Schollen E, Vandenberk P, Cassiman J J, Matthijs G
Center for Human Genetics, University of Leuven, Belgium.
Clin Chem. 1997 Jan;43(1):18-23.
We developed a diagnostic test based on the reverse dot-blot principle, in which five mitochondrial point mutations responsible for Leber hereditary optic neuropathy (LHON) were screened simultaneously. A series of wild-type and mutant oligonucleotides representing each mutation were covalently bound to a single nylon membrane strip. The target sites were amplified in a multiplex PCR and the products were hybridized to the membrane. Detection is based on chemiluminescence. To test the developed assay, 47 patients suspected of having LHON were screened. In 11 cases (23%) the diagnosis of LHON could be confirmed (3460, 1; 9804, 1; 11778, 5; 14484, 3; 15257, 1). The results suggest that the clinical identification of the mitochondrial defect is not trivial and the availability of a rapid screening method simplifies the molecular analysis of these cases.
我们基于反向斑点杂交原理开发了一种诊断测试方法，可同时筛查导致Leber遗传性视神经病变（LHON）的五种线粒体点突变。一系列代表每种突变的野生型和突变型寡核苷酸被共价结合到单个尼龙膜条上。通过多重PCR扩增靶位点，产物与膜杂交。检测基于化学发光。为测试所开发的检测方法，对47例疑似LHON的患者进行了筛查。在11例（23%）中确诊为LHON（3460，1例；9804，1例；11778，5例；14484，3例；15257，1例）。结果表明，线粒体缺陷的临床鉴定并非易事，快速筛查方法的可用性简化了这些病例的分子分析。
