Colocalization of cellular nanostructure using confocal fluorescence and partial wave spectroscopy

A new multimodal confocal microscope has been developed, which includes a parallel Partial Wave Spectroscopic (PWS) microscopy path. This combination of modalities allows molecular‐specific sensing of nanoscale intracellular structure using fluorescent labels. Combining molecular specificity and sen...

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Published in:	Journal of biophotonics Vol. 10; no. 3; pp. 377 - 384
Main Authors:	Chandler, John E., Stypula‐Cyrus, Yolanda, Almassalha, Luay, Bauer, Greta, Bowen, Leah, Subramanian, Hariharan, Szleifer, Igal, Backman, Vadim
Format:	Journal Article
Language:	English
Published:	Weinheim WILEY‐VCH Verlag 01.03.2017 Wiley Subscription Services, Inc
Subjects:	Cancer Cancer screening Cell Nucleus - drug effects Cell Nucleus - ultrastructure Cheek chromatin Confocal confocal microscopy Fluorescence HeLa Cells Heterogeneity Humans hyperspectral microscopy Image Processing, Computer-Assisted Intracellular Ionophores - pharmacology Localization Medical imaging Medical screening Microscopy, Confocal - instrumentation Microscopy, Confocal - methods Microscopy, Fluorescence - instrumentation Microscopy, Fluorescence - methods Mitochondria Mitochondria - drug effects Mitochondria - ultrastructure Molecular structure Multimodal Imaging nanocytology Nanostructure Oxidative phosphorylation Phosphorylation Spectroscopy Spectrum Analysis - instrumentation Spectrum Analysis - methods Valinomycin Valinomycin - pharmacology hyperspectral microscopy chromatin confocal microscopy nanocytology mitochondria
ISSN:	1864-063X, 1864-0648, 1864-0648
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Abstract	A new multimodal confocal microscope has been developed, which includes a parallel Partial Wave Spectroscopic (PWS) microscopy path. This combination of modalities allows molecular‐specific sensing of nanoscale intracellular structure using fluorescent labels. Combining molecular specificity and sensitivity to nanoscale structure allows localization of nanostructural intracellular changes, which is critical for understanding the mechanisms of diseases such as cancer. To demonstrate the capabilities of this multimodal instrument, we imaged HeLa cells treated with valinomycin, a potassium ionophore that uncouples oxidative phosphorylation. Colocalization of fluorescence images of the nuclei (Hoechst 33342) and mitochondria (anti‐mitochondria conjugated to Alexa Fluor 488) with PWS measurements allowed us to detect a significant decrease in nuclear nanoscale heterogeneity (Σ), while no significant change in Σ was observed at mitochondrial sites. In addition, application of the new multimodal imaging approach was demonstrated on human buccal samples prepared using a cancer screening protocol. These images demonstrate that nanoscale intracellular structure can be studied in healthy and diseased cells at molecular‐specific sites. Instrumentation has been developed enabling colocalization of confocal fluorescence and nanoscale‐sensitive spectroscopic microscopy modalities. This combination allows sensing of the heterogeneity of nanoscale intracellular structure within specific fluorescently labeled organelles or molecular structures. To demonstrate the capabilities of this multimodal instrument, the effects on the heterogeneity of nanoscale structure were examined in the nuclei and mitochondria of HeLa cells treated with valinomycin, a potassium ionophore that uncouples oxidative phosphorylation.
