hoefer SE600 Chroma
Contents
1. no of comb thickness mm wells 0 75 1 0 15 10 6 2 8 3 12 4 12 5 8 7 7 11 5 15 4 3 5 7 8 6 20 3 1 4 1 6 2 28 2 1 2 7 4 1 1 1 ref prep 4 90 6 121 9 183 1 2 ref prep 4 85 6 112 9 171 e p17 Fig 6 Attaching gel sandwiches to the upper buffer chamber If the assembly leaks take it to a sink and partially release the cams to allow buffer to drain out of the upper chamber Disassemble check alignment of all sandwich components and adjust if necessary A Remove cams from the lower cam holes Place the upper chamber onto the sandwiches and then insert the cams into the upper cam holes ridge short end pointing down B The final cam position not shown must be vertical so that the assembly fits into the lower buffer chamber e pl8 Final Assembly Upper buffer chamber Rinse both buffer chambers with water and distilled water thoroughly before each use Note Before using the first time disassemble the unit and wash with a dilute solution of a laboratory detergent and rinse thoroughly first with water and then with distilled water Clean away any gel adhering to the exterior of the gel sandwiches a If running only one gel Block the second upper buffer chamber slot by installing the acrylic buffer dam included with the unit Fit clamps onto the dam taking care to align the clamp ends and dam edges Install the dummy gel screws facing out in the second cradle in the d2. A tampa de seguran a 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 agua ou 50 50 glicol de gua ethylene pelo exchanger de calor se for assim equiparam 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 instrumento Org nico solvente causar agress o irrepar vel unidade Nao opera com temperaturas de buffer acima do m ximo especificou especificac es t cnicas Superaquecer causar agress o irrepar vel a unidade pv Informaci n Importante Spanish Si este equipo es utilizado en una manera no especificado por Hoefer Inc la protecci n proporcionado por el eguipo 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 La tapa de la seguridad debe estar en el lugar antes de conectar la alimentaci n lleva a una alimentaci n Apaga
3. product qty code no Gel casters For 1 or 2 gels Dual Gel Caster 1 SE6015 basic 2 gels 18 cm wide Includes 2 blank gaskets for 1 or 2 gels One included with each SE600 Chroma unit For up to 4 gels Gel Caster Kit 1 SE675 4 gels 18 x 16 cm Includes 8 glass plates 3 space saver plates 5 filler sheets 100 sheets of wax paper Spacer Mate alignment template and filler plugs Order combs and spacers separately For up to 10 gels Multiple Gel Caster Kit 1 SE615 10 gels 18 x 16 cm Includes 20 glass plates space saver plate 5 filler sheets 100 sheets of wax paper Spacer Mate alignment template and filler plugs Order combs and spacers separately p35 product qty code no Clamps and cams Clamp and Cam Kit four 16 cm clamps and 8 black cams 1 SE6003UK Replacement thumbscrews for clamps 12 SE6003U 2 Cams black for clamps with cam holes 4 SE6005L Clamp assemblies 16 cm SE6003U Clamp assemblies 8 cm SE6403U Glass plates 18 x 8 cm Glass plates 2 SE6402 Glass plate club sandwich divider notched 1 SE6402D 18 x 16 cm Glass plates 2 SE6102 Glass plate club sandwich divider notched 1 SE6102D SER11 e p36 choose the appropriate spacer and plate length for your unit universal clamp SE6003U basic caster SE6015 cam Combs number thic
4. Casting stand The casting stand holds assembled gel sandwiches upright for casting gels Adjustable feet level the caster A laminated gasket in the bottom of each casting cradle seals the bottom of the sandwich when it is clamped into the stand Cams Cams are used twice first to secure the assembled sandwich in the casting stand and second to attach the sandwich to the upper buffer chamber Rubber gaskets There are two sets of two gaskets The solid laminated gaskets fit into the bottom of the casting stand and form the seal for casting the gel The slotted gaskets fit under the upper buffer chamber and form the seal between the upper and lower chambers The ridges on the upper gasket align the gasket slot to maintain an open channel between the top of the gel and the buffer in the upper chamber Spacers Spacers determine the thickness of the gel and are available in three thicknesses 0 75 1 0 and 1 5 mm and two widths 1 and 2 cm May be ordered separately Spacer Mate alignment template This template aligns spacers during sandwich assembly Combs Combs are available in sizes that form 10 12 15 20 or 28 wells Preparative combs include one or two reference wells in addition to a preparative well Most combs are available in all three thicknesses 0 75 1 0 and 1 5 mm May be ordered separately All preparative combs and the 10 12 15 and 20 well combs form wells that are 25 mm deep The 28
5. rana Hoefer SE600 Chroma Standard Dual Cooled Gel Electrophoresis Unit um SE600X IM Rev CO 04 12 Ad O e fe r e Contents Important Information Waste Electrical and Electronic Equipment WEEE Gel Electrophoresis Unit Function and Description wawa aa aaa AAA iere el Unpacking and Inventonm Operating Insiruchons eee Prepare the Gel Sandwich Acrylamide GelS Gradient GelS REESEN EEN a sunan Sample Preparation and Loading Final Assembly eee Separating the Sample Care and Maintenance 11111111 Troubleshooting iminente BiDWOSPAaPhY wez EA alk Ordering Information pi Important Information English If this eguipment is used in a manner not specified by Hoefer Inc the protection provided by the eguipment 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 maintaining 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 eguipped Do not connec
6. System Second Edition Large Scale Biology Press 1991 Bravo R Schafer R Willecke K MacDonald Bravo H Fey S J and Celis J E More than one third of the discernible mouse polypeptides are not expressed in a Chinese hamster mouse embryo fibroblast hybrid that retains all mouse chromosomes Proc Natl Acad Sci USA Apr 79 7 2281 2285 1982 Hurkman W J and Tanaka C K Solubilization of Plant Membrane Proteins for Analysis by Two Dimensional Gel Electrophoresis Plant Physiology 81 802 806 1986 Mets L J and Bogorad L Two dimensional polyacrylamide gel electrophoresis an improved method for ribosomal proteins Anal Biochem Jan 57 1 200 210 1974 O Farrell P H High resolution two dimensional electrophoresis of proteins J Biol Chem May 25 250 10 4007 4021 1975 Bjellgvist B et al Isoelectric focusing in immobilized pH gradients principle methodology and some applications J Biochem Biophys Methods 6 317 339 1982 Gorg A et al The current state of two dimensional electrophoresis with immobilized pH gradients Electrophoresis 9 531 546 1988 Gorg A Two dimensional electrophoresis with immobilized pH gradients current state Biochem Soc Trans 21 130 132 1993 Bjellqvist B et al Micropreparative two dimensional electrophoresis allowing the separation of samples containing milligram amounts of proteins Electrophoresis 14 1375 1378 1993 Blomb
