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Creative Biolabs offers single-cell proteomics mass spectrometry profiling service employing heavy metal isotopes as antibody labels to detect cellular proteins quantitatively using mass spectrometry.

Single Cell Proteomics Mass Spectrometry

Although MS-based proteome profiling has gotten more comprehensive and quantitative, the comparatively large samples required for an in-depth measurement has remained a chronic drawback. Bulk samples, which often contain thousands or more cells, provide a population average while masking significant cellular variability. Single-cell proteomics has the potential to revolutionize biomedical research and provide a new degree of granularity in biological system comprehension. More than 1000 proteins can currently be quantified from individual mammalian cells because to recent developments in sample processing, separations, and MS instruments.

Elemental analysis by a mass spectrometer.Fig.1. Elemental analysis by a mass spectrometer. (Zhang, 2018)

Methods of Single Cell Mass Spectrometry

For single-cell MS, there are two essentially different techniques. In vacuum-based ion sources, the cells are frequently dehydrated and interrogated by an ion or laser beam for examination. The studied cells are far from their native state due to the loss of water, yet these ion sources provide excellent sensitivity and often high throughput for the analysis. Ambient ionization technologies, on the other hand, cause minimal cell disturbance and can be used in situ or even in vivo. But this method has reduced sensitivity and limited throughput.

  • Vacuum-based ion sources method
  • Vacuum-based single-cell MS techniques include SIMS, MALDI, and matrix-free LDI methods. Each of these methods can study tissue-embedded cells, and they can be paired with cell trapping and/or microscope-guided cell targeting methods for isolated, floating, or circulating cells.

  • Ambient ionization method
  • The analysis of live cells in their natural environment became possible thanks to the ambient ionization methods for single-cell MS. This has major implications for single-cell analysis because of the much reduced mechanical and chemical disturbances and greatly simpler sample preparation, this has major implications for single-cell analysis.

Single Cell Proteomics Mass Spectrometry Workflow

The single cell proteomics carrier technique employs tandem mass tags to label lowly abundant samples of interest and a carrier sample, before combining all labeled samples for analysis by liquid chromatography tandem MS. Tandem mass tag (TMT) ensures that copies of a given peptide sequence from single cells and bulk samples have the same mass-to-charge ratio during survey scans, allowing them to be separated together for fragmentation and MS2 analysis. The precursor ions produce sample-specific reporter ions during fragmentation, whose abundances allow for relative measurement.

Workflow of single-cell proteomics mass spectrometry.Fig.2. Workflow of single-cell proteomics mass spectrometry. (Slavov, 2021)

Feature & Benefits

The single cell proteomics mass spectrometry profiling service has a high signal resolution and can simultaneously detect hundreds of distinct labels. Mass cytometry can collect data on single-cell surface marker, signaling, transcription factors, cytokines, cell cycle proliferation, and other information at the same time. It can be used in a variety of scientific domains, including hematopoiesis, immunology, stem cells, cancer, and drug screening.

Published Data

Paper Title Streamlined single-cell proteomics by an integrated microfluidic chip and data-independent acquisition mass spectrometry
Journal Nature communications
IF 14.919
Published 2022
Abstract In this study, they profile on average 1,500 protein groups across 20 single mammalian cells using chip-based sample processing and DIA-MS with project-specific mass spectrum libraries. They cover a dynamic range of 5 orders of magnitude with strong consistency and <16% missing values across runs using the chip-DIA methodology to profile the proteomes of adherent and non-adherent malignant cells.
Result

Using built-in cell traps, exact numbers of cells, including 1, 5, 10, 50, and 100 cells, were caught to operate the iProChip for proteomic workflow. Dispensing and incubating cells trapped in capture chambers with cocktail buffer containing RapiGest, tris(2-carboxyethyl)phosphine hydrochloride (TCEP), and chloroacetamide (CAA), which was specifically adapted to achieve one-pot cell lysis, protein reduction, and alkylation to minimize sample loss, was done in parallel. Following that, the reaction vessel was used to perform protein digestion and acidification, and the digested peptides were then passed over the C18 column for 15 minutes to undergo multiplexed desalting

 Comparison of identification coverage and quantitation performance of proteomic profiling of PC-9 cells by DIA and DDA methods.Fig.3. Comparison of identification coverage and quantitation performance of proteomic profiling of PC-9 cells by DIA and DDA methods. (Gebreyesus, 2022)

A. Comparison of data-dependent acquisition (DDA, light blue) and data-independent acquisition (DIA, light green) modes.
B. Overlap of protein groups identified by DDA and DIA.
C. Distribution of coefficient of variation (CV%) for quantified protein groups by DDA and DIA.
D. Evaluation of missing values (%) of proteins identified and quantified in triplicate LC–MS/MS runs by DDA and DIA.
E. Assessment of dynamic range based on protein abundance rank and annotation of selected proteins related to lung cancer.

Creative Biolabs has methodically developed every stage of the process, from antibody creation through data analysis, to assure high-quality study outcomes. Please contact us for more details about our single-cell proteomics mass spectrometry profiling service.

References

  1. Zhang, L., Vertes, A. Single-cell mass spectrometry approaches to explore cellular heterogeneity. Angewandte Chemie International Edition. 2018; 57(17): 4466-4477.
  2. Slavov, N. Single-cell protein analysis by mass spectrometry. Current Opinion in Chemical Biology. 2021; 60: 1-9.
  3. Gebreyesus, S.T., et al. Streamlined single-cell proteomics by an integrated microfluidic chip and data-independent acquisition mass spectrometry. Nature communications. 2022; 13(1): 1-13.
! ! For Research Use Only. Not for diagnostic or therapeutic purposes.

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