In situ molecular imaging of adsorbed protein films in water indicating hydrophobicity and hydrophilicity

In situ molecular imaging of protein films adsorbed on a solid surface in water was realized by using a vacuum compatible microfluidic interface and time-of-flight secondary ion mass spectrometry (ToF-SIMS). Amino acid fragments from such hydrated protein films are observed and identified in the positive ion mode and the results are in agreement with reported works on dry protein films. Moreover, water clusters from the hydrated protein films have been observed and identified in both the positive and negative ion mode for a series protein films. Thus, the detailed composition of amino acids and water molecules in the hydrated protein films can be characterized, and the protein water microstructures can be revealed by the distinct three-dimensional spatial distribution reconstructed from in situ liquid ToF-SIMS molecular imaging. Furthermore, spectral principal component analysis of amino acid fragment peaks and water cluster peaks provides unique insights into the water cluster distribution, hydrophilicity, and hydrophobicity of hydrated adsorbed protein films in water.