The rapid detection of procalcitonin in septic serum using immunoaffinity MALDI chips

Clinical Proteomics

Research

Jaroslav Hrabak¹  Lucie Kraftova¹  Vendula Studentova¹  Martin Matejovic²  Thomas Karvunidis²  Jan Horak²  Jaroslav Radej²  Zuzana Kalaninova³  Petr Novak³  Josef Dvorak³  Petr Pompach⁴  Marcela Kralovcova⁵  Jana Novakova⁶  Michael Volny⁶ 

[1] Biomedical Center, Faculty of Medicine in Pilsen, Charles University, Pilsen, Czech Republic;Biomedical Center, Faculty of Medicine in Pilsen, Charles University, Pilsen, Czech Republic;Department of Internal Medicine, Faculty of Medicine in Pilsen, Pilsen University Hospital, Charles University, Pilsen, Czech Republic;Department of Biochemistry, Faculty of Science, Charles University, Prague, Czech Republic;Institute of Microbiology, The Czech Academy of Science, Prague, Czech Republic;Department of Biochemistry, Faculty of Science, Charles University, Prague, Czech Republic;Institute of Microbiology, The Czech Academy of Science, Prague, Czech Republic;Institute of Biotechnology, The Czech Academy of Science, Prague, Czech Republic;Department of Internal Medicine, Faculty of Medicine in Pilsen, Pilsen University Hospital, Charles University, Pilsen, Czech Republic;Institute of Microbiology, The Czech Academy of Science, Prague, Czech Republic;

关键词: Procalcitonin;    MALDI-TOF;    Sepsis;    Immunoaffinity;    Ion soft landing;   

DOI  :  10.1186/s12014-023-09410-3

 received in 2022-08-18, accepted in 2023-04-27,  发布年份 2023

来源: Springer

【 摘 要 】

BackgroundSepsis is a common worldwide health condition with high mortality. It is caused by a dysregulated immune response to the pathogen. Severe infections resulting in sepsis can be also determined by monitoring several bloodstream biomarkers, one of them being pro-hormone procalcitonin (PCT). PCT concentration in the bloodstream correlates well with sepsis and in severe cases increases up to a thousand times from the healthy physiological values in a short time. In this study, we developed a rapid technique for PCT detection by MALDI-TOF mass spectrometry, that uses in-situ enrichment directly on the specialized immuno MALDI chips that are utilized as MALDI plates. The method’s ability to detect PCT was confirmed by comparing the results with LC–MS bottom-up workflow. The new method detects intact PCT by its m/z and uncovers its alternations in septic serum.MethodsThe MALDI chips used for the detection of PCT were prepared by ambient ion soft landing of anti-PCT antibody on an ITO glass slide. The chips were used for the development of the rapid MALDI-TOF MS method. A parallel method based on affinity enrichment on magnetic beads followed by LC–MS/MS data-dependent peptide microsequencing was used to prove PCT presence in the sample. All samples were also tested by ELISA to determine PCT concentration prior to analyzing them by mass spectrometry methods.ResultsThe MALDI chip method was optimized using recombinant PCT spiked into the human serum. The PCT detection limit was 10 ng/mL. The optimized method was used to analyze 13 sera from patients suffering sepsis. The PCT results were confirmed by LC–MS/MS. The measurement of the intact PCT by the MALDI chip method revealed that sera of patients with severe sepsis have other forms of PCT present, which show post-processing of the primary sequence by cleavage of PCT, resulting in the formation of N and C termini fragments.ConclusionsProcalcitonin from human serum was successfully enriched and detected using immunoaffinity MALDI chips. The intact PCT was characterized in 13 septic patients. The method is more specific compared to non-MS-based immunoaffinity techniques and allows observation of different variants of PCT in septic patients.

【 授权许可】

CC BY   
© The Author(s) 2023

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