Table 3. Reproducibility of detecting differences in dystrophin expression in muscle biopsies with a wide range of dystrophin levels from healthy controls to BMD and DMD samples by immunofluorescence using MANDYS106 antibody and Definiens intensity measurements (at ‘control’ microscope settings).

Sample	Experiment	Number of sections	Number of fibers analyzed	Cross sectional area of individual fibers; Mean ± SD (x103 µm2)	Dystrophin membrane intensity MANDYS106 (au)
					Mean	Intra-assay precision CV% sections	Inter-assay precision CV% experiments	Q90 mean	Maximum	Ranking
Control 2	1	2	625	4.4±1.8	929	13%	13%	2397	3609	1
	2	2	544	4.3±1.9	768	3%		2169	3640	1
BMD 1	1	2	262	4.8±2.7	415	3%	9%	1224	2425	2
	2	2	337	4.9±2.2	367	6%		1076	2195	3
BMD 3	1	2	173	7.2±3.6	387	13%	4%	1125	2366	3
	2	2	221	7.2±3.8	412	7%		1195	2439	2
BMD 4	1	2	250	5.8±3.9	359	4%	7%	1062	2152	4
	2	2	102	7.7±4.9	323	6%		993	2122	4
BMD 2	1	2	300	4.4±3.3	200	7%	4%	668	1553	5
	2	2	197	6.4±4.2	213	4%		689	1552	5
DMD 1	1	2	1102	2.3±1.1	58	6%	24%	154	341	6
	2	2	739	3.0±1.8	82	13%		198	399	6
DMD 2	1	2	937	2.6±1.5	44	10%	43%	115	243	7
	2	2	816	2.5±1.4	82	6%		184	334	7

The intra-assay precision (CV%) was calculated for the mean dystrophin intensity of multiple sections of a biopsy/sample analyzed in the same experiment. Inter-assay precision between experiments performed on different days was assessed in two ways by ranking of the samples based on the dystrophin intensity in each experiment and by calculating the CV% from the mean dystrophin intensity in each experiment (experiment 1 and 2 performed 102 days apart).

BMD, Becker muscular dystrophy; CV, coefficient of variation; DMD, Duchenne muscular dystrophy; SD, standard deviation.