AbstractList	A new multimodal confocal microscope has been developed, which includes a parallel Partial Wave Spectroscopic (PWS) microscopy path. This combination of modalities allows molecular‐specific sensing of nanoscale intracellular structure using fluorescent labels. Combining molecular specificity and sensitivity to nanoscale structure allows localization of nanostructural intracellular changes, which is critical for understanding the mechanisms of diseases such as cancer. To demonstrate the capabilities of this multimodal instrument, we imaged HeLa cells treated with valinomycin, a potassium ionophore that uncouples oxidative phosphorylation. Colocalization of fluorescence images of the nuclei (Hoechst 33342) and mitochondria (anti‐mitochondria conjugated to Alexa Fluor 488) with PWS measurements allowed us to detect a significant decrease in nuclear nanoscale heterogeneity (Σ), while no significant change in Σ was observed at mitochondrial sites. In addition, application of the new multimodal imaging approach was demonstrated on human buccal samples prepared using a cancer screening protocol. These images demonstrate that nanoscale intracellular structure can be studied in healthy and diseased cells at molecular‐specific sites. Instrumentation has been developed enabling colocalization of confocal fluorescence and nanoscale‐sensitive spectroscopic microscopy modalities. This combination allows sensing of the heterogeneity of nanoscale intracellular structure within specific fluorescently labeled organelles or molecular structures. To demonstrate the capabilities of this multimodal instrument, the effects on the heterogeneity of nanoscale structure were examined in the nuclei and mitochondria of HeLa cells treated with valinomycin, a potassium ionophore that uncouples oxidative phosphorylation. A new multimodal confocal microscope has been developed, which includes a parallel Partial Wave Spectroscopic (PWS) microscopy path. This combination of modalities allows molecular-specific sensing of nanoscale intracellular structure using fluorescent labels. Combining molecular specificity and sensitivity to nanoscale structure allows localization of nanostructural intracellular changes, which is critical for understanding the mechanisms of diseases such as cancer. To demonstrate the capabilities of this multimodal instrument, we imaged HeLa cells treated with valinomycin, a potassium ionophore that uncouples oxidative phosphorylation. Colocalization of fluorescence images of the nuclei (Hoechst 33342) and mitochondria (anti-mitochondria conjugated to Alexa Fluor 488) with PWS measurements allowed us to detect a significant decrease in nuclear nanoscale heterogeneity (Σ), while no significant change in Σ was observed at mitochondrial sites. In addition, application of the new multimodal imaging approach was demonstrated on human buccal samples prepared using a cancer screening protocol. These images demonstrate that nanoscale intracellular structure can be studied in healthy and diseased cells at molecular-specific sites. A new multimodal confocal microscope has been developed, which includes a parallel Partial Wave Spectroscopic (PWS) microscopy path. This combination of modalities allows molecular-specific sensing of nanoscale intracellular structure using fluorescent labels. Combining molecular specificity and sensitivity to nanoscale structure allows localization of nanostructural intracellular changes, which is critical for understanding the mechanisms of diseases such as cancer. To demonstrate the capabilities of this multimodal instrument, we imaged HeLa cells treated with valinomycin, a potassium ionophore that uncouples oxidative phosphorylation. Colocalization of fluorescence images of the nuclei (Hoechst 33342) and mitochondria (anti-mitochondria conjugated to Alexa Fluor 488) with PWS measurements allowed us to detect a significant decrease in nuclear nanoscale heterogeneity ( capital sigma ), while no significant change in capital sigma was observed at mitochondrial sites. In addition, application of the new multimodal imaging approach was demonstrated on human buccal samples prepared using a cancer screening protocol. These images demonstrate that nanoscale intracellular structure can be studied in healthy and diseased cells at molecular-specific sites. Instrumentation has been developed enabling colocalization of confocal fluorescence and nanoscale-sensitive spectroscopic microscopy modalities. This combination allows sensing of the heterogeneity of nanoscale intracellular structure within specific fluorescently labeled organelles or molecular structures. To demonstrate the capabilities of this multimodal instrument, the effects on the heterogeneity of nanoscale structure were examined in the nuclei and mitochondria of HeLa cells treated with valinomycin, a potassium ionophore that uncouples oxidative phosphorylation. A new multimodal confocal microscope has been developed, which includes a parallel Partial Wave Spectroscopic (PWS) microscopy path. This combination of modalities allows molecular-specific sensing of nanoscale intracellular structure using fluorescent labels. Combining molecular specificity and sensitivity to nanoscale structure allows localization of nanostructural intracellular changes, which is critical for understanding the mechanisms of diseases such as cancer. To demonstrate the capabilities of this multimodal instrument, we imaged HeLa cells treated with valinomycin, a