Purpose: To compare the accuracy of a commercially available MLPA kit with a laboratory developed RT-PCR assay for the detection of SMN1 and SMN2 copy numbers in clinical samples. Methods: We developed and validated a...Purpose: To compare the accuracy of a commercially available MLPA kit with a laboratory developed RT-PCR assay for the detection of SMN1 and SMN2 copy numbers in clinical samples. Methods: We developed and validated a laboratory developed real time PCR based test capable of detecting SMN1 and SMN2 copy numbers in individuals. We also validated an MLPA kit purchased from MRC Holland for the same purpose. We then analyzed a series of 1027 anonymized samples using both technologies. When discrepant results were obtained, each sample was re-analyzed at least twice using both platforms. Results: Five samples did not yield results in either assay. For SMN1 copy number analysis, 2 RT-PCR assays revealed indeterminant results and all 1020 other samples were concordant for SMN1 copy number. There were 9 discrepancies in SMN2 copy number determination mostly due to a variability in MLPA analysis. Conclusion: Both MLPA and RTPCR assays give a reliable estimate of SMN1 copy number and are therefore appropriate for population based carrier screening for Spinal Muscular Atrophy Type 1. The MLPA kit has a low incidence (<1%) of underestimating the SMN2 copy number by 1 copy, but this inconsistency is of little clinical significance and can be overcome by replicate testing.展开更多
Purpose: To compare the sensitivity of Hexosaminidase A (HexA) enzyme-based testing to gene sequencing for carrier detection in non-Jewish individuals. Methods: Blood samples were obtained from parents and relatives o...Purpose: To compare the sensitivity of Hexosaminidase A (HexA) enzyme-based testing to gene sequencing for carrier detection in non-Jewish individuals. Methods: Blood samples were obtained from parents and relatives of affected patients at an annual Tay-Sachs and Allied Diseases Foundation meeting. A family history was taken for each individual. Samples were analyzed for leukocyte HexA activity, serum HexA activity and subjected to extensive gene sequencing. The results from these analyses were combined with our previously published data describing 34 obligate Tay-Sachs disease (TSD) carriers. Results: Twelve additional TSD carriers were detected in this study. Gene sequencing successfully identified all 12 carriers whereas enzyme analysis identified 11 of 12 carriers. This individual is a carrier of the B1 variant that is known to cause false negative results with enzyme testing. Combined data from 46 non-Jewish TSD carriers revealed that gene sequencing had a higher sensitivity rate than HexA enzyme-based testing (94% versus 87%) in non-Jewish TSD carriers. In our series, approximately 4% of non-Jewish TSD carriers have this mutation. Conclusions: HexA gene sequencing provides a higher sensitivity for TSD carrier detection than HexA based enzyme analysis in non-Jewish patients primarily due to the presence of individuals with the B1 variant.展开更多
文摘Purpose: To compare the accuracy of a commercially available MLPA kit with a laboratory developed RT-PCR assay for the detection of SMN1 and SMN2 copy numbers in clinical samples. Methods: We developed and validated a laboratory developed real time PCR based test capable of detecting SMN1 and SMN2 copy numbers in individuals. We also validated an MLPA kit purchased from MRC Holland for the same purpose. We then analyzed a series of 1027 anonymized samples using both technologies. When discrepant results were obtained, each sample was re-analyzed at least twice using both platforms. Results: Five samples did not yield results in either assay. For SMN1 copy number analysis, 2 RT-PCR assays revealed indeterminant results and all 1020 other samples were concordant for SMN1 copy number. There were 9 discrepancies in SMN2 copy number determination mostly due to a variability in MLPA analysis. Conclusion: Both MLPA and RTPCR assays give a reliable estimate of SMN1 copy number and are therefore appropriate for population based carrier screening for Spinal Muscular Atrophy Type 1. The MLPA kit has a low incidence (<1%) of underestimating the SMN2 copy number by 1 copy, but this inconsistency is of little clinical significance and can be overcome by replicate testing.
文摘Purpose: To compare the sensitivity of Hexosaminidase A (HexA) enzyme-based testing to gene sequencing for carrier detection in non-Jewish individuals. Methods: Blood samples were obtained from parents and relatives of affected patients at an annual Tay-Sachs and Allied Diseases Foundation meeting. A family history was taken for each individual. Samples were analyzed for leukocyte HexA activity, serum HexA activity and subjected to extensive gene sequencing. The results from these analyses were combined with our previously published data describing 34 obligate Tay-Sachs disease (TSD) carriers. Results: Twelve additional TSD carriers were detected in this study. Gene sequencing successfully identified all 12 carriers whereas enzyme analysis identified 11 of 12 carriers. This individual is a carrier of the B1 variant that is known to cause false negative results with enzyme testing. Combined data from 46 non-Jewish TSD carriers revealed that gene sequencing had a higher sensitivity rate than HexA enzyme-based testing (94% versus 87%) in non-Jewish TSD carriers. In our series, approximately 4% of non-Jewish TSD carriers have this mutation. Conclusions: HexA gene sequencing provides a higher sensitivity for TSD carrier detection than HexA based enzyme analysis in non-Jewish patients primarily due to the presence of individuals with the B1 variant.