7. ce oznakowanie 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 ch odziwo r d a je eli ci nienie wody jest nieuregulowanych Nigdy nie wprowadza rozpuszczalnika organicznego 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 Informac es Importantes Portuguese Se este equipamento usado numa maneira n o especificada por Hoefer Inc que a protecc o fornecida pelo equipamento pode ser comprometida Este instrumento projectado para uso de interior de laborat rio s S 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 seguranca registrada por um nacionalmente reconhecido testando laborat rio
8. concentrations high salt concentrations or inconsistent chemical quality Check band progress after 5 min and again after 1 h keeping an eye on the migration rate of the tracking dye The run is complete when the tracking dye reaches the bottom of the gel Watch the buffer level and if necessary replenish it as required to keep the top electrode submerged A small volume of buffer may leak past a nicked plate or gasket or buffer may pass through the gel After electrophoresis 0 Once the tracking dye reaches the bottom of the gel turn off the power supply disconnect the leads and remove the safety lid using finger leverage between the lid and the top of the heat exchanger Lift straight up to avoid bending the banana plugs a If coolant is circulating stop the flow and disconnect the fittings or tubing Pull out the upper buffer chamber assembly Pour the buffer into a sink Install the assembly in the dual gel caster and then release the sandwiches by turning and removing the cams o Unscrew the clamps from the sandwiches and remove Gently loosen and then slide away both spacers Use the Hoefer Wonder Wedge Gel Plate Separation tool to separate the plates Carefully lift the glass plate with the gel attached Handle the gel with care to avoid damaging it Invert the plate and position the gel low over the staining tray Pry one corner of the gel away from the glass and al
9. pointing up Seal the gel sandwich against the casting gasket by turning both cams as far as needed usually 90 150 up to 180 The cam action presses the plates down into the gasket to seal the bottom of the sandwich The seal is complete once the glass edge appears darker and nearly transparent against the gasket Do not turn past this point glass plate spacer cam hole gasket foam side down SG 2 le cam hole p casting cradles 2 cam hole leveling feet h spirit lev cam install ridge cam hole end up e pll e pl2 Acrylamide Gels Prepare the monomer solution and pour the gel Prepare the reguired amount of monomer solution Deaerate and add the initiator and catalyst just prior to pouring the gel Pipette the solution into one corner of the sandwich taking care not to introduce any air bubbles See below for the appropriate solution level according to the application No stacking gel Continuous system Fill solution to just below the top of the upper plate edge If bubbles are trapped remove with a pipette or syringe Introduce a comb at a slight angle into each sandwich taking care not to trap air bubbles under the teeth Club sandwich Pipette the solution into both sandwiches filling each to the same level below the notched edge Stacking gel Fill solution to 3 4 cm below the top of the glass plate This height allows 1 cm of stacking gel below the wells Pou
10. R and Smith J A Electrophoretic separation of proteins In Current Protocols in Molecular Biology Ausubel F A eds OSC 10 2 1 10 2 21 1991 Hames B D and Rickwood D Gel Electrophoresis of Proteins A Practical Approach Second edition City IRL Press 1990 Sambrook J and Russell D W Molecular Cloning A Laboratory Manual Cold Spring Harbor Laboratory Cold Spring Harbor NY 2001 Sasse J and Gallagher S R Staining proteins in gels Current Protocols in Molecular Biology Ausubel F A et al eds OSC 10 6 1 10 6 8 1991 SDS Polyacrylamide Gel Electrophoresis and Isoelectric Focusing Handbook 80 6013 88 Hoefer Inc 2001 Non denaturing gel systems Reisfeld R A et al Acidic buffer system for resolution of cationic proteins Nature 195 281 1962 McLellan T Electrophoresis buffers for polyacrylamide gels at various pH values Anal Biochem 126 94 1982 Hedrick J L and Smith A J Size and charge isomer separation and estimation of molecular weights of proteins by discontinuous gel electrophoresis Arch Biochem Biophys 126 155 1968 Denaturing gel systems Laemmli U K Cleavage of structural proteins during the assembly of the head of bacteriophage T Nature 227 680 685 1970 Matsudaira P T and Burgess D R SDS microslab linear gradient polyacrylamide gel electrophoresis Anal Biochem 87 386 396 1978 Schreier M H Erni B a
11. exchanger Replace worn grommets Fill the lower buffer chamber to the level appropriate for at edges the run See Fig 8 page 21 Use magnetic stirrer and stir bar to keep buffer well mixed Excessive heat Particulates in sample Circulate ext coolant Decrease the current or voltage setting Prechill the buffer Run the gel in the cold room Centrifuge or filter sample before loading to remove particulates Overloading Load less sample Degradation Current leakage around gel Add protease inhibitor such as PMSF Check for leaks all plates and spacers must be aligned and free of grease and cracks If used the buffer dam must be secure Sample or reagent preparation If the required pH of a solution is overshot do not back titrate Discard and prepare fresh buffer Check recipes gel concentrations and buffer dilution For instance do not use Tris HCI instead of Tris for Laemmli tank buffer Decrease the salt concentration of samples Reagent quality Dispose of older acrylamide solutions and use only stock of the highest quality Use only freshly deionized urea Voltage or current settings To increase or decrease the migration rate adjust the voltage or current by 25 50 problem Bands are skewed or distorted possible cause Incomplete gel preparation and polymerization remedy Degas the stacking gel solution and avoid trapping air bubbles under the com
12. prevent gel exposure to oxygen Slowly deliver the overlay solution from a glass syringe fitted with a 22 gauge needle Apply the solution near the spacer at one side of the sandwich and allow it to flow across the surface unaided o Allow the gels to polymerize for a minimum of 1 h After polymerization pour off the overlay and rinse the gel surface several times with distilled water e p15 _ __ __Z A Note With Coomassie Blue it is possible to detect 1 ug of protein in a single band With the more sensitive silver stains it is possible to detect as little as 10 ng of protein e p16 Prepare the stacking gel monomer solution pour the stacking gel and introduce a comb at a slight angle into the sandwich taking care not to trap air under the teeth Allow a minimum of 1 h for the gel to polymerize Sample Preparation and Loading The sample can be loaded either while the sandwich is in the caster or after the upper buffer chamber is attached When loading samples while using divider plates the samples must be loaded without the upper buffer chamber in place The amount of sample loaded depends on the thickness of the gel the sensitivity of the detection method used and the amount of sample expected in each band In a continuous buffer system the protein sample should be relatively concentrated because no stacking gel is used In a discontinuous buffer system the zone into which e
