title: COLD-PCR: improving the sensitivity of molecular diagnostics assays.
abstract: The detection of low-abundance DNA variants or mutations is of particular interest to medical diagnostics, individualized patient treatment and cancer prognosis; however, detection sensitivity for low-abundance variants is a pronounced limitation of most currently available molecular assays. We have recently developed coamplification at lower denaturation temperature-PCR (COLD-PCR) to resolve this limitation. This novel form of PCR selectively amplifies low-abundance DNA variants from mixtures of wild-type and mutant-containing (or variant-containing) sequences, irrespective of the mutation type or position on the amplicon, by using a critical denaturation temperature. The use of a lower denaturation temperature in COLD-PCR results in selective denaturation of amplicons with mutation-containing molecules within wild-type mutant heteroduplexes or with a lower melting temperature. COLD-PCR can be used in lieu of conventional PCR in several molecular applications, thus enriching the mutant fraction and improving the sensitivity of downstream mutation detection by up to 100-fold.
authors.affiliation: Division of DNA Repair and Genome Stability, Department of Radiation Oncology, Dana-Farber/Brigham and Women's Cancer Center, Harvard Medical School, Boston, MA, USA.
authors.forename: Coren A
authors.initials: CA
authors.lastname: Milbury
authors.forename: Jin
authors.initials: J
authors.lastname: Li
authors.forename: Pingfang
authors.initials: P
authors.lastname: Liu
authors.forename: G Mike
authors.initials: GM
authors.lastname: Makrigiorgos
mesh_terms: D003080:Cold Temperature; D004247:DNA / Q000032:analysis; D006801:Humans; D009154:Mutation; D009691:Nucleic Acid Denaturation; D057089:Pathology, Molecular / Q000379:methods* / Q000592:standards*; D016133:Polymerase Chain Reaction / Q000379:methods* / Q000592:standards*; D012680:Sensitivity and Specificity; D016159:Tumor Suppressor Protein p53 / Q000235:genetics
chemical_list: D016159:Tumor Suppressor Protein p53; D004247:DNA