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Related Experiment Video

Updated: Apr 15, 2026

Selected Reaction Monitoring Mass Spectrometry for Absolute Protein Quantification
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Antibody-free workflows for protein quantification by LC-MS/MS.

Daniel Wilffert1, Rainer Bischoff, Nico C van de Merbel

  • 1Analytical Biochemistry, Department of Pharmacy, University of Groningen, Antonius Deusinglaan 1, 9713 AV Groningen, The Netherlands.

Bioanalysis
|April 15, 2015
PubMed
Summary

This review critically evaluates antibody-free methods for quantitative liquid chromatography-tandem mass spectrometry (LC-MS/MS) protein bioanalysis. It compares these novel techniques with traditional immunoaffinity approaches, highlighting four distinct antibody-free workflow categories.

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Area of Science:

  • Analytical Chemistry
  • Biochemistry
  • Pharmacology

Background:

  • Immunoaffinity-based approaches are widely used for quantitative protein bioanalysis via LC-MS/MS.
  • Limitations of antibody-dependent methods necessitate the exploration of alternative strategies.
  • The development of antibody-free techniques offers potential advantages in protein quantification.

Purpose of the Study:

  • To critically review and evaluate antibody-free methods for quantitative LC-MS/MS protein bioanalysis.
  • To compare the performance and applicability of antibody-free workflows against immunoaffinity-based approaches.
  • To categorize and discuss various antibody-free strategies for protein and peptide enrichment.

Main Methods:

  • Review and critical evaluation of existing literature on antibody-free protein bioanalysis.
  • Categorization of antibody-free workflows into four distinct groups based on enrichment strategies.
  • Comparative analysis of direct analysis, protein precipitation, solid-phase extraction, and affinity-based SPE methods.

Main Results:

  • Four main categories of antibody-free workflows were identified and discussed.
  • Direct analysis, fractionated protein precipitation, reversed-phase/ion-exchange SPE, and affinity-SPE represent key strategies.
  • Each method offers unique advantages and challenges for quantitative protein bioanalysis.

Conclusions:

  • Antibody-free approaches provide viable alternatives to immunoaffinity-based methods for quantitative LC-MS/MS protein bioanalysis.
  • The choice of antibody-free workflow depends on specific analytical needs and target protein characteristics.
  • Further development and validation of these methods are crucial for their broader adoption.