by Asim Siddiqui, SVP, Research & Tech Development, Seer
Today, scientists are looking to mass spectrometry-based proteomics to help them uncover novel insights into health and disease with new approaches enabling depth of protein coverage not previously possible. Advanced technologies are becoming more automated, cost-effective, and widely available to the broader life sciences community. With this progress, mass spec-based proteomics has evolved, allowing researchers to perform population-scale studies with deep molecular profiling, better throughput and flexible workflows.
Mass spec is considered the gold standard in proteomics, so it is essential to understand how it performs compared with other approaches, the evidence demonstrating its utility in definitively identifying peptides, challenges inherent to the methods, and what the future holds.
Basics of mass spec-based proteomics
Mass spectrometry allows researchers to accurately and reproducibly measure the mass of ions. A fundamental innovation that enabled proteomics in mass spec was the invention of electrospray ionization, enabling efficient ionization of peptides and the subsequent measurement of their mass as they fly through the mass spec.
In general, a majority of mass spec-based proteomics are performed using a method referred to as “bottom-up proteomics”, where the proteins are digested by an enzyme, typically trypsin, that leads to cleavage at particular sites along the peptide backbone. This approach leads to defined shorter peptides that are more generally compatible with currently available technologies. In the mass spec, each peptide is fragmented further. Analyzing a sub-fragment of the parent peptide chain allows scientists to gather more information about sub-components of that chain, reconstructing peptide sequences more accurately. Top-down proteomics analyzes intact proteins, which enables the identification of the complete protein sequences. However, its experimental challenges currently make it unsuitable for deep coverage, large-scale population studies.
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