The picture shows guide for choosing gel % but there is overlap between protein sizes. Also don't know which to choose if have proteins of different sizes. Tahnks
If you are interested in separating proteins on the same gel that have greatly different molecular weights, it may be necessary to prepare (or preferably buy if you can) a gradient gel, in which the acrylamide concentration varies from top to bottom. It's obviously more difficult to prepare such a gel.
An alternative is to run two gels, a high percentage gel to separate the small proteins and a low percentage gel to separate the large proteins. However, if the sizes of the proteins you wish to separate are not too far apart, you should be able to separate them on a single gel.
Remember, if this is denaturing, reducing SDS-PAGE, only the molecular weights of the individual polypeptide chains matter. Oligomers will be dissociated into monomers. For example, if your protein is a 400 kDa tetramer of 100 kDa monomers, you will only see the 100 kDa monomer band.
If you are interested in separating proteins on the same gel that have greatly different molecular weights, it may be necessary to prepare (or preferably buy if you can) a gradient gel, in which the acrylamide concentration varies from top to bottom. It's obviously more difficult to prepare such a gel.
An alternative is to run two gels, a high percentage gel to separate the small proteins and a low percentage gel to separate the large proteins. However, if the sizes of the proteins you wish to separate are not too far apart, you should be able to separate them on a single gel.
Remember, if this is denaturing, reducing SDS-PAGE, only the molecular weights of the individual polypeptide chains matter. Oligomers will be dissociated into monomers. For example, if your protein is a 400 kDa tetramer of 100 kDa monomers, you will only see the 100 kDa monomer band.
It actually depends on the difference of your protein sizes. Suppose, your proteins are of 15 kDa and 50 kDa then you need to use 15% and 12.5% gels respectivly.
Hello Heba,
Please check the attached protocol.
http://www.abcam.com/protocols/general-western-blot-protocol
Best regards,
It completely depends on the the protein molecular weight. Tell your protein size if you want exact gel percentage.
http://www.bio-rad.com/webroot/web/pdf/lsr/literature/Bulletin_6040.pdf
- Badges
- Science method