Journal
NEUROLOGY
Volume 83, Issue 22, Pages 2062-2069Publisher
LIPPINCOTT WILLIAMS & WILKINS
DOI: 10.1212/WNL.0000000000001025
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Funding
- European Union's seventh framework programme (FP7) under SKIP-NMD grant [305370]
- Association Francaise contre les Myopathies (AFM)
- Wellcome Trust [HICF-1009-025]
- TREAT-NMD Neuromuscular Network
- Great Ormond Street Hospital Children's Charity
- National Institute for Health Research Biomedical Research Centre at Great Ormond Street Hospital for Children NHS Foundation Trust
- University College London
- Wellcome Trust University Award
- National Institute of Neurological Diseases and Stroke [R01 NS043264]
- CureDuchenne
- MRC [MR/K000608/1] Funding Source: UKRI
- Medical Research Council [MR/K000608/1] Funding Source: researchfish
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Objective:We formed a multi-institution collaboration in order to compare dystrophin quantification methods, reach a consensus on the most reliable method, and report its biological significance in the context of clinical trials.Methods:Five laboratories with expertise in dystrophin quantification performed a data-driven comparative analysis of a single reference set of normal and dystrophinopathy muscle biopsies using quantitative immunohistochemistry and Western blotting. We developed standardized protocols and assessed inter- and intralaboratory variability over a wide range of dystrophin expression levels.Results:Results from the different laboratories were highly concordant with minimal inter- and intralaboratory variability, particularly with quantitative immunohistochemistry. There was a good level of agreement between data generated by immunohistochemistry and Western blotting, although immunohistochemistry was more sensitive. Furthermore, mean dystrophin levels determined by alternative quantitative immunohistochemistry methods were highly comparable.Conclusions:Considering the biological function of dystrophin at the sarcolemma, our data indicate that the combined use of quantitative immunohistochemistry and Western blotting are reliable biochemical outcome measures for Duchenne muscular dystrophy clinical trials, and that standardized protocols can be comparable between competent laboratories. The methodology validated in our study will facilitate the development of experimental therapies focused on dystrophin production and their regulatory approval.
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