The process and performance of phosphoprotein enrichment using optimized large-scale functionalized NPs. (a) Workflow illustrating capture and enrichment process of phosphoproteins from a mixture of standard proteins or a biological sample using the Fe3O4-GAPT-Zn NPs. SDS-PAGE analysis with (b) SYPRO Ruby and (c) Pro-Q Diamond stains of a standard protein mixture containing 300 μg BSA, 100 μg β-casein, and 100 μg pepsin, respectively. Equal amount (10 μg) of the loading mixture (LM), flow-through (FT), and elution mixture after enrichment (EM) was loaded on the gel. Phosphorylated pepsin and β-casein are labeled in the Pro-Q EM showing the highly specific enrichment of phosphoproteins from the standard protein mixture.