Abstract
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.
Highlights
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)
Absolute quantification has been challenging. This is an important reason that ToF-SIMS has not been a popular analysis tool, though it was developed more than 50 years ago
Because all protein films form in water, it is reasonable to expect that the effect of the matrix can be minimized when comparing in situ liquid ToF-SIMS data from different protein films in data analysis
Summary
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). 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. (c) ToF-SIMS profiles of an amino acid fragment (CH5N3+, m/z 59, red curve) and a water cluster ((H2O)3H+, m/z 55, green curve) from the adsorbed and hydrated BSA film during two measuring periods: using the Bi3+ primary ion beam with (I) long pulse width and (II) short pulse width. The SIMS m/z spectra and 3D images have provided a detailed evaluation on the composition and structure of these hydrated adsorbed protein films for the first time
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