Super‐Resolution Microscopy Using a Bioorthogonal‐Based Cholesterol Probe Provides Unprecedented Capabilities for Imaging Nanoscale Lipid Heterogeneity in Living Cells
Despite more than 20 years of work since the lipid raft concept was proposed, the existence of these nanostructures remains highly controversial due to the lack of noninvasive methods to investigate their native nanorganization in living unperturbed cells. There is an unmet need for probes for direc...
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| Veröffentlicht in: | Small methods 2021-09, Vol.5 (9), p.e2100430-n/a |
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| creator | Lorizate, Maier Terrones, Oihana Nieto‐Garai, Jon Ander Rojo‐Bartolomé, Iratxe Ciceri, Dalila Morana, Ornella Olazar‐Intxausti, June Arboleya, Aroa Martin, Alexia Szynkiewicz, Marta Calleja‐Felipe, Maria Bernardino de la Serna, Jorge Contreras, F.‐Xabier |
| description | Despite more than 20 years of work since the lipid raft concept was proposed, the existence of these nanostructures remains highly controversial due to the lack of noninvasive methods to investigate their native nanorganization in living unperturbed cells. There is an unmet need for probes for direct imaging of nanoscale membrane dynamics with high spatial and temporal resolution in living cells. In this paper, a bioorthogonal‐based cholesterol probe (chol‐N3) is developed that, combined with nanoscopy, becomes a new powerful method for direct visualization and characterization of lipid raft at unprecedented resolution in living cells. The chol‐N3 probe mimics cholesterol in synthetic and cellular membranes without perturbation. When combined with live‐cell super‐resolution microscopy, chol‐N3 demonstrates the existence of cholesterol‐rich nanodomains of |
| doi_str_mv | 10.1002/smtd.202100430 |
| format | Article |
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Direct imaging and characterization of nanoscale lipid heterogeneity in living cells with high temporal and spatial resolution using developed bioorthogonal cholesterol probe and nanoscopy are introduced. This probe enables tackling biological questions that were previously beyond reach. Visualization of cholesterol distribution through thick tissues outlines the tool's potential to investigate the role of cholesterol in health and disease.</description><identifier>ISSN: 2366-9608</identifier><identifier>EISSN: 2366-9608</identifier><identifier>DOI: 10.1002/smtd.202100430</identifier><identifier>PMID: 34928061</identifier><language>eng</language><publisher>Germany</publisher><subject>Animals ; bioorthogonal reactions ; Cells, Cultured ; cholesterol ; Cholesterol - analysis ; Cholesterol - chemistry ; HeLa Cells ; Humans ; lipid raft ; Membrane Microdomains - chemistry ; membranes ; Microscopy, Fluorescence ; Models, Molecular ; Molecular Conformation ; Molecular Imaging - methods ; Molecular Probes - chemistry ; nanoprobes ; nanoscale lipid heterogeneity ; Neurons - chemistry ; Neurons - cytology ; Rats ; Spatio-Temporal Analysis ; super‐resolution microscopy</subject><ispartof>Small methods, 2021-09, Vol.5 (9), p.e2100430-n/a</ispartof><rights>2021 The Authors. Small Methods published by Wiley‐VCH GmbH</rights><rights>2021 The Authors. Small Methods published by Wiley-VCH GmbH.