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bioRxiv · 10.1101/2021.01.19.426900

In Vivo Subcellular Mass Spectrometry Enables Proteo-Metabolomic Single-cell Systems Biology in a Chordate Embryo Developing to a Normally Behaving Tadpole (X. laevis)

Lombard-Banek, C.·Li, J.·Portero, E. P.·Onjiko, R. M.·Singer, C. D.·Plotnick, D.·Al Shabeeb, R. Q.·Nemes, P.
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Abstract

We present the first example of in vivo high-resolution mass spectrometry (HRMS) for subcellular molecular systems biology of proteins and metabolites. With light microscopy, we identified the left-dorsal and left-ventral animal cells in cleavage-stage non-sentient Xenopus laevis embryos. Using precision-translated fabricated microcapillaries, the subcellular content of each cell was double-probed, each time collecting <5% of cell volume (~10 nL) swiftly (<5 s/event). The proteins and metabolites were analyzed by custom-built ultrasensitive capillary electrophoresis electrospray ionization employing Orbitrap and time-of-flight HRMS. Label-free detection of ~150 metabolites (57 identified) and 738 proteins found proteo-metabolomic networks with differential quantitative activities between the cell types. Spatially and temporally scalable sampling the technology preserved the integrity of the analyzed cells, the neighboring cells, and the embryo. 95% of the analyzed embryos developed into sentient tadpoles that were indistinguishable from their wild-type siblings based on anatomy and visual function in a background color preference assay.

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Lombard-Banek, C., Li, J., Portero, E. P., Onjiko, R. M., Singer, C. D., Plotnick, D., Al Shabeeb, R. Q., Nemes, P.. 2021-01-20. In Vivo Subcellular Mass Spectrometry Enables Proteo-Metabolomic Single-cell Systems Biology in a Chordate Embryo Developing to a Normally Behaving Tadpole (X. laevis). https://doi.org/10.1101/2021.01.19.426900

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