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

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Semidry Electroblotter 5-1 Thermo Scientific
Section 5
Technical Tips
How long will it take to blot the proteins from my gel?
Transfer times have to be determined experimentally. This is because
transfer time is dependent upon:
• Percentage of gel
• Type and amount of cross linking in the gel
• Type of protein: cytoplasmic, membrane, nuclear
• Size of protein
There is no formula for determining transfer time. There are too many
variables involved to give specific transfer conditions that will work for
every protocol.
Guidelines are:
• HEP-l: 2mA/cm2 of gel for 1 hour
These guidelines are just a starting point and exact conditions have to be
determined.
Different kinds of blotting
Western Blotting is a blotting method for proteins that use specific
antibodies attached to particular protein to help identify it. It is often
performed after SDS-PAGE or some other form of polyacrylamide gel
electrophoresis.
Southern Blotting is a method sometiems called hybridization because a
radioactive probe is "hybridized" or attached to specific pieces of DNA.
Northern Blotting is a similar method but the molecules involved are
RNA.
Both Southern and Northern blotting generally require the DNA or RNA
to first be separated out on an agarose gel.