13. to subtract from the desired run temperature 10 W intersects the graph at about 1 C If the desired temperature is 23 C set the bath to 23 12 22 C If the desired temperature is 4 C set the bath to 4 1 3 C Fig 8 Upper and lower buffer chamber fill levels Use the chart Fig 7 on page 20 to estimate a starting point for the circulator bath temperature setting Adjust as necessary for variables such as ambient temperature changes in power output and circulator bath efficiency If accurate temperature control is critical measure the temperature and adjust as necessary Optional Prechill the buffer Fit the upper buffer chamber assembly into the lower buffer chamber Use a steady hand to avoid disturbing the samples Grasp the assembly in the casting stand by the upper buffer chamber and carefully lower it into the lower chamber o Inspect the installation and check the buffer levels Upper buffer chamber UBC The electrode along the upper chamber ridge must be submerged about 1 cm This level reguires 450 600 ml of buffer just enough to cover the upper chamber ribs but not high enough to contact the banana plug Do not fill above UBC MAX fill line Lower buffer chamber LBC Fill to LBC MAX fill line Upper chamber bt buffer max fill line MAX
14. todos controles de alimentaci n y desconecta 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 Recalentar 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 tillhandah ll vid utrustningen kan skadas Detta instrument formges f r inomhuslaboratorium anv ndning bara Bara medhj lpare och delar godkande eller levererade 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 erkand testande laboratorium Sakerheten locket maste vara pa platsen fore koppla kraften tillgangen blyen till en kraft tillgang Vander sig alla kraft tillgang kontroller av och kopplar bort kraften blyen fore flytta s kerheten locket Cirkulerar bara vatten eller 50 50 vatten ethyl
15. 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 laboratoriota Turvallisuuskansi t ytyy olla paikallaan k yttoj nnitteeseen Kiert kaikki kaytt jannitevalvonnat ja irrottaa valtalyijyt ennen poistaminen turvallisuuskantta Kiert vain vesi tai 50 50 vesi ethylene glycol siin tapauksessa varustetun limm nvaihtimen l pi l yhdist l mm nvaihdinta vesinapautukseen eik j hdytysnestel hteeseen miss vesipaine 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 Ylikuumeneminen 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 tre diminu e Cet instrument est con u pour l usage de laborat
16. 999 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 2012 Hoefer Inc All rights reserved Printed in the USA Hoefer
17. Lower chamber buffer max fill line Buffer level i label MAX e p21 p22 Place the safety lid on the unit by engaging the safety interlock pins before lowering the electrode connections on to the banana plugs Plug the color coded leads into the jacks of an approved power supply Plug the red lead into the red output jack and the black lead into the black output jack In most systems the red lead which is connected to the bottom electrode is the anode and the black lead connected to the top electrode is the cathode Important assembly notes e IEF Runs The buffer level in the lower buffer chamber must never reach the upper buffer chamber maintain at least 2 cm of clearance e Do not fill the upper or lower chamber above the recommended levels illustrated in Fig 8 Remove buffer in contact with the electrode posts e Pour buffer slowly and away from the slots in the upper buffer chamber to avoid disturbing the samples e Use only water or 50 50 water ethylene glycol as coolant Never use a commercial antifreeze or any alcohol based mixture or irreparable damage to the heat exchanger will result e Do not connect the heat exchanger to a water tap or any other source where the water pressure is unregulated Note SE600 Chroma unit uses 18 cm wide plates The gel thickness determines the cross section and current reguirement for constant current runs The length of the plate deter
18. ach molecular species migrates is sharpened by the stacking gel so the sample need not be as concentrated Prepare the wells Remove the comb by gently rocking it side to side and then lifting it straight up to avoid damaging the well walls Carefully rinse each well with distilled water to remove unpolymerized acrylamide and then drain by inverting the gel sandwich or caster Fill each well with electrophoresis buffer O Note Once the samples are in the wells take care to not jar the sandwiches so that the samples are not spilled or mixed Prepare the sample Increase liquid sample density with 10 glycerol or sucrose Add a tracking dye such as phenol red bromophenol blue or pyronin Y For SDS protein gels use 2X treatment buffer to denature both liquid and dry samples in a test tube To liquid protein solutions add an equal volume of 2X buffer To dry protein samples add equal volumes of 2X sample buffer and high purity water to achieve the desired concentration Heat the tube in boiling water for 90 seconds then allow to cool to room temperature Treated samples can be stored at 40 to 80 C for future runs Heat membrane proteins to 60 C for 20 minutes Store unused sample at 4 C o Underlay the sample into the wells using a fine tipped microsyringe or gel loading pipette tip Table 1 Sample volume for standard comb sizes volume of sample pl per 1 mm depth
19. alt concentrations e p29 problem possible cause Poor band Running resolution conditions remedy Begin electrophoresis as soon as the sample is loaded to prevent low molecular weight species from diffusing Conduct the separation at a lower current or voltage setting to reduce Joule heating Reagent guality Use only the highest guality reagents Poor stacking Use only gels that were recently prepared Add a stacking gel or increase height of the stacking gel Prepare the resolving gel surface by first rinsing it with stacking gel monomer before pouring the stacking gel to ensure continuity between the gels Check pH values of the resolving and stacking gel solutions Do not back titrate buffers Incomplete gel polymerization Allow gel to polymerize fully Sample preparation e p30 Store sample on ice before it is denatured Dialyze or desalt the sample Heat samples in SDS sample buffer for no more than 1 2 min at 100 C to improve dissociation of subunits Store on ice after heating Adjust the sample volume or concentration Add more mercaptoethanol or dithiothreitol check sample treatment Add protease inhibitors such as PMSF if necessary to prevent proteolytic degradation of sample Increase glycerol or sucrose to increase sample density Store samples to be frozen in aliquots to avoid repeated freeze thawing Store at 40 to 80 C Bibliography General Gallagher S
