Ion exchange chromatography: Difference between revisions
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Ion-exchange [[Chromatography|chromatography]] separates [[Proteins|proteins]] according to their net charge. The stationary phase of the column will have a specific charge. | Ion-exchange [[Chromatography|chromatography]] separates [[Proteins|proteins]] and/or molecules according to their net charge. The stationary phase of the column will have a specific charge which is created by ion-exhcnage resins which come in two forms: polyanions (negatively charged) and polycations (positively charged). | ||
For example negatively charged [[Proteins|proteins]] ([[Anions|anions]]) will bind to a positively charged [[ | For example negatively charged [[Proteins|proteins]] ([[Anions|anions]]) will bind to a positively charged [[Diethylaminoethylcellulose|diethylaminoethylcellulose ]](DEAE-cellulose) columns. The negative proteins bound to column can by eluted by adding a negatively charged [[Buffer|buffer]] which will compete with the [[Protein|protein]] for binding to the column. Proteins that have lower charge density will be eluted first from the column <ref>Berg J., Tymoczko J and Stryer L. (2007) Biochemistry, 6th edition, New York: WH Freeman</ref>. | ||
The stationary phase is an insoluble matrix which will either carry a positve or negative charge. Whilst diethylaminoethylcellulose (DEAE-cellulose) is positively charged, an negatively charged matrix includes [[ | The stationary phase is an insoluble matrix which will either carry a positve or negative charge. Whilst diethylaminoethylcellulose (DEAE-cellulose) is positively charged, an negatively charged matrix includes [[Carboxymethylcellulose|carboxymethylcellulose]] (CM-cellulose) <ref>Alberts et al. (2008) Molecular Biology Of The Cell, 5th Edition. Chapter 8, page 514.</ref>.<br> | ||
=== References === | === References === | ||
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<references /> | <references /> | ||
3. Matthews, Van Holde, Ahern (2000) Biochemistry; Third Edition; P151 | |||
Revision as of 11:45, 29 November 2012
Ion-exchange chromatography separates proteins and/or molecules according to their net charge. The stationary phase of the column will have a specific charge which is created by ion-exhcnage resins which come in two forms: polyanions (negatively charged) and polycations (positively charged).
For example negatively charged proteins (anions) will bind to a positively charged diethylaminoethylcellulose (DEAE-cellulose) columns. The negative proteins bound to column can by eluted by adding a negatively charged buffer which will compete with the protein for binding to the column. Proteins that have lower charge density will be eluted first from the column [1].
The stationary phase is an insoluble matrix which will either carry a positve or negative charge. Whilst diethylaminoethylcellulose (DEAE-cellulose) is positively charged, an negatively charged matrix includes carboxymethylcellulose (CM-cellulose) [2].
References
3. Matthews, Van Holde, Ahern (2000) Biochemistry; Third Edition; P151