Isoelectric focusing
Isoelectric focusing is an analytical purification technique that separates proteins, electrophoretically, according to their acidic or basic intrinsic charges i.e. their isoelectric point [1].
A polyacrylamide gel apparatus containing a pH gradient is used. The pH gradient is established by the subjection of a buffer-like mixture of many polyampholytes, each with a different isoelectric point, to electrophoresis. The protein sample is then loaded and a suitable voltage is applied to the electric field. The protein migrates to its characteristic isoelectric pH forming a band and, once reached, its electrophoretic mobility becomes zero; movement in the positive or negative direction is inhibited [2].
Isoelectric focusing can distinguish between proteins that differ by as little as one net charge i.e. a pI value of 0.01 [3].
References
- ↑ Alberts, B. Johnson, A. Lewis, J. Raff, M. Roberts, K. Walter, P. (2008) Molecular Biology of The Cell, 5th edition, New York: Garland Science. Page 521.
- ↑ Berg, J. Stryer, L. Tymoczko, J. (2007) Biochemistry, 6th Edition, New York: W.H Freeman and Company. Page 73.
- ↑ Berg, J. Stryer, L. Tymoczko, J. (2007) Biochemistry, 6th Edition, New York: W.H Freeman and Company. Page 73.