A1 Refereed original research article in a scientific journal
Targeted and Untargeted Amine Metabolite Quantitation in Single Cells with Isobaric Multiplexing
Authors: Heininen, Juho; Movahedi, Parisa; Kotiaho, Tapio; Kostiainen, Risto; Pahikkala, Tapio; Teppo, Jaakko
Publisher: Wiley-VCH
Publication year: 2024
Journal: Chemistry - A European Journal
Journal acronym: Chemistry
Article number: e202403278
Volume: 30
Issue: 72
ISSN: 0947-6539
eISSN: 1521-3765
DOI: https://doi.org/10.1002/chem.202403278
Web address : https://doi.org/10.1002/chem.202403278
Self-archived copy’s web address: https://research.utu.fi/converis/portal/detail/Publication/459037549
We developed a single cell amine analysis approach utilizing isobarically multiplexed samples of 6 individual cells along with analyte abundant carrier. This methodology was applied for absolute quantitation of amino acids and untargeted relative quantitation of amines in a total of 108 individual cells using nanoflow LC with high-resolution mass spectrometry. Together with individually determined cell sizes, this provides quantification of intracellular concentrations within individual cells. The targeted method was partially validated for 10 amino acids with limits of detection in low attomoles, linear calibration range covering analyte amounts typically from 30 amol to 120 fmol, and correlation coefficients (R) above 0.99. This was applied with cell sizes recorded during dispensing to determine millimolar intracellular amino acid concentrations. The untargeted approach yielded 249 features that were detected in at least 25% of the single cells, providing modest cell type separation on principal component analysis. Using Greedy forward selection with regularized least squares, a sub-selection of 100 features explaining most of the difference was determined. These features were annotated using MS2 from analyte standards and accurate mass with library search. The approach provides accessible, sensitive, and high-throughput method with the potential to be expanded also to other forms of ultrasensitive analysis.
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Funding information in the publication:
Research Council of Finland (project #321472) is acknowledged for funding.