Amersham Biosciences Hoefer DALT User manual

80-6431-50 Rev A/12-98
Hoefer DALT System
DALT Electrophoresis Tank
DALT Gradient Maker
DALT Multiple Gel Caster
DALT Blotting Kit
User Manual

AmershamBiosciences
Important User Information
Please read this entire manual to fully understand the safe
and effective use of this product.
The exclamation mark within an equilateral triangle is intended to alert the
user to the presence of important operating and maintenance instructions in
the literature accompanying the instrument.
If you have any comments on this manual, please send them to us at:
Amersham Biosciences Inc.
Marketing Department
654 Minnesota Street
San Francisco, CA 94107 USA
AmershamBiosciences reservesthe righttomake changesinthe specifications
without priornotice.
Warranty and Liability
Amersham Biosciences guaranteesthat the product deliveredhasbeen thoroughly
testedto assurethat it meetsitspublished specifications.Thewarranty included inthe
conditions of delivery is valid only if the product has been installed and used according to
theinstructionssuppliedby Amersham Biosciences.
Amersham Biosciences shall in noevent beliableforincidental orconsequential
damages,including without limitation,lostprofits, loss ofincome,lossofbusinessoppor-
tunities, loss of use and other related exposures, however caused, arising from the faulty
and incorrect use of the product.
©1998Amersham Biosciences Limited.
All rights reserved.
No part of this publication may be reproduced, stored in a retrieval system or transmitted
in any form by any means, without permission in written form from the company.
English

