Fig. 2.

Alternative imaging for subcellular proteomics and/or transcriptomics, which couple technologies in MS, microfluidics, and/or microdissection.A, instrumentation coupling flow cytometry and microscopy allows for multiplexing of several protein–RNA targets using fluorescent labels, gaining both spatial and single-cell information. B, microlaser ablation and ionization of molecules, such as peptides, lipids, or metabolites, directly from tissue or cell culture sample enables label-free acquisition of mass spectra across each “pixel” of sample. Very rich datasets but still have poor resolution because of current technical limitations. C, similar to MSI, microlaser ablation allows for acquisition of spectra per “pixel” of a sample. Though, this method has improved subcellular resolution and uses labeling of antibodies conjugated to non-naturally occurring metal isotopes to quantify ∼40 target proteins/RNAs of interest. The metal isotope signals have less signal overlap than fluorescent methods allowing improved multiplexing than traditional antibody probing. MSI, MS imaging.