| Title | Comparison of IMAC and MOAC for phosphopeptide enrichment by column chromatography |
| Publication Type | Journal Article |
| Year of Publication | 2012 |
| Authors | Negroni L, Claverol S, Rosenbaum J, Chevet E, Bonneu M, Schmitter J-M |
| Journal | Journal of Chromatography B |
| Volume | 891-892 |
| Pagination | 109 - 112 |
| Date Published | 4/2012 |
| ISSN | 15700232 |
| Abstract | Automated phosphopeptide enrichment prior to MS analysis by means of Immobilized Metal Affinity Chromatography (IMAC) and Metal Oxide Affinity Chromatography (MOAC) has been probed with packed columns. We compared POROS-Fe(3+) and TiO(2) (respectively IMAC and MOAC media), using a simple mixture of peptides from casein-albumin and a complex mixture of peptides isolated from mouse liver. With theses samples, selectivity of POROS-Fe(3+) and TiO(2) were pH dependant. In the case of liver extract, selectivity increased from 12-18% to 58-60% when loading buffer contained 0.1M acetic acid or 0.1M trifluoroacetic acid, respectively. However, with POROS-Fe(3+) column, the number of identifications decreased from 356 phosphopeptides with 0.1M acetic acid to 119 phosphopeptides with 0.1M TFA. This decrease of binding capacity of POROS-Fe(3+) was associated with strong Fe(3+) leaching. Furthermore, repetitive use of IMAC-Fe(3+) with the 0.5M NH(4)OH solution required for phosphopeptide elution induced Fe(2)O(3) accumulation in the column. By comparison, MOAC columns packed with TiO(2) support do not present any problem of stability in the same conditions and provide a reliable solution for packed column phosphopeptide enrichment. |
| DOI | 10.1016/j.jchromb.2012.02.028 |
| Short Title | Journal of Chromatography B |
