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Li et al. Microbiome Res Rep 2024;3:26 Microbiome Research
DOI: 10.20517/mrr.2023.57
Reports

Original Article Open Access

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

xenobiotics

#
Leyuan Li # , Janice Mayne , Adrian Beltran, Xu Zhang , Zhibin Ning , Daniel Figeys
School of Pharmaceutical Sciences, Ottawa Institute of Systems Biology and Department of Biochemistry, Microbiology and
Immunology, Faculty of Medicine, University of Ottawa, Ottawa K1H8M5, Ontario, Canada.
#
Authors contributed equally.
Correspondence to: Prof. Daniel Figeys, School of Pharmaceutical Sciences, Ottawa Institute of Systems Biology and Department
of Biochemistry, Microbiology and Immunology, Faculty of Medicine, University of Ottawa, 451 Smyth Road, Ottawa K1H 8M5,
Ontario, Canada. E-mail: dfigeys@uottawa.ca

How to cite this article: Li L, Mayne J, Beltran A, Zhang X, Ning Z, Figeys D. RapidAIM 2.0: a high-throughput assay to study
functional response of human gut microbiome to xenobiotics. Microbiome Res Rep 2024;3:26. https://dx.doi.org/10.20517/mrr.
2023.57

Received: 2 Oct 2023 First Decision: 21 Nov 2023 Revised: 3 Mar 2024 Accepted: 25 Mar 2024 Published: 3 Apr 2024

Academic Editor: Marco Ventura Copy Editor: Dong-Li Li Production Editor: Dong-Li Li

Abstract
Aim: Our gut microbiome has its own functionalities which can be modulated by various xenobiotic and biotic
components. The development and application of a high-throughput functional screening approach of individual
gut microbiomes accelerates drug discovery and our understanding of microbiome-drug interactions. We
previously developed the rapid assay of individual microbiome (RapidAIM), which combined an optimized
culturing model with metaproteomics to study gut microbiome responses to xenobiotics. In this study, we aim to
incorporate automation and multiplexing techniques into RapidAIM to develop a high-throughput protocol.

Methods: To develop a 2.0 version of RapidAIM, we automated the protein analysis protocol, and introduced a
tandem mass tag (TMT) multiplexing technique. To demonstrate the typical outcome of the protocol, we used
RapidAIM 2.0 to evaluate the effect of prebiotic kestose on ex vivo individual human gut microbiomes biobanked
with five different workflows.

Results: We describe the protocol of RapidAIM 2.0 with extensive details on stool sample collection, biobanking,
in vitro culturing and stimulation, sample processing, metaproteomics measurement, and data analysis. The
analysis depth of 5,014 ± 142 protein groups per multiplexed sample was achieved. A test on five biobanking

© The Author(s) 2024. Open Access This article is licensed under a Creative Commons Attribution 4.0
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adaptation, distribution and reproduction in any medium or format, for any purpose, even commercially, as
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