I have yet to see myosin heavy chain (MW at 200000) in the 2D gels I have run using the in-gel rehydration method. A recent newsletter from Pharmacia indicated that high molecular weight proteins are difficult to get into the IPG strip. Pharmacia used their IPGphor to apply 30V to the strip during sample rehydration to get the heavy proteins into the first dimension - from their gel images it looks like it worked very well.
The Multiphor unit seems to work well. It is quite versatile in that one could run a second dimension horizontal gel on the same basic apparatus. One would probably need to buy an extra kit to do this. For high throughput the IPGphor would be the best choice, however, it costs a bundle.
I have noticed that the IPG strips are not always consistent from batch to batch. Sometimes one batch of strips is half a cm longer than another batch. We are thinking of making our own IPG strips so that we can control the quality and gradient. Do not use the protocol from the manual supplied with the Immobile Drystrip kit - it is very outdated. A. Gorg has a great manual on 2D electrophoresis and making IPGs at:
http://www.edv.agrar.tu-muenchen.de/blm/deg/
I too would be very interested in other peoples experience in making their own IPG strips.
Michael Johns
Lacombe Research Centre
Agriculture and Agri-Food Canada
6000 C & E Trail
Lacombe, Alberta
Canada
T4L 1W1
Phone: 403-782-8100 ext. 268
Fax: 403-782-6120
johnsm@em.agr.ca
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