Question

How is PAGE (Polyacrylamide Gel Electrophoresis) different from other protein purification techniques?

Answer 1

Polyacrylamide Gel Electrophoresis, is an analytical technique used to separate components of a protein mixture based on their size.The technique is based upon the principle that a charged molecule will migrate when placed in the presence of an electric field towards an electrode with opposite sign. The general electrophoresis techniques cannot be used to determine the molecular weight of biological molecules because the mobility of a substance in the gel medium depends on both charge and size.

To overcome this problem the biological samples are treated so that they can obtain a uniform charge, then the electrophoretic mobility depends only on size. For this purpose different protein molecules having different shapes and sizes is denatured . Denaturation leads to lose their secondary, tertiary or quaternary structure .The proteins are covered by SDS and are negatively charged and when loaded onto a gel and placed in an electric field, it will migrate towards the anode are separated by a molecular sieving effect based on size. After the visualization by a staining technique, the size of a protein can be calculated by comparing its migration distance with that of a known molecular weight.

The main difference between page and other purification techniques is all the matter of size.

Agarose gel electrophoresis can resolve much larger molecules, such as DNA after PCR .

PAGE is good for analysis of small nucleic acids such as tRNAs, oligonucleotides, miRNA and proteins.