</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c3850-84ca59f022f5bcb8878768b2e04b19287deaec67e64276058067a5172d9d7cff3</citedby><cites>FETCH-LOGICAL-c3850-84ca59f022f5bcb8878768b2e04b19287deaec67e64276058067a5172d9d7cff3</cites><orcidid>0000-0003-0875-634X</orcidid></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://onlinelibrary.wiley.com/doi/pdf/10.1002%2Fsmtd.202100430$$EPDF$$P50$$Gwiley$$Hfree_for_read</linktopdf><linktohtml>$$Uhttps://onlinelibrary.wiley.com/doi/full/10.1002%2Fsmtd.202100430$$EHTML$$P50$$Gwiley$$Hfree_for_read</linktohtml><link.rule.ids>314,777,781,1412,27905,27906,45555,45556</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/34928061$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Lorizate, Maier</creatorcontrib><creatorcontrib>Terrones, Oihana</creatorcontrib><creatorcontrib>Nieto‐Garai, Jon Ander</creatorcontrib><creatorcontrib>Rojo‐Bartolomé, Iratxe</creatorcontrib><creatorcontrib>Ciceri, Dalila</creatorcontrib><creatorcontrib>Morana, Ornella</creatorcontrib><creatorcontrib>Olazar‐Intxausti, June</creatorcontrib><creatorcontrib>Arboleya, Aroa</creatorcontrib><creatorcontrib>Martin, Alexia</creatorcontrib><creatorcontrib>Szynkiewicz, Marta</creatorcontrib><creatorcontrib>Calleja‐Felipe, Maria</creatorcontrib><creatorcontrib>Bernardino de la Serna, Jorge</creatorcontrib><creatorcontrib>Contreras, F.‐Xabier</creatorcontrib><title>Super‐Resolution Microscopy Using a Bioorthogonal‐Based Cholesterol Probe Provides Unprecedented Capabilities for Imaging Nanoscale Lipid Heterogeneity in Living Cells</title><title>Small methods</title><addtitle>Small Methods</addtitle><description>Despite more than 20 years of work since the lipid raft concept was proposed, the existence of these nanostructures remains highly controversial due to the lack of noninvasive methods to investigate their native nanorganization in living unperturbed cells. There is an unmet need for probes for direct imaging of nanoscale membrane dynamics with high spatial and temporal resolution in living cells. In this paper, a bioorthogonal‐based cholesterol probe (chol‐N3) is developed that, combined with nanoscopy, becomes a new powerful method for direct visualization and characterization of lipid raft at unprecedented resolution in living cells. The chol‐N3 probe mimics cholesterol in synthetic and cellular membranes without perturbation. When combined with live‐cell super‐resolution microscopy, chol‐N3 demonstrates the existence of cholesterol‐rich nanodomains of <50 nm at the plasma membrane of resting living cells. Using this tool, the lipid membrane structure of such subdiffraction limit domains is identified, and the nanoscale spatiotemporal organization of cholesterol in the plasma membrane of living cells reveals multiple cholesterol diffusion modes at different spatial localizations. Finally, imaging across thick organ samples outlines the potential of this new method to address essential biological questions that were previously beyond reach.
Direct imaging and characterization of nanoscale lipid heterogeneity in living cells with high temporal and spatial resolution using developed bioorthogonal cholesterol probe and nanoscopy are introduced. This probe enables tackling biological questions that were previously beyond reach. Visualization of cholesterol distribution through thick tissues outlines the tool's potential to investigate the role of cholesterol in health and disease.</description><subject>Animals</subject><subject>bioorthogonal reactions</subject><subject>Cells, Cultured</subject><subject>cholesterol</subject><subject>Cholesterol - analysis</subject><subject>Cholesterol - chemistry</subject><subject>HeLa Cells</subject><subject>Humans</subject><subject>lipid raft</subject><subject>Membrane Microdomains - chemistry</subject><subject>membranes</subject><subject>Microscopy, Fluorescence</subject><subject>Models, Molecular</subject><subject>Molecular Conformation</subject><subject>Molecular Imaging - methods</subject><subject>Molecular Probes - chemistry</subject><subject>nanoprobes</subject><subject>nanoscale lipid heterogeneity</subject><subject>Neurons - chemistry</subject><subject>Neurons - cytology</subject><subject>Rats</subject><subject>Spatio-Temporal