Molecular Combing compared to Southern Blot in Measuring D4Z4 Contractions in FSHD (P2.027)

OBJECTIVE: To compare the analysis of the facioscapulohumeral muscular dystrophy type 1 locus (FSHD1) on chromosome 4q35-qter (chr 4q) by Southern blot (SB) to molecular combing (MC) in genomic DNA specimens referred to a clinical laboratory for FSHD testing. BACKGROUND: Current diagnostic testing for FSHD1 by SB may lead to indeterminate results in up to 23[percnt] of cases. MC may represent an improvement over SB by its analytical ability to directly visualize D4Z4 macrosatellite repeat arrays on specific chr 4q and chr 10q alleles. DESIGN/METHODS: A de-identified set of 87 genomic DNA specimens determined by SB as normal (n=71), abnormal with D4Z4 macrosatellite repeat array contractions (n=7), indeterminate (n=6), borderline (n=2), and mosaic (n=1) were blinded and independently re-analyzed by MC. RESULTS: The results were the same on SB and MC in 86.2[percnt] of cases. All contractions (n=7) and mosaics (n=1) detected by SB were confirmed by MC. Sixty-seven of 71 normal SB results were concordant with MC (94.4[percnt]). The four discrepancies were either mosaic (n=2), rearranged (n=1), or borderline by MC (n=1). All indeterminate SB results (n=6) were resolved by MC as either normal (n=4), borderline (n=1), or rearranged (n=1). The two borderline SB results showed a D4Z4 contraction on the chr 4qA allele and a normal result by MC. CONCLUSIONS: MC overcomes a number of technical limitations of SB including the direct visualization of D4Z4 macrosatellite repeat arrays on specific chr 4q and chr 10q alleles, and more precise D4Z4 repeat sizing. This study suggests that the analytical validity of MC is superior to SB in determining D4Z4 contraction size, detecting mosaicism and resolving borderline and indeterminate SB results. Further studies are needed to establish the clinical validity of these findings. Study Supported by: Quest Diagnostics Disclosure: Dr. Vasale has received personal compensation for activities with Quest Diagnostics. Dr. Jocson has received personal compensation for activities with Quest Diagnostics. Dr. Boyar has received personal compensation for activities with Quest Diagnostics. Dr. Liaquat has received personal compensation for activities with Quest Diagnostics. Dr. Hoffman has received personal compensation for activities with Quest Diagnostics. Dr. Sulcova has received personal compensation for activities with Quest Diagnostics as an employee. Dr. Tsao has received personal compensation for activities with Quest Diagnostics. Dr. Hensley has received personal compensation for activities with Quest Diagnostics. Dr. Liu has received personal compensation for activities with Quest Diagnostics. Dr. Chan has received personal compensation for activities with Quest Diagnostics as an employee. Dr. Owen has received personal compensation for activities with Quest Diagnostics. Dr. Braastad has received personal compensation for activities with Quest Diagnostics. Dr. Sun has received personal compensation for activities with Quest Diagnostics as an employee. Dr. Anguiano has received personal compensation for activities with Quest Diagnostics. Dr. Jaremko has received personal compensation for activities with Quest Diagnostics. Dr. Wang has received personal compensation for activities with Quest Diagnostics. Dr. Suer has received personal compensation for activities with Quest Diagnostics. Dr. Batish has received personal compensation for activities with Quest Diagnostics. Dr. Strom has received personal compensation for activities with Quest Diagnostics. Dr. Higgins has received personal compensation for activities with Quest Diagnostics.