Hoefer DALT System
Amersham Biosciences
Renseignements importants
d’utilization
Pour une bonne compréhension et une utilisation
en sécurité maximale, il convient de lire entièrement ce manuel.
Dans la documentation qui accompagne l’instrument un point d’excla-
mation dans un triangle équilatéral a pour but d’attirer l’attention de
l’utilisateur sur des instructions importantes de fonctionnment ou de
maintenance.
Tous vos commentaires sur ce manuel seront les bienvenus et veuillez les adresser
à:
Amersham Biosciences Inc.
Marketing Department
654 Minnesota Street
San Francisco, CA 94107 USA
Amersham Biosciences seréserveledroitd’effectuerdesmodificationsde
ces spécifications sans aucun préavis.
Garantieet responsabilité
Amersham Biosciences garantit àl’utilisateurqueleproduitlivréa subi
avec succèstouslesessaisprévuspours’assurerqu’il estconforme aux spécifica-
tionsetnormesenvigueur. Lagarantieinclusedanslesconditionsdelivraison
n’est valablequesi leproduit a étéinstalléetutiliséconformément aux instruc-
tions fournies parAmershamBiosciences.
Lasociété AmershamBiosciences nesera enaucuncasresponsable de
tout dommagecausédirectementouindirectementpartouteutilisation incor-
recteou nonapprouvée du produit oudécoulant decetteutilisation, y compris
toute perte de bénéfice ou de recettes, toute perte de perspectives commerciales,
tout empêchement d’utilisation ettoutautrerisquesayantunrapportavec l’util-
isation du produit, mais sans aucune limitation quant à la nature de ces
dommages.
©1998AmershamBiosciences Limited.
Tous droits réservés.
La reproduction, le stockage dans un système de récupération d’informations ou
la transmission sous quelque forme que ce soit et par quelque moyen que ce soit
de la présente publication en totalité ou en partie sont strictement interdits sans
autorisation préalable écrite de la société.
Información importante para el usuario
Para comprender el producto y utilizarlo con segu-
ridad es necesario leer este manual en su totalidad.
El signo de admiración en un triángulo
equilátero en el manual, advierte al usuario sobre la presencia de
instrucciones importantes de operación y mantenimiento del aparato.
Si desearan hacer algún comentario sobre este manual, tengan la amabilidad de
remitirlo a:
Amersham Biosciences Inc.
Marketing Department
654 Minnesota Street
San Francisco, CA 94107 USA
Amersham Biosciences sereserva el derecho amodificarlasespecifica-
ciones sin previo aviso.
Garantía y responsabilidad
Amersham Biosciences garantizaqueel productoentregado hasidopro-
badoafondo paracomprobarelcumplimientodelas especificacionespublica-
das. La garantía incluida en las condiciones de entrega sólo es válida si el
producto se ha instalado y utilizado de acuerdo con las instrucciones entregadas
porAmersham Biosciences.
Amersham Biosciences noseráresponsable, bajo ningún concepto, de
daños directos o indirectos, incluyendo sin limitación la pérdida de beneficios, la
pérdida de ingresos, la pérdida de oportunidades de negocio, la pérdida de uti-
lización y otras consecuencias relacionadas, cualquiera que sea la causa, que se
deban a lautilización defectuosaeincorrecta del producto.
©1998Amersham Biosciences Limited.Reservadostodos
los derechos.
No está permitida la reproducción, ni el almacenaje en un sistema de recuper-
ación, ni la transmisión de parte alguna de esta publicación sin la autorización
por escrito de la empresa.
Wichtige Benutzerinformationen
Für ein vollständiges Verständnis und eine sichere
Handhabung dieses Produktes ist es notwendig, daß
der Benutzer dieses Handbuch vollständig durchliest.
Ein Ausrufezeichen in einem gleichseitigen Dreieck soll den
Benutzer auf die Anwesenheit wichtiger Betriebs- und Wartungsan-
weisungen in der dem Gerät beiliegenden Dokumentation hin-
weisen.
Wenn Sie Anmerkungen zu diesem Handbuch haben, dann senden Sie diese bitte
an:
Amersham Biosciences Inc.
Marketing Department
654 Minnesota Street
San Francisco, CA94107 USA
AmershamBiosciences behältsichdas Rechtvor,die Spezifikationen
ohne vorhergehende Ankündigung zu ändern.
Gewährleistung and Haftung
Amersham Biosciences garantiert, daß dasgelieferte Produkt sorgfältig
aufdieEinhaltungderveröffentlichtenSpezifikationen getestetwurde.Diein
den Lieferbedingungen näher erläuterten Gewährleistungsansprüche gelten nur
dann,wenn dasProdukt gemäß denvon AmershamBiosciences
geliefertenAnweisungeninstalliertund benutzt wurde.
Amersham Biosciencces übernimmt keinerlei Haftung fürSchädenoder
Folgeschäden, einschließlich, aber nicht begrenzt auf Gewinneinbußen, Einkom-
mensverluste,entgangeneGeschäftsabschlüsse,VerlustderGebrauchsfähigkeit
oder andere Verluste, die wie auch immer durch eine fehlerhafte oder
unsachgemäßeVerwendung desProduktsverursachtwurden.
©1998AmershamBiosciences Limited.
Alle Rechte vorbehalten.
Die vorliegende Veröffentlichung darf nur mit vorhergehender schriftlicher Gene-
hmigung durch das Unternehmen vervielfältigt, in einem Abrufsystem gespe-
ichertoderin irgendeinerForm odermit irgendwelchenMitteln übertragen
werden.
Informazioni importanti per l’operatore
Per un utilizzo sicuro del prodotto, leggere attentam-
ente l’intero contenuto del presente manuale.
Il punto esclamativo all’interno di un
triangolo equilatero indica all’operatore la presenza di importanti
istruzioni di funzionamento e manutenzione nella documentazione
allegata al prodotto.
Si prega di inviare eventualicommential presentemanuale a:
Amersham Biosciences Inc.
Marketing Department
654 Minnesota Street
San Francisco, CA 94107 USA
Amersham Biosciecnes si riserva il dirittodi apportaremodificheai dati
tecnici senza preavviso.
Garanziae responsabilitá
Amersham Biosciences garantiscecheprimadellaconsegna il prodottoè
stato collaudato a fondo per soddisfare i requisiti specificati. La garanzia inclusa
nelle condizioni di consegna risulta valida solamente se il prodotto è stato instal-
lato edutilizzatonel rispetto delleistruzioni fornitedaAmersham
Biosciences
Amersham Biosciences non potrà essereritenuta responsabiledi incidenti
o danni consequenziali, inclusi’ma non limitati’a perdite di profitti, mancato
guadagno, perditedi affari,difetti di funzionamento erelativeesposizioni, dovuti
ad un utilizzo non corretto del prodotto.
©1998AmershamBiosciences Limited.
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Hoefer DALT System
AmershamBiosciences
CONTENTS
Hoefer DALT System Function and Description 3
DALT System Components 3
Unpacking 7
Important Information/Informations Importantes 8
Specifications 9
Required or Convenient, But not Supplied 11
Preparing the Gel Caster 12
Casting Homogeneous (Non-Gradient) Gels 14
Preparation for Gradient Gel Casting 16
Configuring the Gradient Divider 16
Calibrating the Peristaltic Pump 17
Casting Gradient Gels 18
Gradient Casting Setup 18
Pouring Gel Solutions for Gradient Gels 19
Applying Overlay to DALT Slab Gels 22
Polymerization 22
Unloading the Gel Caster 23
Preparing the DALT Tank for Electrophoresis 24
Connecting a Refrigerated Circulating Bath 24
Filling the Tank with SDS Electrophoresis Buffer 24
Loading and Running Second Dimension Gels 27
Equilibrating IPG Strips 27
Loading the IPG Strips onto the DALT Slab Gels 29
Loading Cassettes into the DALT Tank 31
Electrophoresis Conditions in the DALT Tank 32
Unloading the DALT Cassettes 33
Cleaning DALT Cassettes 33
The DALT Blotting Kit 34
Preparing the DALT Tank for Transfer 34
Assemble the Transfer Cassette 35
Loading the Cassettes 37