20. b teeth Irregular interface between stacking and running gels Overlay the running gel with water saturated butanol before polymerization begins to avoid forming an uneven gel surface Sample preparation Stained sample collects Near the buffer front Gel concentration Degradation Dialyze or desalt the sample Molecules are not sufficiently restricted by the resolving gel pore size increase the T Proteins may be degraded by endogenous proteases use protease inhibitors during the isolation step Near the top of the gel when the buffer front has reached the bottom Gel concentration Precipitation The gel pore size is too small decrease the T of the resolving or stacking gel The protein has precipitated Heat the sample at a lower temperature 70 C or less for 1 2 min At both top and bottom of the gel Tracking dye doesn t sharpen into a concentrated Gel concentration Poor stacking The molecular weight range of the sample requires an acrylamide concentration gradient to resolve the full range of protein sizes 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 zone in the stacking gel Reagent quality Sample preparation gt gt gt When preparing samples avoid using solutions with high s
21. cations Overheding vil for rsage uboelig skade til enheden O 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 ethyleenglycol 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 oplosmiddelen 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 Jos tata
22. e and terminates at the banana plug The upper chamber reguires 0 5 0 8 Liters of buffer fill no higher than the top of the plastic ribs Heat exchanger The heat exchanger must be installed for every use because it houses the bottom electrode anode which runs along the bottom of the frame When connected to a circulator bath the heat exchanger regulates the buffer temperature in the lower chamber Coolant passes through the glass tubes which are secured with silicone rubber grommets The heat exchanger connector ports are 13 mm o d The heat exchanger is rated to a maximum of 0 8 atmospheres above ambient 12 psig Connect only to coolant sources with regulated pressure Do not connect to the water tap Safety lid The banana plug on the heat exchanger connects to the red lead and the plug on the upper buffer chamber connects into the black lead The 4 mm shrouded color coded leads plug into color coded jacks in the power supply Engage interlock pins before lowering electrode connections on to banana plugs Always install the safety lid before use Glass plates The SE600 Chroma accommodates 18 cm wide plates 16 or 8 cm long Notched divider plates ordered separately divide gel sandwiches to form club sandwiches of two gels each so up to four gels can be run at one time Clamps Two 16 cm clamps are used to secure the gel sandwich The clamp pressure bar adjusted with screws distributes pressure evenly
23. e sandwich upright on a flat surface set it into the casting cradle Fig 3 Club sandwich assembly Side clamps will accommodate two spacers up to 1 5 mm thick glass plates at the outer sides of the sandwich spacers notched center plate e plo o Club sandwich A 16 cm long notched center divider plate ordered separately pairs two sandwiches to double the number of gels that can be cast and run Assemble a club sandwich in the same manner as a regular sandwich except before placing the top glass plate lay the divider plate and a second set of spacers on the stack Place the notch so that it will be at the top of the gels It is essential that the spacers and plates align perfectly in order to seal Remove the sandwich and inspect the bottom to make sure that edges are aligned flush to ensure a complete seal Adjust if necessary Optional Apply a light film of Gel Seal compound only on the bottom corner surfaces created by the spacers and plates if the sandwiches tend to leak Place the laminated gasket into the casting cradle See Fig 4 with the foam side down Place the clamp assembly in the casting cradle screw side facing out Note When turning the cams it is easier to keep the caster balanced if you turn both toward the center of the caster Fig 4 Caster components and setup o Insert a cam into the hole on each side of the casting tray with the ridge short end
24. e 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 piv nasjonalt ha blitt anerkjent prover laboratorium Sikkerheten lokket m v re p plass far forbinding kraftforsyningene blyene til en kraftforsyning Vender all kraftforsyningsstyring av og frakopler kreftene blyene for 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 kjalemiddelkilde hvor vannet trykket er unregulated Introduserer Aldri antifreeze eller noe organisk losemiddel inn i noe del av instrumentet Organiske losemiddler vil for rsake irreparabel skade p enheten Driver med buffertemperaturer over maksimum ikke spesifiserte teknisk spesifikasjoner A overoppheting vil for rsake irreparabel skade p enheten Wazne Informacje Polish Jezeli ten sprzet jest wykorzystywany w spos b nie okreslone przez Hoefer Inc do ochrony przewidzianej przez urz dzenie moze zosta obni ony Instrument ten jest przeznaczony do u ytku w laboratoriach kryty tylko Tylko akcesori w i cz ci zatwierdzone lub dostarczone przez Hoefer Inc mog by wykorzystane do eksploatacji utrzymania i obs ugi tego produktu korzysta jedynie zasilacza e jest nosz
25. embly Inspect glass plates for nicks Use only unchipped plates to prevent leaking clamp spacer glass plates ridges pressure plate Note The glass plates and spacers must be flush with the clamp ridges at both top and bottom for a good seal Note Do not use silicone grease or petroleum jelly to seal the sandwich These substances are difficult to remove and ultimately cause artifacts Construct the gel sandwich and insert into caster Prepare the caster and clamps Place the spirit level into the caster center and adjust the leveling feet Loosen all clamp screws and make space for the sandwich by sliding the pressure plates toward the screws a Construct gel sandwiches For each sandwich choose two perfectly clean unchipped glass plates and two spacers Lay one plate on a flat surface lay the Spacer Mate alignment template onto the plate wide side at the top of the plate place a spacer along each edge and lay the second glass plate on top Secure the sandwich with clamps Slide one clamp at a time along the sandwich sides Finger tighten one screw on each clamp set the sandwich upright on a flat surface and loosen the screw to align the stack Taking great care in alignment will ensure a good seal Finger tighten all screws Remove the Spacer Mate Tip Use the casting cradle to hold the sandwich during alignment Remove the laminated gasket from the cradle and instead of setting th
