TY - JOUR
T1 - Chemical imaging of cellular ultrastructure by null-deflection infrared spectroscopic measurements
AU - Kenkel, Seth
AU - Gryka, Mark
AU - Chen, Lin
AU - Confer, Matthew P.
AU - Rao, Anirudha
AU - Robinson, Scott
AU - Prasanth, Kannanganattu V.
AU - Bhargava, Rohit
N1 - ACKNOWLEDGMENTS. Research reported in this publication was supported in part by the National Institute of Biomedical Imaging and Bioengineering of the NIH under award Nos. T32EB019944 and R01EB009745 as well as by the National Science Foundation via award number 2153032. KVP laboratory is supported by grants from NIH (AG065748 and GM132458) and National Science Foundation (EAGER, 1723008), and Cancer Center at Illinois seed grants and Prairie Dragon Paddlers. The content is solely the responsibility of the authors and does not necessarily represent the official views of the NIH.
PY - 2022/11/22
Y1 - 2022/11/22
N2 - 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.
AB - 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.
KW - atomic force microscopy
KW - chemical imaging
KW - infrared imaging
KW - nanoscale imaging
KW - spectroscopy
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U2 - 10.1073/pnas.2210516119
DO - 10.1073/pnas.2210516119
M3 - Article
C2 - 36375054
AN - SCOPUS:85141940261
SN - 0027-8424
VL - 119
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
IS - 47
M1 - e2210516119
ER -