Absolute quantification of dystrophin protein in human muscle biopsies using parallel reaction monitoring (PRM). (8th November 2019)

• Record Type: Journal Article
• Authors: Canessa, Emily H.; Goswami, Mansi V.; Alayi, Tchilabalo D.; Hoffman, Eric P.; Hathout, Yetrib
• Other Names: Fabris Dan guestEditor.
• Is Part Of: Journal of mass spectrometry. Volume 55:Number 2(2020)
• Page Start: n/a
• Page End: n/a
• Publication Date: 2019-11-08
• Subjects: dystrophin -- muscular dystrophy -- parallel reaction monitoring -- skeletal muscle -- targeted mass spectrometry quantification; Mass spectrometry -- Periodicals; 543.65
• DOI: 10.1002/jms.4437
• Languages: English
• ISSNs: 1076-5174

Abstract

Abstract: The need for a reliable and accurate method to quantify dystrophin proteins in human skeletal muscle biopsies has become crucial in order to assess the efficacy of dystrophin replacement therapies in Duchenne muscular dystrophy as well as to gain insight into the relationship between dystrophin levels and disease severity in Becker's muscular dystrophy. Current methods to measure dystrophin such as western blot and immunofluorescence, while straightforward and simple, lack precision and sometimes specificity. Here, we standardized a targeted mass spectrometry method to determine the absolute amount of dystrophin in ng/mg of muscle using full‐length 13 C6–Arg– and 13 C6, 15 N2–Lys–labeled dystrophin and parallel reaction monitoring (PRM). The method was found to be reproducible with a limit of quantification as low as 30 pg of dystrophin protein per mg of total muscle proteins. The method was then tested to measure levels of dystrophin in muscle biopsies from a healthy donor and from Duchenne and Becker's muscular dystrophy patients.