EASY-Spray and double nanoViper PepMap Neo columns - PRODUCT SPECIFICATIONS

Significance of Low-Flow Peptide Mapping

Low-flow nanoLC-MS peptide mapping has emerged as a cornerstone technique in proteomics, offering heightened sensitivity, improved chromatographic resolution and deep proteome coverage. By minimizing dead volume and reducing solvent consumption, laboratories can achieve sharper peaks, higher identification rates and more reliable quantification across large sample sets.

Objectives and Study Overview

This document presents the performance attributes of Thermo Scientific EASY-Spray and double nanoViper PepMap Neo columns when paired with the Vanquish Neo UHPLC System and an Orbitrap Exploris 480 mass spectrometer. The primary goals are to demonstrate:

• High column-to-column reproducibility over multiple injections
• Robust operation at pressures up to 1500 bar
• Improved peptide and protein identification and quantification consistency

Methodology and Instrumentation

Two column formats were evaluated using a direct injection workflow with a 200 ng HeLa cell digest:

• EASY-Spray PepMap Neo, 75 µm I.D. × 750 mm, C18, 2 µm, 100 Å, temperature-controlled emitter
• Double nanoViper PepMap Neo, 75 µm I.D. × 500 mm, C18, 2 µm, 100 Å, zero-dead-volume union

The UHPLC platform was the Thermo Scientific Vanquish Neo system, and detection was performed on a Thermo Scientific Orbitrap Exploris 480 mass spectrometer. Key design features included:

• Integrated ZDV connections to minimize extra-column volume
• High-pressure packing for uniform bed density
• Fused-silica emitters with a 7 µm bore for stable nano-spray

Main Results and Discussion

Over four replicate EASY-Spray columns, peptide identifications ranged from ~87 000 to 94 000 and protein identifications around 7 000, demonstrating high reproducibility. In a ruggedness test of 500 injections on a double nanoViper column, retention time drift remained below 0.1 min, peak width and symmetry metrics showed negligible variation, and system pressure held stable near 350 bar. These outcomes confirm that both column formats deliver consistent chromatographic performance over extended use.

Benefits and Practical Applications

The EASY-Spray and double nanoViper PepMap Neo columns offer:

• Exceptional column-to-column consistency for large-scale proteomics
• High pressure tolerance (up to 1500 bar) enabling longer columns or smaller particles for deeper coverage
• Zero-dead-volume connections for sharper peaks and improved sensitivity

These attributes support workflows in biomarker discovery, quantitative proteomics and multi-site studies requiring strict reproducibility.

Future Trends and Potential Uses

With continued advances in column packing technologies and UHPLC systems, future developments may include:

• Sub-2 µm and core-shell particle chemistries to further boost separation power
• Integrated microfluidic cartridges for automated sample preparation and desalting
• Expanded temperature control ranges to optimize viscosity and resolution

Conclusion

Thermo Scientific EASY-Spray and double nanoViper PepMap Neo columns, when used with the Vanquish Neo UHPLC and an Orbitrap Exploris 480, deliver robust, high-pressure, low-dead-volume performance. They provide reproducible peptide and protein identifications and are well suited for demanding proteomics applications.

Reference

Thermo Scientific Technical Note TN74152 and Product Specification PS000155-EN 0821C