NanoLC

Nano liquid chromatography (nanoLC) is a variant of high-performance liquid chromatography that operates at nanoliter per minute flow rates. By reducing flow and column volume, nanoLC increases the concentration of analytes entering the detector, improving sensitivity, especially when coupled to mass spectrometry.

Typical nanoLC systems use narrow-bore columns with inner diameters around 75 μm and lengths from 15 to 60 cm. They employ high-efficiency stationary phases such as C18 to separate complex mixtures. The system requires precise pumping and low dead volumes, and is often coupled to nanoelectrospray ionization (nanoESI) for MS detection.

Solvent systems usually involve a binary gradient with solvents such as water with 0.1% formic acid and an organic solvent like acetonitrile or methanol, enabling gradient elution of peptides and other analytes.

NanoLC provides high separation efficiency and strong concentration of analytes at the detector, yielding enhanced mass spectrometric signal and lower limits of detection. The technique is particularly sensitive to leaks, blockages, and solvent composition, requiring careful maintenance and specialized hardware.

Applications include proteomics for peptide mapping and shotgun sequencing, as well as metabolomics, glycomics, and other low-volume analyses.

Limitations include higher equipment cost, complexity of method development, and potential sample carryover or column fouling.

NanoLC-MS has become a standard tool in many proteomics workflows, particularly when maximum sensitivity is required to analyze tiny sample amounts.