Analysis</subject><subject>super‐resolution microscopy</subject><issn>2366-9608</issn><issn>2366-9608</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2021</creationdate><recordtype>article</recordtype><sourceid>24P</sourceid><sourceid>WIN</sourceid><sourceid>EIF</sourceid><recordid>eNqFkUtu2zAQhomiQWM42XZZcJmNXZKSKGkZu3kBTls08VqgyJHDgCIVUnLgXY6Qe_RWPUkpOE2y64aP4Tc_Z-ZH6DMlc0oI-xraXs0ZYfGSJuQDmrCE81nJSfHx3fkQHYdwT2ICoUnG6Cd0mKQlKwinE_T7ZujA_3l6_gXBmaHXzuJrLb0L0nU7vA7abrDAC-2c7-_cxllhIr0QARRe3jkDoQfvDP7pXQ3jutUKAl7bzoMEBbYfQdGJWhvd6_jUOI-vWrEZlb8LG38SBvBKd1rhSxjVNmBB9zusbQxvR24JxoQjdNAIE-D4ZZ-i9fnZ7fJytvpxcbU8Xc1kUmRkVqRSZGVDGGuyWtZFkRc5L2oGJK1p7DtXIEDyHHjKck6yOIhcZDRnqlS5bJpkik72up13D0NssGp1kLECYcENoWKcMpJQnvGIzvfoOLHgoak6r1vhdxUl1ehRNXpUvXoUE768aA91C-oV_-dIBMo98KgN7P4jV91c3357E_8L6B6jSA</recordid><startdate>20210901</startdate><enddate>20210901</enddate><creator>Lorizate, Maier</creator><creator>Terrones, Oihana</creator><creator>Nieto‐Garai, Jon Ander</creator><creator>Rojo‐Bartolomé, Iratxe</creator><creator>Ciceri, Dalila</creator><creator>Morana, Ornella</creator><creator>Olazar‐Intxausti, June</creator><creator>Arboleya, Aroa</creator><creator>Martin, Alexia</creator><creator>Szynkiewicz, Marta</creator><creator>Calleja‐Felipe, Maria</creator><creator>Bernardino de la Serna, Jorge</creator><creator>Contreras, F.‐Xabier</creator><scope>24P</scope><scope>WIN</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7X8</scope><orcidid>https://orcid.org/0000-0003-0875-634X</orcidid></search><sort><creationdate>20210901</creationdate><title>Super‐Resolution Microscopy Using a Bioorthogonal‐Based Cholesterol Probe Provides Unprecedented Capabilities for Imaging Nanoscale Lipid Heterogeneity in Living Cells</title><author>Lorizate, Maier ; Terrones, Oihana ; Nieto‐Garai, Jon Ander ; Rojo‐Bartolomé, Iratxe ; Ciceri, Dalila ; Morana, Ornella ; Olazar‐Intxausti, June ; Arboleya, Aroa ; Martin, Alexia ; Szynkiewicz, Marta ; Calleja‐Felipe, Maria ; Bernardino de la Serna, Jorge ; Contreras, F.‐Xabier</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c3850-84ca59f022f5bcb8878768b2e04b19287deaec67e64276058067a5172d9d7cff3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2021</creationdate><topic>Animals</topic><topic>bioorthogonal reactions</topic><topic>Cells, Cultured</topic><topic>cholesterol</topic><topic>Cholesterol - analysis</topic><topic>Cholesterol - chemistry</topic><topic>HeLa Cells</topic><topic>Humans</topic><topic>lipid raft</topic><topic>Membrane Microdomains - chemistry</topic><topic>membranes</topic><topic>Microscopy, Fluorescence</topic><topic>Models, Molecular</topic><topic>Molecular Conformation</topic><topic>Molecular Imaging - methods</topic><topic>Molecular Probes - chemistry</topic><topic>nanoprobes</topic><topic>nanoscale lipid heterogeneity</topic><topic>Neurons - chemistry</topic><topic>Neurons - cytology</topic><topic>Rats</topic><topic>Spatio-Temporal Analysis</topic><topic>super‐resolution microscopy</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Lorizate, Maier</creatorcontrib><creatorcontrib>Terrones, Oihana</creatorcontrib><creatorcontrib>Nieto‐Garai, Jon Ander</creatorcontrib><creatorcontrib>Rojo‐Bartolomé, Iratxe</creatorcontrib><creatorcontrib>Ciceri, Dalila</creatorcontrib><creatorcontrib>Morana, Ornella</creatorcontrib><creatorcontrib>Olazar‐Intxausti, June</creatorcontrib><creatorcontrib>Arboleya, Aroa</creatorcontrib><creatorcontrib>Martin, Alexia</creatorcontrib><creatorcontrib>Szynkiewicz, Marta</creatorcontrib><creatorcontrib>Calleja‐Felipe, Maria</creatorcontrib><creatorcontrib>Bernardino de la Serna, Jorge</creatorcontrib><creatorcontrib>Contreras, F.