Mass Cytometry

Mass cytometry is a mass spectrometry technique based on inductively coupled plasma mass spectrometry combined with mass tags that allows accurate measuring of various molecular species on single cells in a highly multiplexed fashion. In this approach, antibodies are tagged with isotopically pure rare earth elements and these are used to tag the components of cells.

Cells are nebulized and sent into argon plasma, breaking molecular bonds and ionizing metal tags, which are then analyzed by a time-of-flight mass spectrometer. The approach overcomes limitations of spectral overlap that limit flow cytometry, allowing simultaneous quantification of 50 mass tag channels.

Tagging technology and instrument development occurred at the University of Toronto and DVS Sciences, Inc, which has been acquired by Fludigm in 2014. Our lab was one of the pioneer users of mass cytometry, for the first time applying it to profile the hematopoietic continuum and assess signaling responses to drug treatments on a single cell level. We are continuously developing methods and applications and using the technology to profile a variety of human healthy and disease conditions.

Multiplexed Ion Beam Imaging (MIBI)

We have developed a method that uses secondary ion mass spectrometry to image antibodies that are tagged with isotopically pure elemental metal reporters.

Multiplexed ion beam imaging (MIBI) allows analyzing up to 100 targets simultaneously over a five-log dynamic range in a way similar to CyTOF, but in addition to measuring protein levels on individual cells, it also provides the information about cell morphology and localization.

We used MIBI to analyze paraffin-embedded human breast cancer sections stained with 10 labels simultaneously, providing new insights into disease pathogenesis that could be valuable for basic research and clinical diagnostics.

 

Phospho flow

Phospho-specific flow cytometry, or phospho flow, measures the phosphorylation state of intracellular proteins at the single cell level. This is achieved by a special cell permebealization strategy and staining intracellular targets with phosphoepitope-specific antibodies.

Many phosphorylation events can be analyzed simultaneously in each cell, along with cell surface markers, enabling complex biochemical signaling networks to be resolved in heterogeneous cell populations.

The method has been applied to many diverse areas of biology, including the characterization of signaling pathways in normal immune responses to antigenic stimulation and microbial challenge, alteration of signaling networks that occur in cancer and autoimmune diseases, and high- throughput, high-content drug discovery. We also showed the information about the distribution of phosphoepitope levels in single cells can be also used to infer the causal structure of intracellular signalling networks de novo using machine learning methods.