Live cell metabolic imaging of cancer cell lines using multiphoton fluorescence polarization

Einstein Gnanatheepam; Anandh Sundaramoorthy; Bharanidharan Ganesan; Kailasha Purushothaman; Durgalakshmi Dhinasekaran; Aruna Prakasarao; Ganesan Singaravelu

doi:10.1117/12.2545847

14 February 2020 Live cell metabolic imaging of cancer cell lines using multiphoton fluorescence polarization

Einstein Gnanatheepam, Anandh Sundaramoorthy, Bharanidharan Ganesan, Kailasha Purushothaman, Durgalakshmi Dhinasekaran, Aruna Prakasarao, Ganesan Singaravelu

Author Affiliations +

Proceedings Volume 11244, Multiphoton Microscopy in the Biomedical Sciences XX; 112441B (2020) https://doi.org/10.1117/12.2545847
Event: SPIE BiOS, 2020, San Francisco, California, United States

Abstract

Metabolic imaging of live cell may allow in understanding the molecular level changes in cells under various diseased state, including cancer. The intrinsic fluorophores, Nicotinamide adenine dinucleotide (NADH) and flavin adenine dinucleotide (FAD) are crucial for electron transfer in the oxidation-reduction reactions in the cell. Metabolic imaging based on fluorescence polarization enables to analyze both biochemical distribution and their conformation. In this study, multiphoton fluorescence polarization imaging of NADH and FAD from cancer and normal cell lines of epithelial origin were carried out. Spectral deconvolution method was adopted to isolate fluorescence emission from different coenzymes NADH and FAD. The observed heterogeneity of the multiphoton autofluorescence in live cells was used in intensity-toconcentration image conversion. The multiphoton autofluorescence exhibits anisotropy features at the cellular level, that directly indicate the presence of NADH and FAD in two differing conformation states viz; free and protein-bound. Mapping of anisotropy of cellular autofluorescence enables to probe the distribution of population fractions of free and bound forms of NADH and FAD. Further, the redox ratio between normal and cancer cell lines confirms the changes in the metabolic activities between them. These molecular-level studies demonstrate the potential of probing cellular metabolism associated with cancer, without the need for cell destruction as in the case of conventional biochemical assays.

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Einstein Gnanatheepam, Anandh Sundaramoorthy, Bharanidharan Ganesan, Kailasha Purushothaman, Durgalakshmi Dhinasekaran, Aruna Prakasarao, and Ganesan Singaravelu "Live cell metabolic imaging of cancer cell lines using multiphoton fluorescence polarization", Proc. SPIE 11244, Multiphoton Microscopy in the Biomedical Sciences XX, 112441B (14 February 2020); https://doi.org/10.1117/12.2545847

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