Boorman, Dale, Pope, Iestyn ORCID: https://orcid.org/0000-0002-4104-0389, Masia, Francesco ORCID: https://orcid.org/0000-0003-4958-410X, Langbein, Wolfgang ORCID: https://orcid.org/0000-0001-9786-1023, Hood, Steve, Borri, Paola ORCID: https://orcid.org/0000-0002-7873-3314 and Watson, Peter ORCID: https://orcid.org/0000-0003-0250-7852 2021. Hyperspectral CARS microscopy and quantitative unsupervised analysis of deuterated and non-deuterated fatty acid storage in human cells. The Journal of Chemical Physics 155 (22) , 224202. 10.1063/5.0065950 |
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Abstract
Coherent anti-Stokes Raman scattering (CARS) implemented as a vibrational micro-spectroscopy modality eradicates the need for potentially perturbative fluorescent labeling while still providing high-resolution, chemically specific images of biological samples. Isotopic substitution of hydrogen atoms with deuterium introduces minimal change to molecular structures and can be coupled with CARS microscopy to increase chemical contrast. Here, we investigate HeLa cells incubated with non-deuterated or deuterium-labeled fatty acids, using an in-house-developed hyperspectral CARS microscope coupled with an unsupervised quantitative data analysis algorithm, to retrieve Raman susceptibility spectra and concentration maps of chemical components in physically meaningful units. We demonstrate that our unsupervised analysis retrieves the susceptibility spectra of the specific fatty acids, both deuterated and non-deuterated, in good agreement with reference Raman spectra measured in pure lipids. Our analysis, using the cell-silent spectral region, achieved excellent chemical specificity despite having no prior knowledge and considering the complex intracellular environment inside cells. The quantitative capabilities of the analysis allowed us to measure the concentration of deuterated and non-deuterated fatty acids stored within cytosolic lipid droplets over a 24 h period. Finally, we explored the potential use of deuterium-labeled lipid droplets for non-invasive cell tracking, demonstrating an effective application of the technique for distinguishing between cells in a mixed population over a 16 h period. These results further demonstrate the chemically specific capabilities of hyperspectral CARS microscopy to characterize and distinguish specific lipid types inside cells using an unbiased quantitative data analysis methodology
Item Type: | Article |
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Date Type: | Publication |
Status: | Published |
Schools: | Physics and Astronomy Biosciences |
Publisher: | American Institute of Physics |
ISSN: | 0021-9606 |
Date of First Compliant Deposit: | 15 December 2021 |
Date of Acceptance: | 14 November 2021 |
Last Modified: | 05 Jan 2024 07:43 |
URI: | https://orca.cardiff.ac.uk/id/eprint/146159 |
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