Chemical imaging of cellular ultrastructure by null-deflection infrared spectroscopic measurements

Seth Kenkel, Mark Gryka, Lin Chen, Matthew P. Confer, Anirudha Rao, Scott Robinson, Kannanganattu V. Prasanth, Rohit Bhargava

Research output: Contribution to journalArticlepeer-review


Nearfield spectroscopic imaging techniques can be a powerful tool to map both cellular ultrastructure and molecular composition simultaneously but are currently limited in measurement capability. Resonance enhanced (RE) atomic force microscopy infrared (AFM-IR) spectroscopic imaging offers high-sensitivity measurements, for example, but probe-sample mechanical coupling, nonmolecular optical gradient forces, and noise overwhelm recorded chemical signals. Here, we analyze the key factors limiting AFM-IR measurements and propose an instrument design that enables high-sensitivity nanoscale IR imaging by combining null-deflection measurements with RE sensitivity. Our developed null-deflection scanning probe IR (NDIR) spectroscopic imaging provides ∼24× improvement in signal-to-noise ratio (SNR) compared with the state of the art, enables optimal signal recording by combining cantilever resonance with maximum laser power, and reduces background nonmolecular signals for improved analytical accuracy. We demonstrate the use of these properties for high-sensitivity, hyperspectral imaging of chemical domains in 100-nm-thick sections of cellular acini of a prototypical cancer model cell line, MCF-10A. NDIR chemical imaging enables facile recording of label-free, chemically accurate, high-SNR vibrational spectroscopic data from nanoscale domains, paving the path for routine studies of biomedical, forensic, and materials samples.

Original languageEnglish (US)
Article numbere2210516119
JournalProceedings of the National Academy of Sciences of the United States of America
Issue number47
StatePublished - Nov 22 2022


  • atomic force microscopy
  • chemical imaging
  • infrared imaging
  • nanoscale imaging
  • spectroscopy

ASJC Scopus subject areas

  • General


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