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HLA-Ig Based Artificial Antigen Presenting Cells for Efficient ex vivo Expansion of Human CTL
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Extending Serum IgG1 Antibody Repertoire Coverage Using DIA-PTCR.

Rien W Leuvenink1, Linus Wollenweber1, Albert Bondt1

  • 1Biomolecular Mass Spectrometry and Proteomics, Bijvoet Centre for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Padualaan 8 3584 CH, Utrecht, The Netherlands.

Journal of the American Society for Mass Spectrometry
|May 11, 2026
PubMed
Summary
This summary is machine-generated.

Gas-phase fractionation using data-independent analysis with proton transfer charge reduction (DIA-PTCR) significantly enhances antibody repertoire profiling. This method doubles the number of detectable antibody clones in serum by reducing spectral congestion in LC-MS analysis.

Keywords:
FAB fragmentsLC-MS based clonal profilingantibody repertoireproton transfer charge reduction (PTCR)spectral congestion

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Area of Science:

  • Biochemistry
  • Immunology
  • Analytical Chemistry

Background:

  • Liquid chromatography-mass spectrometry (LC-MS) based antibody Fab profiling is crucial for understanding immune responses.
  • Extreme spectral congestion in LC-MS hinders the detection and quantification of diverse antibody clones in serum.
  • Current methods face challenges in comprehensively analyzing antibody repertoires due to molecular similarities and ionization complexities.

Purpose of the Study:

  • To develop a method for reducing spectral congestion in LC-MS-based Fab profiling.
  • To enhance the number of detectable antibody clones within a single LC-MS run.
  • To maintain quantitative accuracy for antibody abundance measurements in repertoire profiling.

Main Methods:

  • Implementation of gas-phase fractionation using proton transfer charge reduction (PTCR) in a data-independent analysis (DIA) acquisition mode (DIA-PTCR).
  • Utilizing a limited number of isolation windows (five) within a specific m/z region to cover the total Fab distribution.
  • Integration of reduced noise filtering alongside gas-phase fractionation.

Main Results:

  • The DIA-PTCR method significantly reduced spectral congestion, a major challenge in LC-MS Fab profiling.
  • The number of detected antibody clones per run increased by over 100%, from 1043 to 2063.
  • Quantitative data on antibody abundances were successfully retained, enabling accurate repertoire profiling.

Conclusions:

  • DIA-PTCR is a highly beneficial technique for advancing LC-MS-based antibody repertoire profiling.
  • This approach overcomes limitations of spectral congestion, enabling deeper insights into antibody diversity.
  • The method offers a more comprehensive and quantitative analysis of antibody repertoires.