Single-cell omics with Live-seq

This application note introduces the novel scRNA-seq approach  -  Live-seq – created and developed by Chen et al. (2022) and presents its potential implications for single-cell omics.

Go straight to:

Single-cell omics: definition and existing methods

Single-cell omics definition and existing methods scRNA seq

Schematic Illustration of RNA sequencing by Cytosurge

Omics is known as the study of biological molecules that end with the suffix -omics: Genomics is the study of the genes. Transcriptomics refers to the analysis of RNA transcripts. Proteomics describes the investigation of proteins. Metabolomics focuses on metabolites. 

Single-cell omics addresses cellular heterogeneity head on by isolating and analyzing molecular components from individual cells.

With the development of transcriptomics, several methods for transcriptome profiling were developed, including the RNA sequencing (scRNA-seq) approach.

While many of the existing RNA-seq technologies rely on cell lysis — the Live-seq approach created by Chen et al. (2022), supported by the FluidFM technology is the only non-destructive single-cell RNA sequencing method that allows researchers to take transcriptome snapshots of a cell without killing it. [1]

Do you want more information about Live-seq and FluidFM?

Live-seq: the novel scRNA-seq approach that keeps your cells alive. 

This approach pairs the FluidFM OMNIUM system with a highly sensitive low-input RNA-seq protocol, which stems from a collaboration between ETH Zurich and EPFL. The FluidFM OMNIUM allows you to extract sub-picoliter volumes from cellular compartments of a living single cell and then isolate the extract for further analysis. By avoiding destructive methods, such as cell lysis, you can perform further downstream molecular and phenotypic analysis and even transcriptome profiling over time on the very same cell. 

This approach brings a new paradigm to single-cell transcriptomics by enabling researchers to take transcriptome snapshots of a cell without killing it. Chen et al. (2022) demonstrated that gene expression profiles obtained from single-cell biopsies are suitable representations of lysed cell transcriptomes. [1] This introduces trajectory instead of end-point analysis to your transcriptomics, metabolomics, proteomics, or any other omics studies. 

Non-destructive single-cell biopsy 

Get reliable results without altering gene expression, cell phenotype, or cell-cell interactions.

Continuous monitoring of a single-cell with biopsies 

FluidFM extraction preserves cell viability: extract several times from the same cell within the same run or periodically over time.

Analyze temporal gene expression

Sequential profiling of the single-cell’s transcriptome

FluidFM for Live-seq and Single-cell Omics

Force-sensitive probes with closed microscopic channels are used in our patented FluidFM (Fluidic Force Microscopy) technology. These probes, which have apertures as small as 300 nm, enable the simultaneous delivery of femtoliter volumes and the measurement of interaction forces as low as pN. This technology can realize a broad range of single-cell manipulation methods suited for various applications, including the single-cell biopsy - a gentle single-cell extraction method.

The single-cell extraction of sub-cellular content is supported by a semi-automated application-specific workflows and probe handling. An intuitive software allows for an easy cell selection – point-and-click to target a specific cell.

This new approach presents a number of important outcomes for single-cell omics: 

  • Record a cell transcriptome prior to phenotyping.

  • Record transcriptional events over time to reveal how molecular components influence cell behavior.

  • Directly link an individual cell’s history and trajectory to unravel past cell states and understand lineage decisions. 

  • Take a biopsy from cells of a heterogeneous disease before and after exposure to a specific therapeutic to identify molecular signatures for early drug development.


FluidFM in action: non-destructive extraction of cellular content

Advance your omics research with FluidFM

Explore FluidFM publications

Genome-wide molecular recording using Live-seq

Chen et al. show the establishment of Live-seq, an approach for single-cell transcriptome profiling that preserves cell viability during RNA extraction using FluidFM. By using a model involving exposure of macrophages with lipopolysaccharide (LPS), they were able to apply a genome-wide ranking of genes based on their ability to impact macrophage LPS response heterogeneity.  Furthermore, they show that Live-seq can be used to sequentially profile the transcriptomes of individual macrophages before and after stimulation with LPS. This enables the direct mapping of a cell’s trajectory and transforms scRNA-seq from an end-point to a temporal analysis approach.

[1] W. Chen, O. Guillaume-Gentil, P. Yde Rainer, C. G. Gäbelein, W. Saelens, V. Gardeaux, A. Klaeger, R. Dainese, M. Zachara, T. Zambelli, J. A. Vorholt & B. Deplancke. Live-seq enables temporal transcriptomic recording of single cells. (Aug 2022) Nature, doi:10.1038/s41586-022-05046-9

Single-Cell Mass Spectrometry

In this publication Guillaume-Gentil et al. show non-destructive and quantitative withdrawal of intracellular fluid with sub-picoliter resolution using FluidFM, followed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. By this method they detected and identified several metabolites from the cytoplasm of individual HeLa cells. Validated by 13C-Glucose feeding experiments, this showed that metabolite sampling combined with mass spectrometry analysis was possible while preserving the physiological context and the viability of the analyzed cell. Thus, enabling complementary analysis of the cell. 

[2] O. Guillaume-Gentil, T. Rey, P. Kiefer, A.J. Ibáñez, R. Steinhoff, R. Brönnimann, L. Dorwling-Carter, T. Zambelli, R. Zenobi & J.A. Vorholt. Single-Cell Mass Spectrometry of Metabolites Extracted from Live Cells by Fluidic Force Microscopy. (May 2017) Anal Chem., 89(9), 5017-5023. doi:10.1021/acs.analchem.7b00367

Tunable Single-Cell Extraction for Molecular Analyses

Guillaume-Gentil et al. demonstrate the use of FluidFM for quantitative sampling of cytoplasmic and nucleoplasmic fractions from single cells at a sub-picoliter resolution followed by a comprehensive analysis of the soluble molecules withdrawn from the cytoplasm or the nucleus and dispensed adaptable to a broad range of analytical methods, including the detection of enzyme activities and transcript abundances.

[3] O. Guillaume-Gentil, R.V. Grindberg, R. Kooger, L. Dorwling-Carter, V. Martinez, D. Ossola, M. Pilhofer, T. Zambelli & J.A. Vorholt. Tunable Single-Cell Extraction for Molecular Analyses. (Jul 2016) Cell, 166(2), 506-516. doi: 10.1016/j.cell.2016.06.025.

80+ publications with FluidFM