This Application Note describes the design and performance of an HPLC-Chip with a hybrid TiO2 and C18 enrichment column for selective phosphopeptide enrichment and analysis.
The document details the Agilent Phosphochip, a reusable microfluidic HPLC-Chip designed for the selective enrichment and analysis of phosphopeptides. The chip features a unique sandwiched enrichment column containing titanium dioxide (TiO2) beads flanked by C18 reversed phase material, integrated directly with an analytical column and nanospray emitter to automate the workflow.
The note outlines the experimental setup using the Phosphochip kit (64240-62020) with an Agilent 1200 Series HPLC-Chip/MS system. It describes the operation in two modes: a single mode for phosphopeptide analysis involving trapping and elution steps, and a dual mode for simultaneous analysis of phosphorylated and non-phosphorylated peptides. Results from analyzing bovine protein mixtures and human cell lysates are presented to validate the enrichment performance.
| Kit Part Number | 64240-62020 |
|---|---|
| Compatible MS Systems | Chip/lon Trap, Chip/QQ0 or Chip/QTOF |
The Phosphochip is a reusable microfluidic chip specifically designed for phosphopeptide enrichment and analysis.
The enrichment section contains titanium dioxide beads flanked on both sides with C18 reversed phase material.
The proteins were digested using a protein/protease ratio of 50:1.