26. ene glycol genom varmen exchanger i sa utrustad fall Inte kopplar varmen exchanger till en vatten kran eller nagot kylmedel k lla dar vattnet trycket ar 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 ver det h gsta angivna tekniska specifikationerna verhettning skulle orsaka irreparabla skador p enheten O pvi English Ea French R German R Ea Italian R lt Spanish 14 Swedish R Kee 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 a 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 unsortierten Hausm ll entsorgt werden d rfen sondern separat behandelt werden m ssen Bitte nehme
27. erg A et al Interlaboratory reproducibility of yeast protein patterns analyzed by immobilized pH gradient two dimensional gel electrophoresis Electrophoresis 16 1935 1945 1995 e p33 safety lid with cables SE6056X upper buffer chamber SE6054 heat exchanger SE6160 lower buffer chamber SE6150X e p34 Ordering Information product qty code no SE600 Chroma complete unit 1 SE600X 15 1 Includes 3 sets of glass plates two 15 well combs 2 sets of spacers 1 5 mm thick 6 cams dual gel casting stand with leveling base and level buffer dam Spacer Mate alignment template and Wonder Wedge Gel Plate Separation tool Replacement parts Wonder Wedge Gel Plate Separation tool 1 SE1514 Slotted silicone rubber gaskets 2 SE6008B for upper buffer chamber Laminated silicone rubber gaskets 2 SE6009 for casting stand Buffer dam SE6032 Upper buffer chamber SE6054 for SE600 Chroma Lower buffer chamber for SE6150X SE600 Chroma Lid with high voltage leads SE6056X for SE600 Chroma High voltage safety lead set SE6056 HV Banana plug gold with 2 washers SE6067 SE600 Chroma Heat exchanger SE6160 lower electrode assembly Glass tube with 2 grommets SE6160 5 for heat exchanger lower electrode assembly Grommets for heat exchanger 4 SE6060 6 lower electrode assembly Spirit level SER11 Gel Seal compound 1 4 oz tube SE6070 Spacer Mate 3 SE6119SM
28. kness width of wells mm mm qty code no 10 0 75 8 3 SE511 10 75 10 1 00 8 3 SE511 10 1 0 10 1 50 8 3 SE511 10 1 5 12 0 75 7 6 SE511 12 75 12 1 00 7 6 SE511 12 1 0 12 1 50 7 6 SE511 12 1 5 15 0 75 5 7 SE511 15 75 15 1 00 5 7 SE511 15 1 0 15 1 50 5 7 SE511 15 1 5 20 0 75 4 1 SE511 20 75 20 1 00 4 1 SE511 20 1 0 20 1 50 4 1 SE511 20 1 5 28 0 75 2 7 SE511 28 75 28 1 00 2 7 SE511 28 1 0 28 1 50 2 7 SE511 28 1 5 Comb depth 15 mm all others 25 mm Preparative combs These combs are 25 mm deep adjustable to 10 or 15 mm no of wells thickness width mm prep ref mm prep ref qty code no 1 1 0 75 121 6 1 SE511 R 75 1 1 1 121 6 1 SE511 R 1 0 1 1 1 50 121 6 1 SE511 R 1 5 1 2 0 75 113 6 1 SE511 DR 75 1 2 1 113 6 1 SE511 DR 1 0 1 2 1 50 113 6 1 SE511 DR 1 5 Adjustable comb back 1 SE511 BKA Required to convert any 25 mm deep comb to 10 or 15 mm depth e p37 Spacers thickness mm length cm width cm qty code no 0 75 8 2 2 SE6419 2 75 1 0 8 2 2 SE6419 2 1 0 1 5 8 2 2 SE6419 2 1 5 0 75 16 2 2 SE6119 2 75 1 0 16 2 2 SE6119 2 1 0 5 16 2 2 SE6119 2 1 5 1 0 16 1 2 SE6118 2 1 0 5 16 1 2 SE6118 2 1 5 Companion products Hoefer SE100 Plate Mate washing and storage unit 1 SE100 QuickFit connectors female 3 8 2 QF3 8 QuickFit connectors male 3 8 2 QFX3 8 e p38 Hoefer Inc 84 October Hill Road Holliston MA 01746 Toll Free 1 800 227 4750 Phone 1 508 893 8
29. ks around the gasket Lower buffer chamber Place a magnetic spin bar into the lower buffer chamber LBC and place the unit on a magnetic stirrer Fill the lower chamber with up to 4 liters of buffer a Lower the heat exchanger into the lower chamber fitting the ports into the notches in the rim The heat exchanger must be in place for all runs because the lower electrode is integrated into the heat exchanger If no cooling is reguired skip to step 3 Optional Connect the heat exchanger to a thermostatic circulator Slide hose clamps four total onto each end of two lengths of 10 12 mm i d 3 8 1 2 vinyl or silicone tubing Attach one end of each length of tubing to a heat exchanger port Attach the free ends of each length of tubing to the circulator bath ports one to the inlet and the other to the outlet Secure the connections with the hose clamps e pl9 Fig 7 Approximate circulator bath temperature setting Set the circulator bath temperature setting lower than the desired run temperature by the amount indicated on the graph This should be checked at three points e p20 0 1 v 2 s 3 o 8 a 5 3 4 lt 5 5 o 6 7 0 10 20 30 40 50 60 power supply setting W Example Run parameters 200 V 0 05 A 50 mA 1 Calculate W if your power supply does not display power directly W VxA 10W 200Vx0 05A 2 Interpolate the number of degrees
30. la do jak koli sti z tohoto n stroje Rozpustidl m zp sob nenapraviteln po kozen jednotka Nejsou provozov na s pufru teplot ch nad maxim ln stanovenou technick mi specifikacemi P eh t zp sob nenapraviteln po kozen jednotka Z igtig Information Danish Hvis dette udstyr bruges i en made ikke specificeret ved Hoefer Inc den beskyttelse som er blevet forsynet af udstyret kan maske svaekkes Dette instrument er designet for indend rs laboratoriumbrug bare Bare tilbeh r og del godkendede eller forsynede ved Hoefer Inc kan m ske bruges for drive funktionsfejl og betjening dette produkt bruger Bare en stromforsyning der er CE markerede eller sikkerhed som er blevet attesteret af en som nationalt er blevet anerkendt prove laboratorium Sikkerhedlaget ma veere pa plads for forbinding stramforsyningsblyet til en str mforsyning Drejer alle stramforsyningskontroller 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 kolemiddelkilde hvor vandtrykket er unregulated Introducerer Aldrig antifreeze eller noget organisk oplosningsmiddel ind i nogen del af instrumentet Organiske opl sningsmidler vil for rsage uboelig skade til enheden Driver ikke med st dpudetemperaturer over maksimummet specificerede tekniske specifi
31. low it to drop into the tray or if the gel is thick enough to handle lift it and place it into the tray To avoid splashing add staining or fixative solution to the tray after the gel is transferred Clean the unit as described in the next section e p25 Cleaning e Do not autoclave or heat any part above 45 C e Do not use organic solvents abrasives strong cleaning solutions or strong acids or bases to clean the chambers e Do not soak the laminated gasket Note If the old tube is cracked or broken protect your hand with thick gloves a piece of cloth or paper towels before removing the tube p26 Care and Maintenance Immediately after each use rinse the upper and lower buffer chambers with water and then rinse thoroughly with distilled water Handle the upper buffer chamber with care to prevent damaging the banana plug Clean gaskets with mild detergent and rinse with distilled water Allow to air dry Clean glass plates and spacers with a dilute solution of a laboratory cleanser such as RBS 35 then rinse thoroughly with tap and distilled water Glass plates can also be treated with but not stored in acid cleaning solutions Replacing a heat exchanger glass tube Remove the tube by simultaneously twisting and sliding it down as far as possible until the top end is free of the upper grommet Carefully guide the tube so that it will clear the assembly then lift the tube out of the
