Immobilized metal affinity chromatography, or IMAC, has been employed for many applications making use of its versatile interaction with aromatic nitrogens and phosphates. To this end, NTA matrices have been loaded with a broad range of transition metals, as optimized for the individual interaction. Magnetic beads are ideal for protein purification from dilute supernatants and for pull-down experiments. Fe-NTA MagBeads are often used for the purification of phosphorylated proteins. Alternative matrices are Al-, Ga- or Zr-NTA. PureCube Fe-NTA MagBeads are delivered as a 25% suspension.
PureCube Fe-NTA MagBeads provide:
Special IMAC Magnetic Beads from Cube Biotech were successfully used in the following publications:
Purification of phosphorylated proteins
IMAC methods, in particular Fe-NTA or Ga-NTA, have been widely used to enrich phosphorylated proteins and phosphopeptides as part of the sample preparation for mass spectrometry (1,3,4). Under certain circumstances other transition metals, such as zirconium or aluminium have also been loaded on NTA or IDA matrices for enrichment (5). Magnetic beads have proven to be useful for this kind of sample preparation (2) in a similar manner to their agarose resin counterparts.
||Specific binding of phosphorylated biomolecules (e g. proteins)
||Affinity to phosphorylated biomolecules (e g. proteins)
||Depending on the application
||Delivered as a 25 % suspension
|Metal Ion capacity
||>12 µeqv Fe2+/mL.
1. Albuquerque, C.P. et al. A multidimensional chromatography technology for in-depth phosphoproteome analysis. Mol Cell Proteomics (2008), 7(7), 1389-1396.
2. Herskowitz, J., et al. Phosphoproteomic analysis reveals site-specific changes in GFAP and NDGR2 phosphorylation in frontotemporal lobar degeneration. J. Proteome Res (2010), 9(12):6368-6379.
3. Yu, P. et al. Global analysis of neuronal phosphoproteome regulation by chondroitin sulfate proteoglycans. PLoS One (2013), 8,3, e59285.
4. Aryal, U.K. et al. Optimization of immobilized Gallium (III) ion affinity chromatography for selective binding and recovery of phosphopeptides from protein digests. Journal of Biomolecular Techniques (2008), 19:296-310.
5. Block et. al. Immobilized-metal affinity chromatography (IMAC) a review. Methods Enzymol. (2009), 463:439-73.
6. Searle, B. et. al. (2019). Thesaurus: quantifying phosphopeptide positional isomers. Nature Methods. 16. 703-706. 10.1038/s41592-019-0498-4..
7. Searle, Brian et al. (2018). Chromatogram libraries improve peptide detection and quantification by data independent acquisition mass spectrometry. Nature Communications. 9. 10.1038/s41467-018-07454-w