Neuromusc. Disord., Vol. 3, No. 2, pp. 135-140. 1993 0960-8966/93 $6.00 + 0.00 Printed in Great Britain ©1993 Pergamon Press Ltd SARCOLEMMAL DISTRIBUTION OF ABNORMAL DYSTROPHIN IN Xp21 CARRIERS MARIZ VAINZOF,*tLOUISE V. B. NICHOLSON,~ DENNISE. BULMAN,§ ANA MARIATSANACLIS,II MARIAR1TA PASSOS-BUENO,* RITA C. M. PAVANELLO* and MAYANA ZATZ* *Departamento de Biologia, Instituto de Bioci6ncias, Universidadede S~.oPaulo, S~o Paulo, Brazil; ,Muscular Dystrophy Group Research Laboratories, Newcastle General Hospital, Newcastle upon Tyne, U.K.; §Genetic Department, The Hospital for Sick Children, Toronto, Canada; IIDepartamento de Patologia, Faculdade de Medicina, USP, S~o Paulo, Brazil (Received 3 July 1992; revised 20 October 1992; accepted 23 November 1992) Abstract--Some Becker muscular dystrophy carriers, related to patients with specific DNA deletions, demonstrate both normal and abnormally sized dystrophin bands through qualitative Western blot analysis. The purpose of the present investigation was to assess the sarcolemmal distribution of the altered dystrophin in such carriers. Fibres expressing the normal or deleted dystrophin were identified using specific antibodies which reacted with epitopes from within the deleted region. No negative fibres or patchy immunostaining could be seen when sections from four carriers were labelled with either antibodies (C-terminal and corresponding to the deleted region), although a significant amount of abnormal dystrophin was present in their muscle (as seen on blots). Thus, we were able to confirm that in a proportion of the myonuclei, the defective allele was present on the active X chromosome. Our results suggest that the two types of nuclei were randomly distributed, resulting in normal and abnormal dystrophin molecules which were so intimately mixed that dystrophin-incompetent fibres could not be distinguished in the skeletal muscle from the Xp21 carriers. Key words: Dystrophin Western blot and immunohistochemistry, X-linked muscular dystrophy carriers. INTRODUCTION The protein dystrophin, localized in the sar- colemma of normal muscle fibres by immuno- fluorescence (IF), is absent or very reduced in muscle from Duchenne muscular dystrophy (DMD) patients and of altered size and/or quantity in Becker muscular dystrophy (BMD) [1-8]. Duchenne and Becker muscular dystrophy patients may arise from new mutations, or germinal mosaicism [9, 10], but the majority are inherited from carrier mothers [11, 12]. Im- munohistochemical studies have shown a dystrophin mosaic pattern mainly in DMD carriers with clinical manifestation [13, 14], but little or no alteration was observed among those who were asymptomatic [13, 15, 16]. tAuthor to whom correspondenceshould be addressed at: Departamento de Biologia, Universidade de S~o Paulo, Caixa Postal 11461, S~.o Paulo, Brazil. The proportion of cells with the active X chromosome bearing the abnormal allele responsible for the production of an altered protein is difficult to assess in DMD car- riers. However, some BMD carriers, related to patients with specific deletions, demonstrate both normal and abnormally sized dystrophin bands through qualitative Western blot (WB) analysis [17-19]. In these BMD heterozygotes, the presence of an additional abnormal band demonstrates that in a proportion of nuclei the X chromosome bearing the BMD allele is the active one [19]. The purpose of the present investigation was to assess the sarcolemmal distribution of dys- trophin in carriers who demonstrated both normal and abnormally sized protein on blots (appropriate for the DNA deletions of their affected relatives). Fibres expressing the normal or deleted dystrophin were tested using specific antibodies which reacted with epitopes from within the deleted region. 135