32. lower grommet a Lightly grease the outside of both ends of the new tube with silicone grease Twist and slide one end of the tube into the lower grommet Then slip the other end into the top grommet gently pushing it with a slight twist until it stops Check that the grommet is not pinched Troubleshooting problem Gel sandwich leaks while casting Sample wells damaged or irregular possible cause Dirty or damaged components remedy Plates spacers and the gasket must be completely clean Wash if necessary Replace chipped plates especially if chipped near the spacers Check the caster gasket for cuts or cracks and replace if necessary Mis aligned parts Check plate and spacer alignment realign if necessary Over clamping Air bubbles Turn cam only as far as necessary to create a seal usually 90 1507 but up to 180 On each spacer apply a light film of Gel Seal compound to the bottom outside corner only Do not use silicone grease Remove air bubbles before inserting combs Slide comb into solution at an angle If comb must be removed add more monomer solution before reinserting the comb Incomplete or delayed polymerization Allow acrylamide gels to set for a minimum of 1 h Debris in wells Rinse out unpolymerized gel with sample buffer Comb removal Remove the comb at a slight angle and very slowly to prevent damaging the gel Agarose gels Lowe
33. mines the running time Table 2 Laemmli buffer system starting point guidelines Gel thickness 1 5 mm Current per gel 25 mA constant current Starting voltage 80 90 V 220 250 V Final voltage Thicker or thinner gels require proportionally more or less current For example a 0 75 mm gel which is half as thick as a 1 5 mm gel requires half as much current or 12 5 mA The current must be multiplied by the number of gels For instance if two club sandwiches are installed the four gels require four times as much current The current can be increased for faster runs if active cooling is used and it can be decreased for slower overnight runs At 25 mA per gel Separating the Sample Electrophoresis parameters for discontinuous polyacrylamide gels Gels may be run at either constant current or constant voltage settings A constant current mode is traditionally used with a discontinuous buffer system so that the rate of electrophoretic migration remains unchanged throughout the run Under these conditions voltage increases as the run proceeds A lower current setting is recommended for higher resolution The optimal current level must be determined empirically the main factors that must be balanced include the gel concentration and migration speed and the resulting Joule heating and band distortion Table 2 lists starting point guidelines and adjustments for gel thickness number of gel
34. n 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 Gel Electrophoresis Unit Function and Description The Hoefer SE600 Chroma vertical slab gel electrophoresis unit is intended for protein and nucleic acid electrophoresis under commonly used denaturing and non denaturing conditions Up to 28 samples can be compared on a single slab gel Applications include protein separations nucleic acid fractionation and the second dimension separation of 2 D electrophoresis First dimensi
35. nd Staehelin T Initiation of mammalian protein synthesis I Purification and characterization of seven initiation factors J Mol Biol Nov 116 4 727 753 1977 e p31 e p32 Shapiro A L and Maizel J V Jr Molecular weight estimation of polypeptides by SDS polyacrylamide gel electrophoresis further data concerning resolving power and general considerations Anal Biochem Jun 29 3 505 514 1969 Schaegger H and Von Jagow G Tricine sodium dodecyl sulfate polyacrylamide gel electrophoresis for the separation of proteins in the range from 1 to 100 kDa Anal Biochem 166 368 379 1987 Weber K and Osborn M The reliability of molecular weight determinators by dodecyl sulfate polyacrylamide gel electrophoresis J Biol Chem 224 4406 4412 1969 Two dimensional electrophoresis Adams L D and Gallagher S R Two Dimensional Gel Electrophoresis Using the O Farrell System Current Protocols in Molecular Biology Ausubel F A et al eds OSC pp 10 4 1 10 4 13 1992 Anderson N G Anderson N L and Tollaksen S L Proteins of human urine I Concentration and analysis by two dimensional electrophoresis Clin Chem Jul 25 7 1199 2210 1979 Anderson Leigh and Anderson Norman G High resolution two dimensional electrophoresis of human plasma proteins Proc Natl Acad Sci USA 74 5421 5425 1977 Anderson L Two Dimensional Electrophoresis Operation of the ISO DALT
36. ng through a peristaltic pump Attach one end of the tubing to the gradient maker outlet port and the other end to a 20 cm cannula The o d of the cannula must be less than the spacer thickness Place the cannula so that it rests at the bottom of the sandwich midway between the spacers Optional Adjust the higher percentage acrylamide solution to 15 w v sucrose or 25 v v glycerol to improve layering Prepare the monomer solution Calculate the volume of monomer solution needed Divide the total volume in half and prepare this volume of both the higher and lower percentage acrylamide solutions o Pour the light solution into the reservoir chamber the chamber farthest from the outlet Open the stopcock between the chambers long enough to displace the air and then close Pour the heavy solution into the mixing chamber and place a stirring bar into this chamber Place the gradient maker onto a magnetic stirrer and begin stirring at a rate that mixes well but does not introduce bubbles into the solution Mix the gradient and pump the solution into the sandwich While the solution is stirring begin pumping from the mixing chamber and open the stopcock to the reservoir chamber Raise the cannula as liquid enters the sandwich keeping the tip at the gel surface Prepare more gels as required Overlay each gel with a thin layer of water saturated butanol water or diluted gel buffer to
37. oire int rieur seulement Seulement les accessoires et les parties ont approuv ou ont fourni par Hoefer Inc pourrait tre utilis pour fonctionner maintenir et entretenir ce produit utilise Seulement une alimentation qui est CET a marqu ou la s curit certifi par un nationalement 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 lexchanger de chaleur a un robinet d eau ou a 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 piii instrument Les dissolvants organigues causeront des dommages irr parables a l unit Ne pas fonctionner avec les temp ratures de tampon au dessus du maximum a sp cifi des sp cifications technigues La surchauffe causera des dommages irr parables a 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 Schutz verschlechtert werden Dieses Instrument wird f r den Innenlabo
