Simplified method to perform CLARITY imaging
Imaging methods are used widely to understand structure of brain and other biological objects. However, sample penetration by light microscopy is limited due to light scattering by the tissue. A number of methods have been recently developed to solve this problem. In one approach (SeeDB) simple proc...
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| Veröffentlicht in: | Molecular neurodegeneration 2014-05, Vol.9 (1), p.19-19, Article 19 |
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| container_title | Molecular neurodegeneration |
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| creator | Poguzhelskaya, Ekaterina Artamonov, Dmitry Bolshakova, Anastasia Vlasova, Olga Bezprozvanny, Ilya |
| description | Imaging methods are used widely to understand structure of brain and other biological objects. However, sample penetration by light microscopy is limited due to light scattering by the tissue. A number of methods have been recently developed to solve this problem. In one approach (SeeDB) simple procedure for clarifying brain samples for imaging was described. However, this method is not compatible with immunostaining approach as SeeDB-prepared tissue is not permeable to the antibodies. Another technique for clearing brain tissue (CLARITY) was optimized for immunochemistry, but this method technically much more demanding than SeeDB.
Here we report optimized protocol for imaging of brain samples (CLARITY2). We have simplified and shortened the original protocol. Following hydrogel fixation, we cut brain tissue to 1-1.5 mm thick coronal slices. This additional step enabled us to accelerate and simplify clearing, staining and imaging steps when compared to the original protocol. We validated the modified protocol in imaging experiments with brains from line M Thy1-GFP mouse and in immunostaining experiments with antibodies against postsynaptic protein PSD-95 and striatal-specific protein DARPP32.
The original CLARITY protocol was optimized and simplified. Application of the modified CLARITY2 protocol could be useful for a broad range of scientists working in neurobiology and developmental biology. |
| doi_str_mv | 10.1186/1750-1326-9-19 |
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Here we report optimized protocol for imaging of brain samples (CLARITY2). We have simplified and shortened the original protocol. Following hydrogel fixation, we cut brain tissue to 1-1.5 mm thick coronal slices. This additional step enabled us to accelerate and simplify clearing, staining and imaging steps when compared to the original protocol. We validated the modified protocol in imaging experiments with brains from line M Thy1-GFP mouse and in immunostaining experiments with antibodies against postsynaptic protein PSD-95 and striatal-specific protein DARPP32.
The original CLARITY protocol was optimized and simplified. Application of the modified CLARITY2 protocol could be useful for a broad range of scientists working in neurobiology and developmental biology.</description><identifier>ISSN: 1750-1326</identifier><identifier>EISSN: 1750-1326</identifier><identifier>DOI: 10.1186/1750-1326-9-19</identifier><identifier>PMID: 24885504</identifier><language>eng</language><publisher>England: BioMed Central Ltd</publisher><subject>Animals ; Antibodies ; Brain ; Histocytological Preparation Techniques - methods ; Imaging, Three-Dimensional - methods ; Immunohistochemistry ; Methodology ; Methods ; Mice ; Microscopy, Confocal ; Neuroimaging - methods ; Neurons ; Neurophysiology ; Physiological aspects ; Viral antibodies</subject><ispartof>Molecular neurodegeneration, 2014-05, Vol.9 (1), p.19-19, Article 19</ispartof><rights>COPYRIGHT 2014 BioMed Central Ltd.</rights><rights>2014 Poguzhelskaya et al.; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.</rights><rights>Copyright © 2014 Poguzhelskaya et al.; licensee BioMed Central Ltd. 2014 Poguzhelskaya et al.; licensee BioMed Central Ltd.</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-b576t-2fc68f78a21d1553a77cb822ceea8c8498e928d204ac05a069b3423082f4dc2a3</citedby><cites>FETCH-LOGICAL-b576t-2fc68f78a21d1553a77cb822ceea8c8498e928d204ac05a069b3423082f4dc2a3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC4049387/pdf/$$EPDF$$P50$$Gpubmedcentral$$Hfree_for_read</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC4049387/$$EHTML$$P50$$Gpubmedcentral$$Hfree_for_read</linktohtml><link.rule.ids>230,314,723,776,780,860,881,27903,27904,53770,53772</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/24885504$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Poguzhelskaya, Ekaterina</creatorcontrib><creatorcontrib>Artamonov, Dmitry</creatorcontrib><creatorcontrib>Bolshakova, Anastasia</creatorcontrib><creatorcontrib>Vlasova, Olga</creatorcontrib><creatorcontrib>Bezprozvanny, Ilya</creatorcontrib><title>Simplified method to perform CLARITY imaging</title><title>Molecular neurodegeneration</title><addtitle>Mol Neurodegener</addtitle><description>Imaging methods are used widely to understand structure of brain and other biological objects. However, sample penetration by light microscopy is limited due to light scattering by the tissue. A number of methods have been recently developed to solve this problem. In one approach (SeeDB) simple procedure for clarifying brain samples for imaging was described. However, this method is not compatible with immunostaining approach as SeeDB-prepared tissue is not permeable to the antibodies. Another technique for clearing brain tissue (CLARITY) was optimized for immunochemistry, but this method technically much more demanding than SeeDB.
