ArticleAnalytical chemistry2025
Ultrafast Microdroplet Digestion of Antibodies with Fc-Silencing Mutations.
Article in Analytical chemistry, 2025. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 2 papers.
What it found
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The abstract states no effect estimate the extractor could read, or names no intervention and outcome on the map, so this paper lights no cell and moves no belief. It is still indexed, cited and linked below.
The trial behind it
Trials whose registry record cites this paper, or whose number appears in the abstract. A trial that started after this paper was published is citing it as background, not reporting it.
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Who cites it
2 citing papers in PubMed.
- A Unified Mass Spectrometry Workflow Integrating Bottom-Up and Native Top-Down Proteomics via Nanodroplet Enzymatic Digestion.Analytical chemistry · 2026Article
- Rapid Antibody Structural Characterization and Quantification via Microdroplet Trypsin Digestion.Journal of the American Society for Mass Spectrometry · 2026Article
Corrections and comments
PubMed lists nothing against this paper. Absence here is not a guarantee, only a check that was made.
Authors and funding
10 authors.
Funding
Abstract
Many novel therapeutic monoclonal antibody (mAb) modalities contain mutations that not only silence unwanted binding to Fc-gamma receptors but also could bring resistance to IdeS enzymatic cleavage into mAb subunits, limiting the antibody middle-down analysis by mass spectrometry (MS). Herein we showed, for the first time, the efficient, reproducible, and ultrafast microdroplet digestion (less than 1 ms) of mAbs carrying a series of mutations "LALA", "LAGA", and "LFLE" (e.g., tool antibody LALA-DS, Nivolumab, Pembrolizumab, Vedolizumab, and PD-L1), by a new enzyme: FabRICATOR Xtra (Xtra). The digestion took place during the spray ionization process using an Agilent jet stream (AJS) ion source with a digestion efficiency close to or more than 80%, leading to subunits with high ion abundances for identification and characterization. Increased enzyme/antibody ratio or partial reduction of disulfide bonds increased the digestion efficiency. "One-pot" disulfide reduction and digestion in microdroplets could occur simultaneously by spraying the antibody and enzyme along with the reductant. Notably, for the highly digestion-resistant PD-L1 antibody, the Xtra-based microdroplet digestion process was found to be 9 million times faster than in-solution digestion. Furthermore, a workflow was developed, using a script that can automatically choose a preferred enzyme (Xtra or IdeS) for digestion, based on a target antibody input sequence, which allowed quick analysis of 94 antibody samples within 104 min. Our method is a fully automated microdroplet protein digestion technique that integrates flow injection (FI) and online MS analysis, providing a rapid and robust method for the structural characterization of mAbs with mutations.
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.