Atomic force microscopic detection enabling multiplexed low-cycle-number quantitative polymerase chain reaction for biomarker assays

Andrey Mikheikin, Anita Olsen, Kevin Leslie, Bud Mishra, James K. Gimzewski, Jason Reed

Research output: Contribution to journalArticle

Abstract

Quantitative polymerase chain reaction is the current "golden standard" for quantification of nucleic acids; however, its utility is constrained by an inability to easily and reliably detect multiple targets in a single reaction. We have successfully overcome this problem with a novel combination of two widely used approaches: target-specific multiplex amplification with 15 cycles of polymerase chain reaction (PCR), followed by single-molecule detection of amplicons with atomic force microscopy (AFM). In test experiments comparing the relative expression of ten transcripts in two different human total RNA samples, we find good agreement between our single reaction, multiplexed PCR/AFM data, and data from 20 individual singleplex quantitative PCR reactions. This technique can be applied to virtually any analytical problem requiring sensitive measurement concentrations of multiple nucleic acid targets.

Original languageEnglish (US)
Pages (from-to)6180-6183
Number of pages4
JournalAnalytical chemistry
Volume86
Issue number13
DOIs
StatePublished - Jul 1 2014

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ASJC Scopus subject areas

  • Analytical Chemistry

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