RapidAIM 2.0: a high-throughput assay to study functional response of human gut microbiome to xenobiotics

Our gut microbiome functions like an organ, having its own set of functions and roles which can be modulated by various types of xenobiotic and biotic components. High-throughput screening approaches that are established based on in vitro or ex vivo cell, tissue or organ models greatly accelerate drug discovery and our understanding of biological and pathological processes within these systems. There was a lack of a high-throughput compatible functional screening approach of the gut microbiome until we recently developed the RapidAIM (Rapid Assay of Individual Microbiome). RapidAIM combines an optimized culturing model, which maintains the taxonomic and functional profiles of the human gut microbiome in vitro, and a high-throughput metaproteomics workflow to gain deep functional insights into microbiome responses. This protocol describes the most recently optimized 2.0 version of RapidAIM, consisting of extensive details on stool sample collection, biobanking, in vitro culturing and stimulation, microbiome sample processing, and metaproteomics measurement and data analysis. To demonstrate the typical outcome of the protocol, we show an example of using RapidAIM 2.0 to evaluate the effect of prebiotic kestose on ex vivo individual human gut microbiomes biobanked with five different workflows; we also show that kestose had consistent functional effects across individuals and can be used as positive control in the assay.