Second harmonic super-resolution microscopy for quantification of mRNA at single copy sensitivity

Jing Liu, Il Hoon Cho, Yi Cui, Joseph Irudayaraj

Research output: Contribution to journalArticlepeer-review


Cell-specific information on the quantity and localization of key mRNAs at single copy sensitivity in single cells is critical for evaluating basic cellular process, disease risk, and efficacy of therapy. Quantification of overexpressed mRNAs beyond the diffraction limit is constrained by the optical property of the probes and microscopy techniques. In this report, nanosized barium titanium oxide (BaTiO3, BTO) crystals were utilized as probes for mRNA quantification by a second harmonic super-resolution microscopy (SHaSM). The SHaSM was able to detect a single copy of the human epidermal growth factor receptor 2 (Her2) mRNA at a resolution of 55.6 nm with the ability to resolve multiple mRNA copies in a diffraction-limited spot. Her2 mRNA per cell was counted in SK-BR-3, MCF-7, and HeLa cell lines as 595 ± 79.1, 38.9 ± 8.26, and 1.5 ± 2.8, respectively. Our single-cell quantification results were validated with the fluorescence in situ hybridization studies and quantitative PCR, showing better specificity and selectivity over current single-molecule approaches for transcript detection. The SHaSM is expected to have an upper limit of resolving ∼104 transcripts in a single cell with the ability to monitor intracellular transcriptional dynamics at video rate. The developed approach has strong potential in clinical research and in the early diagnosis of life-threatening diseases such as cancer.

Original languageEnglish (US)
Pages (from-to)12418-12427
Number of pages10
JournalACS Nano
Issue number12
StatePublished - Dec 23 2014
Externally publishedYes


  • BaTiO nanocrystals
  • diagnostics
  • mRNA quantification
  • second harmonic generation
  • single-copy detection
  • super-resolution microscopy

ASJC Scopus subject areas

  • Materials Science(all)
  • Engineering(all)
  • Physics and Astronomy(all)


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