INSTRUCTION MANUAL Electrophoretic Blotting System EBU-102 EBU-102P
TABLE OF CONTENTS Page Important User Information 3-4 Section 1 1.1 1.2 1.3 General Information Introduction Specifications Safety 5 5 5 Section 2 2.1 2.2 2.3 2.4 Description of parts Unpacking EBU-102 or EBU-102P Components/Assembly Unpacking EBU-202 Components/Assembly 6 6 7 7 Section 3 3.1 3.2 3.3 3.4 Instructions for Use Blotting Unit Preparation Preparing the Gel for Transfer Standard Electro-Blot Transfer Removing the Cassettes 8 8-9 10 10 Section 4 4.
ENGLISH IMPORTANT USER INFORMATION This Instruction Manual will explain how to use this product safely and effectively. Please read and carefully follow the instruction manual in its entirety. The triangle/exclamation mark symbol alerts the user of the product to important operational, maintenance, and/or warranty requirements. The triangle/lighting bolt symbol alerts the user of the product to potentially hazardous electrical exposure.
FRANÇAIS INFORMATION IMPORTANTE À L'USAGE DES UTILISATEURS ESPAÑOL Le présent manuel d'utilisation explique la manière de se servir efficacement du produit en conditions de sécurité. Il est recommandé de soigneusement lire la totalité du manuel, avec ses consignes et ses instructions. El presente instructivo explica la manera de usar este producto en forma segura y efectiva. Sírvase leerlo en su totalidad y seguir detenidamente las indicaciones que contiene.
SECTION 1 General Information 1.1 Introduction Electroblotting is a faster, easier, and more efficient method to permanently transfer PAGE gels to nitrocellulose and nucleic acids to nylon, than capillary (passive diffusion) blotting. For a similar transfer, traditional “capillary blotting” can take 6-24 hours to complete compared to only 2 hours or less using electroblotting. The C.B.S.
SECTION 2 Description of Parts 2.1 Unpacking the EBU-102 or EBU-102P Please verify that your unit comes complete with the following components: • Blotting chamber • Blotting cassette with sponge pads • Safety cover with attached power leads • 2 electrode panels • Tubing adapters for cooling base 2.2 Components/Assembly Safety Cover Chamber Blotting Cassette Cooling Ports C.B.S.
2.3 Unpacking the EBU-202 Please verify that your unit comes complete with the following components: • Blotting chamber • Blotting cassette with Scotch-Brite pads • Safety cover with attached power leads • 2 electrode panels • Tubing adapters for cooling base 2.4 Components/Assembly Safety Cover Scotch Brite Pads Blotting Cassette Chamber Cooling Ports C.B.S.
SECTION 3 Instructions for Use 3.1 Blotting Unit Preparation 1. 2. Place the blotting chamber on a level work surface in an authorized work area. Consult applications table 4.1 for type of transfer, buffer system, membrane type and power settings.
the transfer membrane. Disturbing or moving the gel/membrane interface can result in a smeared blot. 7. Quickly follow with 1-2 sheets of saturated 3mm filter paper (optional). 8. Place second saturated Scotch-Brite pad over sandwich assembly and close the cassette. Cassette Assembly 9. Remove entire assembly from pyrex dish and load into desired position (slot) in buffer chamber with black screen facing towards black terminal (cathode) and red screen facing towards red terminal (anode). C.B.S.
Safety Cover Blotting Cassette Chamber C.B.S.
3.3 Standard Electro-Blot Transfer 1. Fill the chamber with transfer buffer. (Buffers should be prepared fresh with reagent grade chemicals and pre-cooled). The transfer buffer should come up to the top of the platinum labyrinth. The buffer should not come in contact with the banana plugs when the gel cassette sandwich(es) are immersed in the unit. 2. Align safety cover over the unit and carefully attach. Begin cycling of coolant. 3.
SECTION 4 Applications & Running Conditions for Tank Type Electro-Blotting 4.1 Recommended Buffers, Membranes, Power Settings and Transfer Times Gel Type Blotting Buffer Transfer Membrane Power denaturing: SDS-PAGE (Western) Towbin (25mM Tris/192mM Glycine/ pH 8.3/Methanol 20% (w/v) nitrocellulose/nylon PVDF: ion-exchange • • • non-denaturing: SDS-PAGE (acidic or neutral proteins) IEF (nativebasic proteins, acid urea gels) 25mMTris/192mM Glycine, pH 8.
4.2 References Ausubel, F.M., Brent, R., Kingston, R.E., Moore, D.D., Seidman, J.G., Smith, J.A., Struhl, K. (ed.) (1993). Current Protocols in Molecular Biology. Vol. 2, Greene Publishing Associates, Inc. and John Wiley & Sons, Inc., Ch.10. Burnette, W.N. (1981). Western blotting: Electrophoretic transfer of proteins from sodium dodecyl sulfate-polyacrylamide gels to unmodified nitrocellulose and radiographic detection with antibody and radioiodinated protein A. Anal. Biochem. 112:195-203. Peluso, R.W.
SECTION 5 Maintenance of Equipment 5.1 Care and Handling The plastic components of the Electrophoretic Blotting units are fabricated from acrylic and polycarbonate. Electrodes and connectors are made from pure platinum, stainless steel, and chrome plated brass. As with any laboratory instrument, adequate care ensures consistent and reliable performance. After each use, rinse buffer chamber, cassettes and Scotch Brite pads with de-ionized water.
SECTION 6 Equipment and Accessories Cat. # EBU-102 EBU-102P EBE-102 EBE-102P EBE-202 EBC-102 EBS-102 Item Electrophoretic Blotting System. Includes chamber with 2 platinum wire electrode panels, 1 color-coded gel cassette, safety cover with power leads Electrophoretic Blotting System, planar electrodes. Includes chamber with 2 planar electrode panels, 1 color-coded gel cassette, safety cover with power leads electrode panels, safety cover and interlocking power leads.
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