Protein phosphorylation, a post-translational modification regulated by kinases and phosphatases, is essential for numerous cellular functions such as gene expression and membrane transport, so identification of phosphorylation sites is vital to understanding many biochemical processes. While mass spectrometry is a useful technique for identifying such sites, low ionization efficiencies of phosphorylated fragments or a low degree of phosphorylation in a given sample can make detection of phosphorylated species difficult. To overcome this challenge, immobilized metal affinity chromatography (IMAC) can be used to enrich phosphopeptides prior to analysis. IMAC isolates phosphopeptides based on their affinity for a metal-ligand complex that is immobilized on a chromatographic support, but the use of this technique complicates analyses and decreases throughput because the sample mixture must be loaded onto a column, rinsed to remove unbound species, and then eluted to collect the fragments of interest. We are developing a similar, but simpler, procedure to capture phosphopeptides directly on a modified gold matrix-assisted laser desorption/ionization sample plate. We immobilize nitrilotriacetate (NTA) on the surface of a MALDI plate using the chemistry shown below and subsequently form the Fe(III)-NTA complex. Addition of unpurified sample to thse modified plates for a 5-10 min incubation period, followed by rinsing of the plate to remove unbound peptide fragments and other contaminants yield a surface that is highly enriched in phosphopeptides. Subsequent addition of matrix allows analysis by MALDI MS. The resulting mass spectra show greatly enhanced signals due to phosphopeptides. To see an example on a phosphopeptide-enriched mass spectrum, click here. This work was recently described in a paper in Analytical Chemistry.