Hoefer DALT System
2AmershamBiosciences
Connecting the Power Supply 38
Transfer Conditions 39
Completing the Transfer 39
Factors Affecting the Transfer 40
Recipes 41
Homogeneous Gel Solutions 46
Gradient Gel Solutions 49
For 25 gels, 1 mm 50
For 12 gels, 1 mm 51
For 23 gels, 1.5 mm 52
For 12 gels, 1.5 mm 53
Gel Identification Numbers 54
Care and Maintenance 55
Cleaning 55
Removing the Electrode Panels from the DALT Tank 55
Troubleshooting 56
References 60
Customer Service Information 61
Technical Service and Repair 61
Ordering information 61
Index 63

Hoefer DALT System Hoefer DALT System Function and Description
Amersham Biosciences 3
Hoefer DALT System Function and Description
In 2-D protein electrophoresis, proteins are separated first according to isoelectric point
by isoelectric focusing, most reliably on immobilized pH gradient (IPG) strips. The
second dimension electrophoresis separates the proteins on the basis of their molecular
mass on a slab gel containing the denaturing detergent, sodium dodecyl sulfate (SDS).
The Hoefer DALT System is designed to simplify the handling of multiple second
dimension gels and improve the reproducibility of the second dimension separation.
DALT System Components
The Hoefer DALT System comprises:
• a multiple vertical slab electrophoresis tank
• a multiple gel caster
• a gradient maker with peristaltic pump
• gel cassettes
• a blotting kit
Electrophoresis Tank
Figure 1. The DALT
Electrophoresis Tank
The DALT Electrophoresis Tank accommodates up to ten 23 ×19 cm slab gels for
separation under identical conditions. The sample, following focusing in an IPG strip, is
attached to the cathodic surface of the gel by embedding in agarose, then the gel is
turned 90° and run in a “sideways” orientation, with the IPG strip standing vertically at
one edge.
The tank is divided into three chambers by silicone rubber flap seals on the barrier
combs. The seals are not designed to be liquid-tight but they do provide a good barrier
to electrical current, causing most current to flow through the gels. Although there is
Hinged lid
Platinum-core
electrode panels (two)
Barrier combs (two) with
silicone rubber flap seals
Buffer
circulation flutes
Alumina
heat exchanger
High-voltage leads
(Siphon pump included,
but not shown)