potassium ionophore that uncouples oxidative phosphorylation. Colocalization of fluorescence images of the nuclei (Hoechst 33342) and mitochondria (anti-mitochondria conjugated to Alexa Fluor 488) with PWS measurements allowed us to detect a significant decrease in nuclear nanoscale heterogeneity (Σ), while no significant change in Σ was observed at mitochondrial sites. In addition, application of the new multimodal imaging approach was demonstrated on human buccal samples prepared using a cancer screening protocol. These images demonstrate that nanoscale intracellular structure can be studied in healthy and diseased cells at molecular-specific sites.A new multimodal confocal microscope has been developed, which includes a parallel Partial Wave Spectroscopic (PWS) microscopy path. This combination of modalities allows molecular-specific sensing of nanoscale intracellular structure using fluorescent labels. Combining molecular specificity and sensitivity to nanoscale structure allows localization of nanostructural intracellular changes, which is critical for understanding the mechanisms of diseases such as cancer. To demonstrate the capabilities of this multimodal instrument, we imaged HeLa cells treated with valinomycin, a potassium ionophore that uncouples oxidative phosphorylation. Colocalization of fluorescence images of the nuclei (Hoechst 33342) and mitochondria (anti-mitochondria conjugated to Alexa Fluor 488) with PWS measurements allowed us to detect a significant decrease in nuclear nanoscale heterogeneity (Σ), while no significant change in Σ was observed at mitochondrial sites. In addition, application of the new multimodal imaging approach was demonstrated on human buccal samples prepared using a cancer screening protocol. These images demonstrate that nanoscale intracellular structure can be studied in healthy and diseased cells at molecular-specific sites. A new multimodal confocal microscope has been developed, which includes a parallel Partial Wave Spectroscopic (PWS) microscopy path. This combination of modalities allows molecular‐specific sensing of nanoscale intracellular structure using fluorescent labels. Combining molecular specificity and sensitivity to nanoscale structure allows localization of nanostructural intracellular changes, which is critical for understanding the mechanisms of diseases such as cancer. To demonstrate the capabilities of this multimodal instrument, we imaged HeLa cells treated with valinomycin, a potassium ionophore that uncouples oxidative phosphorylation. Colocalization of fluorescence images of the nuclei (Hoechst 33342) and mitochondria (anti‐mitochondria conjugated to Alexa Fluor 488) with PWS measurements allowed us to detect a significant decrease in nuclear nanoscale heterogeneity ( Σ ), while no significant change in Σ was observed at mitochondrial sites. In addition, application of the new multimodal imaging approach was demonstrated on human buccal samples prepared using a cancer screening protocol. These images demonstrate that nanoscale intracellular structure can be studied in healthy and diseased cells at molecular‐specific sites. magnified image
Author	Chandler, John E. Stypula‐Cyrus, Yolanda Almassalha, Luay Bauer, Greta Szleifer, Igal Bowen, Leah Subramanian, Hariharan Backman, Vadim
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Cites_doi	10.1117/1.3523618 10.1186/1471-2407-14-189 10.1002/1097-0142(195309)6:5<963::AID-CNCR2820060515>3.0.CO;2-Q 10.1371/journal.pone.0115999 10.1158/0008-5472.CAN-10-1686 10.1073/pnas.0804723105 10.1364/OL.39.004290 10.1007/s10495-011-0642-9 10.1038/nrm3382 10.1038/nmeth.2089 10.1002/ijc.28122 10.1016/j.bpj.2010.05.023 10.1364/OL.34.000518 10.1038/sj.bmt.1701815 10.1016/j.cell.2012.02.035 10.1038/sj.embor.7400488 10.1103/PhysRevLett.111.033903 10.1016/j.mrrev.2003.06.008 10.1158/0008-5472.CAN-08-3895 10.7150/jca.5838 10.1371/journal.pone.0064600
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Snippet	A new multimodal confocal microscope has been developed, which includes a parallel Partial Wave Spectroscopic (PWS) microscopy path. This combination of...
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SubjectTerms	Cancer Cancer screening Cell Nucleus - drug effects Cell Nucleus - ultrastructure Cheek chromatin Confocal confocal microscopy Fluorescence HeLa Cells Heterogeneity Humans hyperspectral microscopy Image Processing, Computer-Assisted Intracellular Ionophores - pharmacology Localization Medical imaging Medical screening Microscopy, Confocal - instrumentation Microscopy, Confocal - methods Microscopy, Fluorescence - instrumentation Microscopy, Fluorescence - methods Mitochondria Mitochondria - drug effects Mitochondria - ultrastructure Molecular structure Multimodal Imaging nanocytology Nanostructure Oxidative phosphorylation Phosphorylation Spectroscopy Spectrum Analysis - instrumentation Spectrum Analysis - methods Valinomycin Valinomycin - pharmacology
Title	Colocalization of cellular nanostructure using confocal fluorescence and partial wave spectroscopy
URI	https://onlinelibrary.wiley.com/doi/abs/10.1002%2Fjbio.201500298 https://www.ncbi.nlm.nih.gov/pubmed/27111884 https://www.proquest.com/docview/1882078133 https://www.proquest.com/docview/2788254630 https://www.proquest.com/docview/1826669073 https://www.proquest.com/docview/1891870265 https://www.proquest.com/docview/1893888349 https://pubmed.ncbi.nlm.nih.gov/PMC5112146
Volume	10
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