Rare Mutation Detection in Blood Plasma Using sRCA Molecule Counting Probes

• Problems in biology and medicine frequently require the ability to observe, evaluate, andcount even extremely rare macromolecules in biological samples. In particular, rare tumorspecific mutations in plasma provide valuable insights in the course of malignant disease andresponses to therapy, but simpler assay techniques are needed. We describe herein a rapidand exquisitely specific means to recognize and magnify detection signals from individualmolecules to easily recorded levels via a process we call super rolling circle amplification(sRCA). We demonstrate the ability of this technique to enumerate tumor-specific sequencevariants in plasma from cancer patients via flow cytometry at very high efficiency, with specificityadequate to detect single nucleotide mutant sequences among 100,000 copies of thenormal sequence in a 3 hr protocol. And the mutation analysis data generated from patientctDNA samples with our sRCA method are in high accordance with the patients’ primary tumorsequencing data.

Subject headings

NATURVETENSKAP  -- Biologi -- Genetik (hsv//swe)

NATURAL SCIENCES  -- Biological Sciences -- Genetics (hsv//eng)

Keyword

Rolling circle amplification

cfDNA

single molecule

digital counting

rare mutation detection

PoC application

Publication and Content Type

vet (subject category)

ovr (subject category)