Rapid detection of allelic losses in brain tumours using microsatellite repeat markers and high-performance liquid chromatography.
High-performance liquid chromatography (HPLC) is a recently introduced high-capacity automated method for detecting unknown mutations (denaturing HPLC) or for sizing DNA fragments under nondenaturing conditions. We have adapted the HPLC method for detection of loss of heterozygosity (LOH) and used glial tumours as a model to evaluate its sensitivity and specificity in comparison to conventional denaturing polyacrylamide gel electrophoresis. A total of 20 oligodendrogliomas (grades II and III), and five astrocytic tumours (grades III and IV) were analysed using 14 polymorphic microsatellite markers mapping to regions on chromosomes 1p, 19q, and 10q using both DNA-HPLC and denaturing gel electrophoresis. This study demonstrated complete concordance between both methods. However, unlike gel electrophoresis, HPLC is automated, does not require post-PCR processing, and does not require hazardous radioactive or expensive fluorescent labelling. Our data suggest that HPLC is a reliable and sensitive method for detection of allelic losses in tumour samples and it is a favourable alternative for high-sensitivity LOH detection in both research and diagnostic environments.
Chernova
O B
OB
Department of Neurosurgery, Cleveland Clinic Foundation, 9500 Euclid Avenue, Cleveland, OH 44195, USA.
Barnett
G H
GH
Cowell
J K
JK
D001932:Brain Neoplasms / Q000235:genetics*; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D019656:Loss of Heterozygosity; D018895:Microsatellite Repeats; D016133:Polymerase Chain Reaction; D012680:Sensitivity and Specificity