An Exfoliated 2D Egyptian Blue Nanosheet for Highly Selective Enrichment of Multi-phosphorylated Peptides in Mass...

Phosphopeptide enrichment is essential for the phosphoprotein profiling, due to the low abundance in complex biological samples. Moreover, selective binding of multi-phosphopeptides over mono-phosphopeptides is rarely established, but strongly needed in real sample analysis, especially for the investigation of cell behaviors with the multisite phosphorylation cascades. Here two-dimensional (2D) nanosheets were synthesized of Egyptian blue (CaCuSi₄ O₁₀ ), the well-known ancient pigment, and its analogues (SrCuSi₄ O₁₀ and BaCuSi₄ O₁₀ ), which were employed in the enrichment of phosphopeptides for the first time. Surprisingly, the 2D CaCuSi₄ O₁₀ nanosheet was highly selective towards multi-phosphopeptides without enriching the mono-phosphorylated peptides in a wide range of acidic conditions or buffer compositions. Meanwhile, the SrCuSi₄ O₁₀ and BaCuSi₄ O₁₀ nanosheet analogues do not exhibit this unique selectivity. Moreover, the ultrathin and well-defined 2D morphology, with abundant Ca^II , of Egyptian blue nanosheet was applied in cortical samples of forebrain specific PDK1 conditional knockout mice and their age-matched littermate controls, that are associated with Alzheimer's disease. The as-prepared 2D CaCuSi₄ O₁₀ nanosheet not only showed specific selectivity, but also exhibited high sensitivity (detection limit of 4×10^-7  m).