Comparison of the techniques for isolating immunoassay-suitable proteins from heterogeneous fecal samples.

Abstract

Immunoassays could provide valuable insights into disease biomarkers and gut health by measuring fecal proteins. However, reliably isolating intact proteins from feces is challenging due to its heterogeneous and variable composition. This paper aims to review and compare different methods for extracting proteins from fecal samples to make them suitable for immunoassay analysis. Mechanical homogenization helps release proteins by disrupting solids, while protease inhibitors preserve protein integrity. Detergents like SDS solubilize proteins by disrupting hydrophobic interactions. Organic solvents such as acetone precipitate proteins and remove contaminants. Thermal treatment denatures proteases. Immunocapture uses antibodies to purify target proteins away from interference selectively. Commercial kits contain optimized buffers but may be cost-prohibitive. Combining mechanical, chemical, and immunological techniques synergistically integrates their advantages, improving the recovery of native proteins with reduced matrix effects. While all methods have merits, tailored protocols integrating multiple mechanisms appear most promising for extracting immunoassay-suitable fecal proteins. Further optimization and standardization of such combination approaches matched to proteins and assays of interest helps expand noninvasive fecal proteome analysis.