1. ¹Leiden University Medical Center, Center for Human and Clinical Genetics, Leiden, The Netherlands
2. ²Department of Molecular and Cellular Biology, University of Utrecht, Utrecht, The Netherlands
3. ³Institute of Human Genetics, International Centre for Life, Central Parkway, Newcastle upon Tyne, UK

Correspondence: Dr SM van der Maarel, Leiden University Medical Center, Center for Human and Clinical Genetics, Wassenaarseweg 72, 2333 AL Leiden, The Netherlands. Tel: +31 71 527 1915; Fax: +31 71 527 6075; E-mail: maarel@lumc.nl

⁴These authors contributed equally to this work

Received 13 October 2004; Revised 8 February 2005; Accepted 22 February 2005; Published online 13 April 2005.

Abstract

Mutations in dysferlin, a member of the fer1-like protein family that plays a role in membrane integrity and repair, can give rise to a spectrum of neuromuscular disorders with phenotypic variability including limb-girdle muscular dystrophy 2B, Myoshi myopathy and distal anterior compartment myopathy. To improve the tools available for understanding the pathogenesis of the dysferlinopathies, we have established a large source of highly specific antibody reagents against dysferlin by selection of heavy-chain antibody fragments originating from a nonimmune llama-derived phage-display library. By utilizing different truncated forms of recombinant dysferlin for selection and diverse selection methodologies, antibody fragments with specificity for two different dysferlin domains could be identified. The selected llama antibody fragments are functional in Western blotting, immunofluorescence microscopy and immunoprecipitation applications. Using these antibody fragments, we found that calpain 3, which shows a secondary reduction in the dysferlinopathies, interacts with dysferlin.