‐Xabier</creatorcontrib><collection>Wiley Online Library Open Access</collection><collection>Wiley Online Library Free Content</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><jtitle>Small methods</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Lorizate, Maier</au><au>Terrones, Oihana</au><au>Nieto‐Garai, Jon Ander</au><au>Rojo‐Bartolomé, Iratxe</au><au>Ciceri, Dalila</au><au>Morana, Ornella</au><au>Olazar‐Intxausti, June</au><au>Arboleya, Aroa</au><au>Martin, Alexia</au><au>Szynkiewicz, Marta</au><au>Calleja‐Felipe, Maria</au><au>Bernardino de la Serna, Jorge</au><au>Contreras, F.‐Xabier</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Super‐Resolution Microscopy Using a Bioorthogonal‐Based Cholesterol Probe Provides Unprecedented Capabilities for Imaging Nanoscale Lipid Heterogeneity in Living Cells</atitle><jtitle>Small methods</jtitle><addtitle>Small Methods</addtitle><date>2021-09-01</date><risdate>2021</risdate><volume>5</volume><issue>9</issue><spage>e2100430</spage><epage>n/a</epage><pages>e2100430-n/a</pages><issn>2366-9608</issn><eissn>2366-9608</eissn><abstract>Despite more than 20 years of work since the lipid raft concept was proposed, the existence of these nanostructures remains highly controversial due to the lack of noninvasive methods to investigate their native nanorganization in living unperturbed cells. There is an unmet need for probes for direct imaging of nanoscale membrane dynamics with high spatial and temporal resolution in living cells. In this paper, a bioorthogonal‐based cholesterol probe (chol‐N3) is developed that, combined with nanoscopy, becomes a new powerful method for direct visualization and characterization of lipid raft at unprecedented resolution in living cells. The chol‐N3 probe mimics cholesterol in synthetic and cellular membranes without perturbation. When combined with live‐cell super‐resolution microscopy, chol‐N3 demonstrates the existence of cholesterol‐rich nanodomains of <50 nm at the plasma membrane of resting living cells. Using this tool, the lipid membrane structure of such subdiffraction limit domains is identified, and the nanoscale spatiotemporal organization of cholesterol in the plasma membrane of living cells reveals multiple cholesterol diffusion modes at different spatial localizations. Finally, imaging across thick organ samples outlines the potential of this new method to address essential biological questions that were previously beyond reach.
Direct imaging and characterization of nanoscale lipid heterogeneity in living cells with high temporal and spatial resolution using developed bioorthogonal cholesterol probe and nanoscopy are introduced. This probe enables tackling biological questions that were previously beyond reach. Visualization of cholesterol distribution through thick tissues outlines the tool's potential to investigate the role of cholesterol in health and disease.</abstract><cop>Germany</cop><pmid>34928061</pmid><doi>10.1002/smtd.202100430</doi><tpages>15</tpages><orcidid>https://orcid.org/0000-0003-0875-634X</orcidid><oa>free_for_read</oa></addata></record> |
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| subjects | Animals bioorthogonal reactions Cells, Cultured cholesterol Cholesterol - analysis Cholesterol - chemistry HeLa Cells Humans lipid raft Membrane Microdomains - chemistry membranes Microscopy, Fluorescence Models, Molecular Molecular Conformation Molecular Imaging - methods Molecular Probes - chemistry nanoprobes nanoscale lipid heterogeneity Neurons - chemistry Neurons - cytology Rats Spatio-Temporal Analysis super‐resolution microscopy |
| title | Super‐Resolution Microscopy Using a Bioorthogonal‐Based Cholesterol Probe Provides Unprecedented Capabilities for Imaging Nanoscale Lipid Heterogeneity in Living Cells |
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