Normalization of organ-on-a-Chip samples for mass spectrometry based proteomics and metabolomics via Dansylation-based assay.

Autor: Gallagher EM; U.S. Army, Threat Agent Sciences Division, Combat Capabilities Development Command (DEVCOM) Chemical Biological Center (CBC), 5183 Blackhawk Rd., Aberdeen Proving Ground, Gunpowder, MD 21010, USA; National Academies of Sciences, Engineering, and Medicine, NRC Research Associateship Program, 500 Fifth Street, NW, Washington, DC, 20001, USA. Electronic address: erin.m.gallagher7.civ@army.mil., Rizzo GM; U.S. Army, BioSciences Division, Combat Capabilities Development Command (DEVCOM) Chemical Biological Center (CBC), 5183 Blackhawk Rd., Aberdeen Proving Ground, Gunpowder, MD 21010, USA., Dorsey R; U.S. Army, Threat Agent Sciences Division, Combat Capabilities Development Command (DEVCOM) Chemical Biological Center (CBC), 5183 Blackhawk Rd., Aberdeen Proving Ground, Gunpowder, MD 21010, USA., Dhummakupt ES; U.S. Army, BioSciences Division, Combat Capabilities Development Command (DEVCOM) Chemical Biological Center (CBC), 5183 Blackhawk Rd., Aberdeen Proving Ground, Gunpowder, MD 21010, USA., Moran TS; U.S. Army, Threat Agent Sciences Division, Combat Capabilities Development Command (DEVCOM) Chemical Biological Center (CBC), 5183 Blackhawk Rd., Aberdeen Proving Ground, Gunpowder, MD 21010, USA., Mach PM; U.S. Army, BioSciences Division, Combat Capabilities Development Command (DEVCOM) Chemical Biological Center (CBC), 5183 Blackhawk Rd., Aberdeen Proving Ground, Gunpowder, MD 21010, USA., Jenkins CC; U.S. Army, BioSciences Division, Combat Capabilities Development Command (DEVCOM) Chemical Biological Center (CBC), 5183 Blackhawk Rd., Aberdeen Proving Ground, Gunpowder, MD 21010, USA.
Jazyk: angličtina
Zdroj: Toxicology in vitro : an international journal published in association with BIBRA [Toxicol In Vitro] 2023 Apr; Vol. 88, pp. 105540. Date of Electronic Publication: 2022 Dec 20.
DOI: 10.1016/j.tiv.2022.105540
Abstrakt: Mass spectrometry based 'omics pairs well with organ-on-a-chip-based investigations, which often have limited cellular material for sampling. However, a common issue with these chip-based platforms is well-to-well or chip-to-chip variability in the proteome and metabolome due to factors such as plate edge effects, cellular asynchronization, effluent flow, and limited cell count. This causes high variability in the quantitative multi-omics analysis of samples, potentially masking true biological changes within the system. Solutions to this have been approached via data processing tools and post-acquisition normalization strategies such as constant median, constant sum, and overall signal normalization. Unfortunately, these methods do not adequately correct for the large variations, resulting in a need for increased biological replicates. The methods in this work utilize a dansylation based assay with a subset of labeled metabolites that allow for pre-acquisition normalization to better correlate the biological perturbations that truly occur in chip-based platforms. BCA protein assays were performed in tandem with a proteomics pipeline to achieve pre-acquisition normalization. The CN Bio PhysioMimix was seeded with primary hepatocytes and challenged with VX after six days of culture, and the metabolome and proteome were analyzed using the described normalization methods. A decreased coefficient of variation percentage is achieved, significant changes are observed through the proteome and metabolome, and better classification of biological replicates acquired because of these strategies.
Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Phillip M. Mach reports financial support was provided by Defense Advanced Research Projects Agency. Phillip M. Mach reports financial support was provided by Defense Threat Reduction Agency.
(Published by Elsevier Ltd.)
Databáze: MEDLINE
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