A microfluidics-enabled automated workflow of sample preparation for MS-based immunopeptidomics.
Cell Rep Methods
; 3(6): 100479, 2023 06 26.
Article
em En
| MEDLINE
| ID: mdl-37426762
ABSTRACT
Mass spectrometry (MS)-based immunopeptidomics is an attractive antigen discovery method with growing clinical implications. However, the current experimental approach to extract HLA-restricted peptides requires a bulky sample source, which remains a challenge for obtaining clinical specimens. We present an innovative workflow that requires a low sample volume, which streamlines the immunoaffinity purification (IP) and C18 peptide cleanup on a single microfluidics platform with automated liquid handling and minimal sample transfers, resulting in higher assay sensitivity. We also demonstrate how the state-of-the-art data-independent acquisition (DIA) method further enhances the depth of tandem MS spectra-based peptide sequencing. Consequently, over 4,000 and 5,000 HLA-I-restricted peptides were identified from as few as 0.2 million RA957 cells and a melanoma tissue of merely 5 mg, respectively. We also identified multiple immunogenic tumor-associated antigens and hundreds of peptides derived from non-canonical protein sources. This workflow represents a powerful tool for identifying the immunopeptidome of sparse samples.
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Base de dados:
MEDLINE
Assunto principal:
Proteômica
/
Microfluídica
Idioma:
En
Revista:
Cell Rep Methods
Ano de publicação:
2023
Tipo de documento:
Article
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