Thermo Scientific™

High-Select™ TiO2 Phosphopeptide Enrichment Kit

Catalog number:  A32993

Related applications:

Protein Mass Spectrometry Analysis

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Description

The Thermo Scientific™ High-Select™ TiO2 Phosphopeptide Enrichment Kit enables efficient enrichment of phosphorylated peptides from complex and fractionated protein digests for analysis by mass spectrometry (MS). (This product replaces Cat. No. 88301.)

Features of the TiO2 Phosphopeptide Enrichment Kit:
• Convenient—pre-formulated buffers for ease of use and spin-tip format to enable parallel processing of multiple samples
• High binding capacity—each column enriches up to 150 μg of phosphopeptides
• High specificity—recovers phosphopeptides with > 90% selectivity
• Complementary—TiO2 enriches a unique set of phosphopeptides that complements the High-Select™ Fe-NTA IMAC Phosphopeptide Enrichment Kit

The complete kit includes 24 titanium dioxide (TiO2) spin tips with optimized buffers to facilitate preparation of enriched phosphopeptides. Each column can enrich phosphopeptides from 0.5 to 3 mg of total protein digest as starting sample. The optimized buffers and simplified procedure eliminate the need for pyrrolidine in the elution buffer and subsequent clean-up using graphite spin columns. Use of this kit results in enrichment of up to 150 µg of phosphopeptides with greater than 90% selectivity.

Mass spectrometry is a key tool for identifying sites of protein phosphorylation and quantifying phosphorylation changes; however, MS analysis of protein phosphorylation is challenging due to the low stoichiometry, high hydrophilicity, poor ionization, and incomplete fragmentation of phosphopeptides. Because of the low relative abundance of phosphorylation modifications in complex protein samples, enrichment is essential for successful MS analysis of phosphopeptides. The improved High-Select TiO2 Phosphopeptide Enrichment Kit complements our lysis, reduction, alkylation, and digestion reagents, along with our C18, graphite spin, and high-pH reversed-phase peptide fractionation columns to provide a complete workflow for phosphopeptide enrichment.

The new procedure and optimized buffers have been shown to maintain the excellent selectivity and yield of the previous kit (Figure 1), while removing hazardous reagents and clean-up. The High-Select TiO2 Phosphopeptide Enrichment Kit is complementary to the High-Select Fe-NTA Phosphopeptide Enrichment Kit, with both kits enriching similar numbers of phosphopeptides but different subpopulations of phosphorylated peptides (Figure 2).

For high resolution analysis of the enriched phosphopeptides, the recommended LC column for Nanospray Flex source is Acclaim PepMap 100 C18 LC Column (Cat. No. 164942 or 164939). For EASY-Spray source, the recommended LC column is EASY-Spray C18 LC Column (Cat. No. ES802 or ES803).

Specifications

Label Type: Unlabeled
Product Line: High-Select™
Shipping Condition: Wet Ice
Workflow Step: Peptide Enrichment
Detection Method: Mass Spectrometry
Final Product Type: Peptide
For Use With (Equipment): Microcentrifuge
Starting Material: Protease-digested protein
Description: Enriches TiO2 phosphopeptide from complex samples
Format: 24 reactions

Contents & storage

• TiO2 Spin Tips (24)
• Centrifuge Column Adadptors (24)
• Binding/Equilibration Buffer (7 mL)
• Wash Buffer (1.8 mL)
• Phosphopeptide Elution Buffer (7 mL)