38. on separation of 2 D protein electrophoresis should be performed on Immobilized pH Gradient Gels The focused strips are easily transferred to the second dimension slab gel for size separation The gel plates are 18 cm wide by 16 cm long Up to four gels can be run at one time if sandwiches are paired into club sandwiches The heat exchanger allows buffer temperature control in the lower chamber Specifications Gel plate size 18 x 16 cm w x h Gel size 14 or 16 cm x 16 cm w x h Maximum watt 50 W Maximum volt 1000 V Maximum amperage 500 mA Maximum temperature 45 C Environmental operating conditions Indoor use 4 40 C Humidity up to 80 Altitude up to 2000 m Installation category II Pollution degree 2 Dimensions 32 x 29x 14 cm w x h x d 12 5 x 11 5 x 5 5 in Product certifications EN 61010 1 UL 61010A 1 CSA C22 2 1010 1 CE Certified This declaration of conformity is valid only when the instrument is e used in laboratory locations e used as delivered from Hoefer Inc except for alterations described in the user manual and e connected to other CE labeled instruments or products recommended or approved by Hoefer Inc Fig 1 Main components of the Hoefer SE600 Chroma see Fig 4 for caster components Included but not shown e Gel Seal compound 1 4 oz e Spacer Mate alignment template e Glass plates 6 e Wonder Wedge plate separation tool e B
39. per l uso di laboratorio 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 riconosciuto testando il laboratorio ll coperchio di sicurezza deve essere nel luogo prima di collegare i piombi di alimentatore a un alimentatore Spegne tutto i controlli di alimentatore e disinserisce i piombi di potere prima di togliere il coperchio di sicurezza Circola solo acgua o 50 50 glicole di acqua etilene attraverso lo scambiatore di calore se cosi equipaggiato Non collegare lo scambiatore di calore a un rubinetto di acqua o qualunque fonte di refrigerante 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 unita Non opera con le temperature di tampone al di sopra del massimo ha specificato le descrizioni tecniche Il surriscaldamento causera il danno irreparabile all unita Viktig Informasjon Norwegian Hvis dette utstyret blir brukt i en m te ikke spesifisert ved Hoefer Inc beskyttelsen som ha blitt git av utstyret kan bli svekket Dette instrumentet er utformet for innend rs laboratoriumbruk bare Bare tilbehar og deler godkjente eller forsynt
40. r solution deaerate it and add catalyst and initiator Pour the stacking gel onto the resolving gel with a disposable or Pasteur pipette to a level about 2 mm from the top of the plate o Introduce a comb at a slight angle into the sandwich taking care not to trap air under the teeth Allow a minimum of 1 h for the gel to polymerize e pl3 Fig 5 Pouring a gradient gel A pipette tip may be used instead of a cannula if the gel solution is delivered at a rate that maintains a continuous stream on the glass surface Note Gradient gels poured in the SE615 or SE675 Multiple Gel Caster are introduced through the bottom Note When pouring an exponential gradient gel position a plunger or sealing plug above the liguid in the mixing chamber to hold the volume constant e pl4 Gradient Gels Both linear and exponential gradient gels can be poured in the dual gel caster We recommend using a Hoefer SG Series Gradient Maker Gradient gels are poured from the top of the caster with a cannula if using the provided dual gel caster or from the bottom if using a Hoefer Multiple Gel Caster see instructions accompanying the caster A stacking gel is then poured over the gradient gel o OO o o oO Pouring a linear gradient gel Assemble sandwich es into the dual gel casters as described on page 9 a Set up the monomer solution flow path Run a length of clear vinyl tubi
41. r the comb no more than 1 cm into the gel Use only recent stocks of the highest quality reagents If the dry ammonium persulfate does not crackle when added to water replace with fresh stock Increase TEMED or APS concentration or both Solutions with extreme pH values especially acidic may not polymerize Remove oxygen from the gel environment Degas the monomer solution 5 10 min before pouring and then overlay the gel surface with water saturated n butanol Incomplete gel Chemicals polymerization pH Oxygen Temperature Adjust the gel solution temperature to a minimum of 20 C especially for low T gels e p27 problem Upper buffer chamber leaks Power supply detects current leak Dye front curves up smiles at edges Protein streaks vertically Unusually slow or fast run e p28 possible cause Mis aligned parts remedy Check that the glass plates spacers and clamps are aligned and fit snugly into the upper chamber gasket Check that both gaskets are centered and that the positioning ridges fit inside the grooves Dirty or damaged components Electrical path to outside ground earth Uneven heat distribution Check that the gasket is not damaged or pinched Replace if necessary Check that the upper buffer chamber is not warped from prior exposure to excessive heat Add more silicone grease to seal heat exchanger grommets Check for leaks or cracks in the heat
42. r the gel and apply an overlay see step 2 After the gel is set prepare the stacking gel as described below 2 D electrophoresis Discontinuous protein system Fill monomer solution to about 1 cm below the top of the glass plate to allow 4 5 mm for the IPG strip or tube gel and an agarose seal A stacking gel will reguire extra space Seal the IPG strip or tube gel in place with agarose dissolved in running buffer Take care to avoid trapping any air bubbles between the first and second dimension gels _ __ __Z A O Overlay each gel with a thin layer of water saturated butanol water or diluted gel buffer to prevent gel exposure to oxygen Slowly deliver the overlay solution from a glass syringe fitted with a 22 gauge needle Apply the solution near the spacer at one side of the sandwich and allow it to flow across the surface unaided Allow the gel to polymerize for a minimum of 1 h Stacking gel preparation Pour the stacking gel while the sandwich is still in the gel caster Stacking gel resolution is optimal when poured just before electrophoresis o Remove the overlay by rinsing the top of the gel several times with distilled water Invert the caster to drain To ensure a seamless contact between the resolving and stacking gels remove residual liguid by blotting one corner with a lab wipe Calculate the stacking gel monomer solution volume Prepare the stacking gel monome
