Thermo Scientific OWL HEP-1, Руководство по эксплуатации и техническому обслуживанию

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Semidry Electroblotter 2-1 Thermo Scientific
Section 2
Setting Up
Once the proteins (or nucleic acids) in a sample aliquot have been
separated on a slab gel, the resulting bands may be transferred to a solid
support membrane. The primary reason for this type of blot is one of
localization and secondarily, concentration of discrete bands. Although
many have used alternative cross linking agents such as DATD
(N,N'dihydroxyethylene-bis Acrylamide) to allow for the accessibility of
gel bound proteins, this still represents an impediment to radio
enumeration due to the quenching by the gel matrix itself.
The most common solid support membrane is nitrocellulose. A second
type of membrane is PVDP (Polyvinylidene difloride) which is generally
used when a transferred protein is to be sequenced, additionally, it has a 2x
binding capacity. Also used for nucleic acid capture are Nylon membranes.
In either case, the proteins are transferred from the gel to the matrix in an
electric field perpendicular to the gel (initial running direction).
Tris based buffers are employed in the transfer. Methanol and SDS are
modifiers often use in protein transfer buffer. These components however
are antithetical in their effects both in terms of movement and adsorption.
Methanol restricts protein movement from the gel but is often required to
support the ionic nature of protein to nitrocellulose binding. SDS aids in
protein elution, but can also inhibit binding of small molecular weight
proteins (Mozdzanowski, J., High yield electroblotting, Electrophoresis,
1992, Vol l3.,p.59-64).
In order to use this blotting device, you will need:
Power supply - Blotting requires a power supply that can operate at a fairly
high current setting and low voltage. If an inappropriate power supply is
used, the power supply may blow a fuse, shut itself off, display a no load or
short load message or even have a short circuit. It is very important to be
sure that the power supply you will be using will work with this device.
Some power supplies that will work with this device are Catalog No. OSP-
135, OSP-300, EC Apparatus EC135 and EC570, and Bio Rad's
PowerPac 200. If your power supply is not among those listed, contact the
manufacturer of your power supply to determine if it will work for tank
blotting applications. Contact Technical Services for power supply
recommendations if you do not have an appropriate power supply.
Materials Needed