Mutation analysis of MLH1 and MSH2 genes performed by denaturing high-performance liquid chromatography

J Biochem Biophys Methods. 2002 Mar 4;51(1):89-100. doi: 10.1016/s0165-022x(02)00003-9.


Mutation analysis of large genes, such as MSH2 and MLH1, is time-consuming and expensive. We investigated the sensitivity and specificity of DHPLC analysis for the detection of mutations within both MSH2 and MLH1. Studies included a series of 46 patients affected by colorectal cancer from HNPCC families. We confirmed 19 changes previously identified by DNA sequencing and, in a blind study, an additional 16 rare alterations including four mutations not previously described. Generally, false negative results were not observed. Elution profiles were highly characteristic for a given change and in 98.5% cases allowed the distinction between novel alterations and previously identified mutations and polymorphisms. For the detection of changes in almost all amplicons, it was sufficient to use just one denaturing temperature. DHPLC was confirmed to be highly sensitive, specific and a cost-effective technique with particularly high potential for the detection of MSH2 and MLH1 gene mutations in the diagnostic setting.

MeSH terms

  • Adaptor Proteins, Signal Transducing
  • Carrier Proteins
  • Chromatography, High Pressure Liquid / methods*
  • DNA Mutational Analysis*
  • DNA-Binding Proteins*
  • False Negative Reactions
  • Family Health
  • Female
  • Humans
  • Male
  • MutL Protein Homolog 1
  • MutS Homolog 2 Protein
  • Mutation
  • Neoplasm Proteins / genetics*
  • Nuclear Proteins
  • Polymorphism, Genetic
  • Proto-Oncogene Proteins / genetics*
  • Sensitivity and Specificity
  • Sequence Analysis, DNA
  • Time Factors


  • Adaptor Proteins, Signal Transducing
  • Carrier Proteins
  • DNA-Binding Proteins
  • MLH1 protein, human
  • Neoplasm Proteins
  • Nuclear Proteins
  • Proto-Oncogene Proteins
  • MSH2 protein, human
  • MutL Protein Homolog 1
  • MutS Homolog 2 Protein