43. rgebrauch nur daftir entworfen Nur Zus tze und Teile genehmigten oder lieferten durch Hoefer Inc kann f r das Funktionieren das Aufrechterhalten und die Wartung dieses Produktes verwendet werden Verwenden Sie nur eine Energieversorgung die CE gekennzeichnet oder durch ein national anerkanntes Probelaboratorium bescheinigte Sicherheit ist Der Sicherheitsdeckel 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 Warmeaustauscher wenn so ausgestattet in Umlauf setzen Verbinden Sie den W rmeaustauscher mit einem Wasserklaps oder jeder K hImittel Quelle nicht wo der Wasserdruck ungeregelt wird F hren Sie nie Frostschutzmittel oder jedes organische L sungsmittel in jeden Teil des Instrumentes ein Organische L sungsmittel werden nicht wiedergutzumachenden Schaden der Einheit verursachen Mit Puffertemperaturen Uber angegebenen technischen Spezifizierungen des Maximums nicht funktionieren Die Uberhitzung wird nicht wiedergutzumachenden Schaden der Einheit verursachen Informazioni Importanti Italian Se quest apparecchiatura usata in un modo specificato da Hoefer Inc la protezione fornito dall apparecchiatura potrebbe essere indebolita Questo strumento disegnato
44. s and migration rate Current Current acts on the total cross section area of all the gels because the gels are connected in parallel in the electrical circuit Thus the current setting for one gel must be multiplied by the number of gels of the same cross section run simultaneously For a gel 1 5 mm thick we suggest a starting current setting of 25 mA Two 1 5 mm gels 50 mA Note Cooling may be required to control Joule heating e p23 Caution After initial monitoring do not leave the unit unattended for more than 1 h before checking the progress of the bands and the buffer level e p24 Voltage The starting voltage for a 1 5 mm slab gel connected to a power supply set to 25 mA is usually 80 90 V using the SE600 Chroma unit with a Laemmli discontinuous buffer system for SDS gels The final voltage is typically 250 400 V depending on the length of the gel See Table 2 Time A run is usually complete when the tracking dye reaches the bottom of the gel In a 16 cm gel SE600 Chroma a 1 5 mm thick Laemmli SDS gel run at 25 mA gel without cooling usually requires 5 hours Record each run Keep a record of the current or voltage setting number and thickness of gels buffer system and the starting and final current or voltage readings for each run so that results can be compared Inconsistent results for the same system and settings indicate potential problems such as leaking current incorrect buffer
45. t the heat exchanger to 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 poskytnut 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 ethylenglykolu prost ednictv m v m n k tepla je li to vybavena 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 d
46. ual gel caster Attach the gel sandwich to the upper buffer chamber Turn the upper buffer chamber upside down and place a slotted gasket into both sandwich holder recesses Both the slot in the gasket and the slot in the recess must align Both slotted gaskets must be used even if running only one gel sandwich Grooves along each slot help keep the gasket in place Additionally a small amount of Gel Seal can be applied at each end of the gasket before install to help hold the gasket against the upper buffer chamber Release the sandwiches from the caster by removing all bottom cams if present Lower the upper buffer chamber onto the gel sandwiches in the casting Note Do not force the cams If you encounter unusual resistance disassemble and inspect clamp and glass alignment along the top of the sandwich Align and reinstall Note If the cooling option is used frequently it is convenient to attach QuickFit connectors to the tubing The valves in these fittings prevent coolant spillage stand Install the cams ridge pointing down into the buffer chamber cam holes Clamp the sandwich in place by simultaneously turning one cam clockwise and the other counterclockwise a full 180 o Use a pipette to carefully fill each slot above the sample wells with buffer to minimize disturbing the samples Then pour 100 ml of buffer into the chamber directing the buffer stream toward the side wall Check that no buffer lea
47. uffer dam Complete unit also includes spacers 4 and combs 2 Required but not included e Magnetic stirrer e Power supply with a minimum rating of 300 V 100 mA constant A or V Optional Circulator bath Note The ordering section lists all accessories and replacement parts color coded leads 2 safety lid interlock pins gt upper buffer La chamber with upper electrode heat exchanger with lower electrode lower buffer chamber Unpacking and Inventory Unwrap all packages carefully and compare contents with the packing list making sure all items arrived If any part is missing contact your local sales office Inspect all components for damage that may have occurred while the unit was in transit If any part appears damaged contact the carrier immediately Be sure to keep all packing material for damage claims or to use should it become necessary to return the unit Lower buffer chamber The lower buffer chamber is transparent which allows visual tracking of electrophoresis process The chamber is chemically resistant to common electrophoretic buffers but not to organic solvents or strong acids or alkali Temperatures above 45 C may cause the chamber to warp Upper buffer chamber The upper buffer chamber is chemically resistant to common electrophoresis buffers but not to organic solvents or strong acids or alkali The upper electrode cathode runs along the center ridg
48. well comb forms wells that are only 15 mm deep so that wells do not collapse when the comb is removed The sample volume held by each well depends on the gel thickness well depth and the number of wells per comb Table 1 lists sample volumes of wells for all combs see page 17 Wonder Wedge Gel Plate Separation tool This tool is used to disassemble gel sandwiches and to check spacer and comb thicknesses Operating Instructions Gel casting and electrophoresis procedures follow Included are instructions for polyacrylamide gels used with continuous or discontinuous buffer systems and gradient gels See page 31 for bibliography Prepare the Gel Sandwich Glass plates spacers and clamp sets are sized so that the assembled sandwich can be easily aligned to create the seal reguired first to cast the gel and then to run it For best results take extra care to align all components when assembling sandwiches One to four gels 18 x 16 cm can be assembled and run in the SE600 Chroma Both precast gels and self cast gels can be used To self cast multiple gels kits can be ordered separately The SE615 Multiple Gel Caster Kit holds up to 10 single gel sandwiches and the SE675 Gel Caster Kit holds up to four sand wiches See the accompanying gel caster User Manual for complete instructions To run four gels concurrently two accessory notched divider plates and two additional pairs of spacers are reguired Fig 2 Sandwich ass
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