Immuno-Affinity Mass Spectrometry: A Novel Approaches with Biomedical Relevance

Ion, Laura; Petre, Brînduşa Alina

doi:10.1007/978-3-030-15950-4_21

Cited by 6 publications

(2 citation statements)

References 59 publications

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“…In proteomic analysis, the ability to handle very small amounts of biological material is crucial. Miniaturized HPLC separation systems have been developed by using fused silica capillary columns [96,97] or chip-based devices [98][99][100]. Monolithic capillary columns (and other columns, such as C8 or C18 columns) can be produced in the laboratory without the need of expensive media, packing solvents, and high-pressure packing instrumentation with the proper training [101].…”

Section: The Goodmentioning

confidence: 99%

A Critical Review of Bottom-Up Proteomics: The Good, the Bad, and the Future of This Field

et al. 2020

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Proteomics is the field of study that includes the analysis of proteins, from either a basic science prospective or a clinical one. Proteins can be investigated for their abundance, variety of proteoforms due to post-translational modifications (PTMs), and their stable or transient protein–protein interactions. This can be especially beneficial in the clinical setting when studying proteins involved in different diseases and conditions. Here, we aim to describe a bottom-up proteomics workflow from sample preparation to data analysis, including all of its benefits and pitfalls. We also describe potential improvements in this type of proteomics workflow for the future.

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Section: The Goodmentioning

confidence: 99%

A Critical Review of Bottom-Up Proteomics: The Good, the Bad, and the Future of This Field

et al. 2020

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“…Moreover, immuno-capturing of both proteins and peptides has been reported to achieve relatively high sensitivity. 16 , 17) In this method, the initial step is the immuno-capturing of the antigen, followed by its elution with an aqueous acid solution. After tryptic hydrolysis, the surrogate peptide is extracted by an antipeptide antibody and quantified by LC–MS/MS.…”

Section: Introductionmentioning

confidence: 99%

Immuno-Mass Spectrometry Workflow for Quantification of Serum α-Fetoprotein Using Antibody-Immobilized Magnetic Beads and Modified Eluents

Takahata

Hara²,

Nishino

et al. 2023

Mass Spectrometry

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Immuno-mass spectrometry (MS) is a powerful method for the quantitative analysis of low-abundance proteins in biological specimens. In these procedures, collecting specifically and efficiently the target protein antigens from the antigen–antibody complex generated on the surface of nanocarrier beads is crucial and can be performed by hydrolyzing the proteins directly on the beads or after elution. Herein, we optimized the conditions of the immunoaffinity purification via elution using serum α-fetoprotein (AFP) as a model and its specific antibody immobilized covalently on magnetic beads. Antibody-coated beads were incubated with human serum spiked with standard AFP for antigen–antibody reaction. AFP was then eluted from the beads using various eluents, including organic solvents, to optimize the elution conditions. After proteolytically hydrolyzing the eluted protein, stable isotope-labeled standard peptides were added to the hydrolysate to quantify the eluted AFP via liquid chromatography–tandem MS. Using an optimized workflow for quantitative analysis afforded a correlation between the amount of spiked AFP and heavy to light ratios calculated based on peptide ion peak areas, from which an endogenous AFP concentration of 2.3±0.6 ng/mL was determined in normal serum; this is consistent with previous reports using radioimmunoassay methods. The present immuno-MS workflow could apply to the detection and quantitation of other low-abundance biofluid biomarkers.

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