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whatis the relationship among voltage, protein size, and length of transfer?

whatis the relationship among voltage, protein size, and length of transfer? - Western Blot Forum

whatis the relationship among voltage, protein size, and length of transfer? - Discuss western blotting and immunoblot in the western blot forum discussion board. Ask Questions about transfers, blocking, membrane antibody incubation and exposures.


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Old 11-09-2011, 02:55 AM
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Default whatis the relationship among voltage, protein size, and length of transfer?



Hi everyone. I am now focusing on a protein which has the size of 44KD. I am using 15% SDS-PAGE for electrophoresis, nitrocellulose membrane, and Bio-Rad semi-dry transfer system. For transfer, I tried 20V for 20 min. The marker transfered well but there were still lots of proteins left on the gel which I saw in coomassie blue staining. Then the bands were wery weak on he film.
Does everyone can give me some advices for the transfer condition or tell me how to calculate the optimal voltage and minutes for transfer? Thanks

William
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Old 11-14-2011, 06:57 AM
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Default Re: whatis the relationship among voltage, protein size, and length of transfer?

Hola Will, in my experience the transference of proteins is similar to the displacement in a PAGE. I explain: as more soft is the gel more displacement of proteins,well you will have more protein transferred (at the same time) , if you use a 10-12% of acrylamide gel, but always is a remaining amount of protein in the gel. Moreover you can observe more transference in the low MW proteins than high ones. Some people recomend doubling SDS conc in the transfer buffer and of course increase the time at this voltage to 30 min.Buena suerte
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