Hoefer DALT System Function and Description Hoefer DALT System
4AmershamBiosciences
some leakage current bypassing the gels, this has little effect on system operation. The
barriers can be raised or removed by loosening the nylon screws which secure them in
place.
The left and right chambers contain platinum wire electrodes, cathode (–) and
anode (+). The center chamber provides cooling via circulating buffer and a tube-type
heat exchanger. Since the gel cassettes are almost entirely exposed to the buffer in the
center chamber, with only their ends protruding into the side chambers, cooling of the
center chamber buffer provides excellent temperature control of the gel slabs during the
run. Electrode panels can be removed by loosening the nylon screws securing them in
place. Removal is not typically necessary unless the platinum wire becomes broken and
requires replacement. See “Removing the Electrode Panels from the DALT Tank” on
page 55.
Figure 2. Rear view of
electrophoresis tank
Buffer moves into the pump through the middle of the three flutes running along the
length of the tank’s center chamber, and is expelled back into the tank through the left
and right flutes. The holes in the output flutes are aligned to generate maximum
circulation over both the cassettes and the heat exchanger on the floor of the tank's
centre chamber (beneath the flutes). As described on page 24, run the pump only when
the tank is filled with liquid, to prevent damage to the pump.
The Quick-fit connectors on the ends of the tank port tubing provide a leak-proof seal
when disengaged, eliminating spillage of coolant solution.
Pump shield, with foot
for open lid support
Buffer
circulation pump
Pump priming line*
*WARNING The liquid in the tank must cover the opening to the pump priming
line when the circulation pump is in operation.
Quick-fit connectors,
attach to circulating
water bath

Hoefer DALT System Hoefer DALT System Function and Description
AmershamBiosciences 5
Gel Cassettes
Figure 3. DALT gel
cassettes
The DALT gel cassettes are pre-assembled. Two glass plates are held together along one
edge by a strip of silicone rubber, and the glass spacers (1.5 or 1.0 mm thick) are glued
in position. To complete assembly, close the two plates like a book. Gels are removed by
opening the book after the run and lifting out the slab. The cassette is easily cleaned as a
unit, and can be stood upright in the open to dry. The cassettes are dishwasher-safe.
Cassettes are 20.4 ×25.5 cm and produce a gel about 19 ×23 cm: 46 ml for 1-mm
thick, 68 ml for 1.5-mm thick. The gels make the best use of the standard 20 ×25 cm X-
ray film.
Multiple Gel Caster
Figure 4. The DALT Gel
Caster
The DALT Gel Caster holds up to 25 gel cassettes, 1 mm thick, or up to 23 cassettes,
1.5 mm thick, with separating sheets, for casting homogeneous or gradient gels. Fewer
cassettes can be filled at one time by inserting filler blocks to occupy unneeded volume.

Hoefer DALT System Function and Description Hoefer DALT System
6AmershamBiosciences
A removable front plate and cassette separating sheets simplify loading and unloading
cassettes from the unit. A hydrostatic balance chamber allows accurate gradient
positioning and provides a means to flush the fill tubing before polymerization occurs.
The chamber also provides a “make up solution” to accommodate the volume
shrinkage during polymerization.
Pump-Assisted Gradient Maker
Figure 5. The DALT
Gradient Maker
The DALT Gradient Maker casts gradients of arbitrary shape, up to 2.0 liters total
volume. The solution is delivered at a controlled rate by a peristaltic pump. A flexible
liquid-tight divider partitions the gradient maker into two chambers of complementary
shapes for casting both linear and non-linear gradients. With the divider adjusting rod,
you can configure the gradient divider to adapt to your gradient gel requirements.
Divider adjusting rod