title: Quantification of Fewer than Ten Copies of a DNA Biomarker without Amplification or Labeling.
abstract: Polymerase chain reaction (PCR) is a highly sensitive diagnosis technique for detection of nucleic acids and for monitoring residual disease; however, PCR can be unreliable for samples containing very few target molecules. Here, we describe a quantification method, using force-distance (FD) curve based atomic force microscopy (AFM) to detect a target DNA bound to small (1.4-1.9 μm diameter) probe DNA spots, allowing mapping of entire spots to nanometer resolution. Using a synthetic BCR-ABL fusion gene sequence target, we examined samples containing between one and 10 target copies. A high degree of correlation (r(2) = 0.994) between numbers of target copies and detected probe clusters was observed, and the approach could detect the BCR-ABL biomarker when only a single copy was present, although multiple screens were required. Our results clearly demonstrate that FD curve-based imaging is suitable for quantitative analysis of fewer than 10 copies of DNA biomarkers without amplification, modification, or labeling.
authors.forename: Yoonhee
authors.initials: Y
authors.lastname: Lee
authors.forename: Youngkyu
authors.initials: Y
authors.lastname: Kim
authors.forename: Donggyu
authors.initials: D
authors.lastname: Lee
authors.affiliation: Nanogea Inc. , 6162 Bristol Parkway, Culver City, California 90230, United States.
authors.forename: Dhruvajyoti
authors.initials: D
authors.lastname: Roy
authors.forename: Joon Won
authors.initials: JW
authors.lastname: Park
mesh_terms: D015415:Biomarkers / Q000032:analysis; D004247:DNA / Q000235:genetics*; D015342:DNA Probes; D016044:Fusion Proteins, bcr-abl / Q000235:genetics*; D057230:Limit of Detection; D018625:Microscopy, Atomic Force / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods*
chemical_list: D015415:Biomarkers; D015342:DNA Probes; D004247:DNA; D016044:Fusion Proteins, bcr-abl