Pushing the boundaries of immunopeptidomics research with trapped ion mobility mass spectrometry (TIMS)
Introduction
Researchers at the Broad Institute of MIT and Harvard’s Proteomics Platform, led by founder and senior director Steven Carr, Ph.D., are using trapped ion mobility spectrometry (TIMS) to drive the technical advancement of proteomics and gain further understanding of disease pathways, targets, and drug effects.
The Broad Institute
Claudia Ctortecka, Ph.D. is a Postdoctoral Fellow at the Proteomics Platform, Broad Institute of MIT and Harvard, which uses state-of-the-art TIMS time of flight (TOF) systems including the Bruker timsTOF Ultra, timsTOF single cell proteomics (SCP) and timsTOF high-throughput (HT) instruments.
The team has built collaborative partnerships across the institute’s expansive network to provide mass spectrometry (MS)-based proteomics analysis capabilities that enhance peptide and protein identifications with the goal of understanding disease biology and drug effects at the mechanistic level.
A core aspect of the group’s work includes the quantification of proteins and their modifications from bulk cells, tissues and biofluids down to the single cell level, which requires cutting-edge high throughput and highly sensitive mass spectrometric methods.
Identification and quantification of immunopeptides
Immunopeptidomics holds immense potential to reveal a comprehensive catalog of peptides recognized by T cells, providing valuable insights into how the immune system identifies these molecules. This research could pave the way towards immunotherapies targeting cancer, autoimmune diseases, and infectious diseases.
A primary challenge in immunopeptidomics research is to selectively extract and identify peptides presented by major histocompatibility complex (MHC) molecules from biological samples, such as tumor tissue or cells, without compromising the integrity and representation of peptides. Tumor tissue for instance is heterogeneous and consists of multiple cell types with varying MHC expression profiles, complicating the analysis.
High-throughput instruments
Dr. Ctortecka is implementing a process of technology and method development to achieve the best immunopeptidomics depth possible at high throughput:
“We have demonstrated that we can increase throughput by overcoming the need to fractionate the samples. I think one of the reasons for this, besides the increased scanning speed, is that we can efficiently leverage singly charged precursors. We are still investigating how well we can do this and how much information we can get out of them, but it is the selection of peptide-like singly charged species using their collisional cross section (CCS) that is helping us unlock this new potential.”
Mass spectrometry offers unique advantages for proteomics, enabling researchers to separate ions according to their CCS values and enhancing sensitivity and peak capacity. This allows for accurate monitoring of low-abundance molecules such as human leukocyte antigens (HLA)-bound peptides to distinguish peptides that are similar and exploiting mobility offset mass aligned (MOMA) capabilities. Furthermore, parallel accumulation serial fragmentation (PASEF) is a TOF MS-based acquisition method enabled by TIMS and employed for fast peptide separation and more confident identification. Together, the specificity, sensitivity, and high speed of MOMA and PASEF results in confident identification of HLA-I and HLA-II bound peptides though TIMS in a single run.
Next steps for the Proteomics Platform
Dr. Ctortecka’s hope for the future is to see MS integrated into the clinic, allowing clinicians to take a tissue biopsy and process it directly with a standardized workflow.
For Research Use Only. Not for use in clinical diagnostic procedures.