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METHODS article

Front. Immunol.

Sec. Vaccines and Molecular Therapeutics

Volume 16 - 2025 | doi: 10.3389/fimmu.2025.1623848

CoreTIA: a modular, statistically robust transduction inhibition assay for AAV neutralization

Provisionally accepted
Beatrix  KovácsBeatrix Kovács1,2Fanni  SomogyiFanni Somogyi1,2Viktória  SzabóViktória Szabó3Zoltan  Zsolt NagyZoltan Zsolt Nagy4Istvan  HernadiIstvan Hernadi5Ferenc  MatyasFerenc Matyas6Wim  VanduffelWim Vanduffel7Zsuzsanna  SzemlakyZsuzsanna Szemlaky8Balázs  RózsaBalázs Rózsa6,9István  UlbertIstván Ulbert1,10,11,2Dániel  HillierDániel Hillier1,10,2*
  • 1HUN-REN Research Centre for Natural Sciences, Institute of Cognitive Neuroscience and Psychology, Budapest, Hungary, Budapest, Hungary
  • 2Semmelweis University Doctoral School, Budapest, Hungary
  • 3Semmelweis University, Department of Ophthalmology,, Budapest, Hungary
  • 4Semmelweis University, Department of Ophthalmology, Budapest, Hungary
  • 5Grastyán Translational Research Center, and Szentágothai Research Center, Center for Neuroscience, University of Pécs, Pécs, Hungary
  • 6HUN-REN Institute of Experimental Medicine, Budapest, Hungary
  • 7Laboratory for Neuro-and Psychophysiology, Leuven Brain Institute, KULeuven, Leuven, Belgium
  • 8South-Pest Central Hospital, National Institute for Infectology and Hematology, Department of Hematology and Stem Cell Transplantation, Budapest, Hungary
  • 9BrainVisionCenter, Budapest, Hungary
  • 10Faculty of Information Technology and Bionics, Pázmány Péter Catholic University, Budapest, Hungary
  • 11Semmelweis University Department of Neurosurgery and Neurointervention, Budapest, Hungary

The final, formatted version of the article will be published soon.

Adeno-associated virus (AAV) gene therapy is often limited by pre-existing neutralizing antibodies (NAbs), yet current assays for NAb detection lack standardization and rarely quantify uncertainty, complicating cross-study comparisons. We present coreTIA (core Transduction Inhibition Assay), a comprehensive framework providing a modular experimental protocol and a statistically robust analysis pipeline. This integrated method delivers precise, reproducible NAb titers with quantified uncertainty for every result. coreTIA’s statistical framework enables robust estimation of neutralization even when dilution series are incomplete, helping to reduce repeat testing and minimizing sample volume requirements. Evaluation and refinement of key assay parameters support consistent performance across AAV serotypes. By providing a protocol and analysis suite as an open resource, coreTIA facilitates more consistent and transparent NAb measurement, potentially aiding assay harmonization and regulatory assessment, addressing a key barrier to progress in gene therapy research and development.

Keywords: Adeno-associated virus (AAV) gene delivery vectors, Pre-existing neutralizing antibodies (NAbs), Assay standardization and harmonization, uncertainty quantification, Bayesian statistical framework, AAV serotype optimization, Transduction inhibition assay (coreTIA - core Transduction Inhibition Assay), Open source assay protocol and analysis pipeline

Received: 06 May 2025; Accepted: 21 Jul 2025.

Copyright: © 2025 Kovács, Somogyi, Szabó, Nagy, Hernadi, Matyas, Vanduffel, Szemlaky, Rózsa, Ulbert and Hillier. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

* Correspondence: Dániel Hillier, HUN-REN Research Centre for Natural Sciences, Institute of Cognitive Neuroscience and Psychology, Budapest, Hungary, Budapest, Hungary

Disclaimer: All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article or claim that may be made by its manufacturer is not guaranteed or endorsed by the publisher.