Related Experiment Video
Updated: Sep 27, 2025
Efficient and Scalable Production of Full-length Human Huntingtin Variants in Mammalian Cells using a Transient Expression System
Published on: December 10, 2021
Development of mAb-based polyglutamine-dependent and polyglutamine length-independent huntingtin quantification
David F Fischer1, Sipke Dijkstra2, Kimberly Lo2
1Charles River, Chesterford Research Park, Saffron Walden, United Kingdom.
Insights
New monoclonal antibody assays accurately measure mutant huntingtin protein (mHTT) levels in Huntington's disease (HD) research. These validated assays offer a sustainable tool for tracking therapeutic progress in HD drug development.
Area of Science:
- Neuroscience
- Genetics
- Biochemistry
Background:
- Huntington's disease (HD) is a neurodegenerative disorder caused by a CAG trinucleotide repeat expansion in the huntingtin gene, leading to mutant huntingtin protein (mHTT) expression.
- Current therapeutic strategies focus on reducing mHTT levels, necessitating reliable methods for its quantification.
Purpose of the Study:
- To develop and validate sensitive and selective assays using monoclonal antibodies for measuring human huntingtin protein (HTT) levels.
- To provide an accessible and sustainable resource for the HD research community.
Main Methods:
- Development of electrochemiluminescence assays on the Meso Scale Discovery platform utilizing monoclonal antibodies.
- Assays were designed to measure HTT in a polyglutamine-independent manner (detecting both mutant and non-mutant forms).
- Validation involved testing in cellular and animal models, as well as human patient biosamples, with cross-site validation performed.
Main Results:
- The newly developed monoclonal antibody-based assays demonstrated sensitivity comparable to previously established polyclonal antibody assays.
- Cross-site validation confirmed the reliability and reproducibility of the assays across different laboratories.
- The assays effectively quantify HTT levels in various biological matrices relevant to HD research.
Conclusions:
- Monoclonal antibody-based assays provide a sensitive, selective, and sustainable method for quantifying HTT in Huntington's disease research.
- These validated assays facilitate direct comparison of results across studies and laboratories, supporting therapeutic development.
- The developed assays represent a valuable resource for advancing the understanding and treatment of Huntington's disease.
Abstract:
Huntington's disease (HD) is caused by an expansion of the CAG trinucleotide repeat domain in the huntingtin gene that results in expression of a mutant huntingtin protein (mHTT) containing an expanded polyglutamine tract in the amino terminus. A number of therapeutic approaches that aim to reduce mHTT expression either locally in the CNS or systemically are in clinical development. We have previously described sensitive and selective assays that measure human HTT proteins either in a polyglutamine-independent (detecting both mutant expanded and non-expanded proteins) or in a polyglutamine length-dependent manner (detecting the disease-causing polyglutamine repeats) on the electrochemiluminescence Meso Scale Discovery detection platform. These original assays relied upon polyclonal antibodies. To ensure an accessible and sustainable resource for the HD field, we developed similar assays employing monoclonal antibodies. We demonstrate that these assays have equivalent sensitivity compared to our previous assays through the evaluation of cellular and animal model systems, as well as HD patient biosamples. We also demonstrate cross-site validation of these assays, allowing direct comparison of studies performed in geographically distinct laboratories.
