Journal Article

Breaking the boundaries of affinity selection-mass spectrometry: From ligand screening to target-ligand interaction insights

Cardoso, Carmen Lucia, do Amaral, Bruno Sérgio, de Oliveira, Pamella Christina Ortega, De Moraes, Marcela Cristina, Cass, Quezia Bezerra

2026

Overview

Affinity selection mass spectrometry (AS-MS) has emerged as a powerful label-free technique for identifying and characterizing ligand-target interactions. This review explores the diverse applications of AS-MS in drug discovery, including its role in selective screening, binding site characterization, and quantitative affinity determination. We discuss the use of AS-MS for determining equilibrium dissociation constants (KD) and competitive binding parameters (affinity competition experiment 50% (ACE50)), highlighting its ability to rank ligand affinities efficiently. The review also examines AS-MS applications in fragment-based drug discovery (FBDD), screening for molecular glues, and investigating interactions with membrane proteins. Moreover, we address key technical challenges, including competitive binding effects, protein stability, and ligand dissociation kinetics, along with recent advancements in automation and artificial intelligence (AI) integration. Rather than providing a comprehensive literature review, this work aims to broaden the applicability of AS-MS assays and encourage researchers to explore its use in underutilized contexts. By providing rapid and high-sensitivity affinity measurements, AS-MS continues to expand its role in drug discovery and structural biology, complementing conventional biophysical techniques. [Display omitted] •AS-MS enables quantitative KD determination using multiple experimental approaches.•Competition assays in AS-MS reveal ligand binding sites and interaction mechanisms.•Multi-target AS-MS assays allow simultaneous ligand affinity profiling.•Structural insights from AS-MS guide synthesis even in impure compound collections.•AS-MS complements biophysical techniques in drug discovery and lead optimization.

• Publisher: Elsevier B.V,Xi'an Jiaotong University,Elsevier
• Subject: Affinity selection mass spectrometry / Competition assays / Drug discovery / Fragment screening / KD determination / Ligand-target interactions / Molecular glues / Review Paper