A validated antibody toolbox for ALS research

A substantial fraction of amyotrophic lateral sclerosis (ALS)-associated proteins remains poorly characterized, in part because of the limited availability of validated research antibodies. We established knockout (KO)-based antibody characterization workflows and demonstrated that widely used antibodies against the major ALS-associated protein C9orf72 lacked specificity (Laflamme et al., 2019). We subsequently scaled this framework to systematically benchmark research antibodies, revealing that up to 61% fail to perform as recommended by manufacturers (Ayoubi et al., 2023b). Here, we extend this approach by establishing the ALS-Reproducible Antibody Platform (ALS-RAP), a comprehensive effort to generate a publicly available dataset of KO-validated antibodies targeting proteins encoded by ALS risk genes. In total, we characterized 303 antibodies against 33 ALS-associated proteins to identify high-quality reagents for use in western blot, immunoprecipitation, and immunofluorescence. Using these antibodies, we profiled protein levels across human induced pluripotent stem cell-derived and primary neurological cell types. These analyses revealed diverse cellular distributions and higher levels of several ALS-associated proteins in glial populations, consistent with emerging evidence for immune contributions to ALS. Together, ALS-RAP provides a validated antibody toolbox and protein expression resource to support the study of ALS-associated proteins.

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Publication Details

Journal
eLife
Published
2026-11-03
DOI
https://doi.org/10.7554/elife.111349.3
Primary Topic
Amyotrophic Lateral Sclerosis Research
Type
article
Field-Weighted Citation Impact
0.00

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article

A validated antibody toolbox for ALS research

Sara González Bolívar, Marie‐France Dorion, Guy Armand Rouleau, Brian D. Marsden et al.
eLife
Amyotrophic Lateral Sclerosis Research
article

A validated antibody toolbox for ALS research

Sara González Bolívar, Marie‐France Dorion, Guy Armand Rouleau, Brian D. Marsden, Michael Biddle, A.M. Edwards, Thomas M. Durcan, Valerio E. C. Piscopo, Carol X.‐Q. Chen, Vincent Soubannier, S. Gräslund, Carl Laflamme, Luke M. Healy, Edward A. Fon, Emma J. MacDougall, Congyao Zha, Patrick A. Dion, Maryam Fotouhi, O. Gileadi, Wen‐Hwa Lee, Ian McDowell, Peter Scottt McPherson, Jay P. Ross, Vincent Francis, Riham Ayoubi, Leonidas Koukouflis, Mathilde Chaineau, Vera Ruíz Moleón, Charles Alende, Wen Hwa Lee, Tracy Keates, Carolyn A. Marks
article en

Abstract

A substantial fraction of amyotrophic lateral sclerosis (ALS)-associated proteins remains poorly characterized, in part because of the limited availability of validated research antibodies. We established knockout (KO)-based antibody characterization workflows and demonstrated that widely used antibodies against the major ALS-associated protein C9orf72 lacked specificity (Laflamme et al., 2019). We subsequently scaled this framework to systematically benchmark research antibodies, revealing that up to 61% fail to perform as recommended by manufacturers (Ayoubi et al., 2023b). Here, we extend this approach by establishing the ALS-Reproducible Antibody Platform (ALS-RAP), a comprehensive effort to generate a publicly available dataset of KO-validated antibodies targeting proteins encoded by ALS risk genes. In total, we characterized 303 antibodies against 33 ALS-associated proteins to identify high-quality reagents for use in western blot, immunoprecipitation, and immunofluorescence. Using these antibodies, we profiled protein levels across human induced pluripotent stem cell-derived and primary neurological cell types. These analyses revealed diverse cellular distributions and higher levels of several ALS-associated proteins in glial populations, consistent with emerging evidence for immune contributions to ALS. Together, ALS-RAP provides a validated antibody toolbox and protein expression resource to support the study of ALS-associated proteins.

eLifeVol. 15
Montreal Neurological Institute and Hospital (CA), Centre for Human Genetics (GB), University of Leicester (GB), University of Ottawa (CA), University of Toronto (CA), Structural Genomics Consortium (CA), Karolinska Institutet (SE), University of Oxford (GB), Stanford Medicine (US), NIHR Leicester Biomedical Research Centre (GB), McGill University (CA), Stanford University (US)
University of Washington, Genome Canada, ALS Society of Canada, Motor Neurone Disease Association, National Centre for the Replacement, Refinement and Reduction of Animals in Research
Good health and well-being
Openalex Percentile: Top 100%
Amyotrophic Lateral Sclerosis Research
DOIOpen Access PDFOpenAlex
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