Here we report optimized protocol for imaging of brain samples (CLARITY2). We have simplified and shortened the original protocol. Following hydrogel fixation, we cut brain tissue to 1-1.5 mm thick coronal slices. This additional step enabled us to accelerate and simplify clearing, staining and imaging steps when compared to the original protocol. We validated the modified protocol in imaging experiments with brains from line M Thy1-GFP mouse and in immunostaining experiments with antibodies against postsynaptic protein PSD-95 and striatal-specific protein DARPP32.
The original CLARITY protocol was optimized and simplified. Application of the modified CLARITY2 protocol could be useful for a broad range of scientists working in neurobiology and developmental biology.</description><subject>Animals</subject><subject>Antibodies</subject><subject>Brain</subject><subject>Histocytological Preparation Techniques - methods</subject><subject>Imaging, Three-Dimensional - methods</subject><subject>Immunohistochemistry</subject><subject>Methodology</subject><subject>Methods</subject><subject>Mice</subject><subject>Microscopy, Confocal</subject><subject>Neuroimaging - methods</subject><subject>Neurons</subject><subject>Neurophysiology</subject><subject>Physiological aspects</subject><subject>Viral antibodies</subject><issn>1750-1326</issn><issn>1750-1326</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2014</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><sourceid>ABUWG</sourceid><sourceid>AFKRA</sourceid><sourceid>AZQEC</sourceid><sourceid>BENPR</sourceid><sourceid>CCPQU</sourceid><sourceid>DWQXO</sourceid><recordid>eNqNks9LHDEUx0NpUWu99lgGevHgaH5OXi6FZdG6sFCwevAUMplkjcxMtplZwf_eLOq6Wgslh4T3Pvm-l28eQl8JPiYEqhMiBS4Jo1WpSqI-oL1N4OPWeRd9HoZbjLnEWOygXcoBhMB8Dx39Dt2yDT64pujceBObYozF0iUfU1dM55OL2eV1ETqzCP3iC_rkTTu4g6d9H12dnV5Oz8v5r5-z6WRe1kJWY0m9rcBLMJQ0RAhmpLQ1UGqdM2CBK3CKQkMxNxYLgytVM04ZBup5Y6lh--jHo-5yVXeusa4fk2n1MuU-0r2OJujXmT7c6EW80xxzxUBmgcmjQB3iPwReZ2zs9NotvXZLK01U1jh8aiLFPys3jLoLg3Vta3oXV4MmgmPKJAj6HyjjXDKoIKPf36C3cZX67OaaAg6CSPVCLUzrdOh9zF3ataie5LKVwJKKTB2_Q-XVuC7Y2Dsfcvy9CzbFYUjObxwh-eV5nP724Nv2R2zw5_lhDxkowao</recordid><startdate>20140526</startdate><enddate>20140526</enddate><creator>Poguzhelskaya, Ekaterina</creator><creator>Artamonov, Dmitry</creator><creator>Bolshakova, Anastasia</creator><creator>Vlasova, Olga</creator><creator>Bezprozvanny, Ilya</creator><general>BioMed Central Ltd</general><general>BioMed Central</general><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>3V.</scope><scope>7TK</scope><scope>7X7</scope><scope>7XB</scope><scope>88E</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>ABUWG</scope><scope>AFKRA</scope><scope>AZQEC</scope><scope>BENPR</scope><scope>CCPQU</scope><scope>DWQXO</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>K9.</scope><scope>M0S</scope><scope>M1P</scope><scope>PIMPY</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>PRINS</scope><scope>7X8</scope><scope>5PM</scope></search><sort><creationdate>20140526</creationdate><title>Simplified method to perform CLARITY imaging</title><author>Poguzhelskaya, Ekaterina ; Artamonov, Dmitry ; Bolshakova, Anastasia ; Vlasova, Olga ; Bezprozvanny, Ilya</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-b576t-2fc68f78a21d1553a77cb822ceea8c8498e928d204ac05a069b3423082f4dc2a3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2014</creationdate><topic>Animals</topic><topic>Antibodies</topic><topic>Brain</topic><topic>Histocytological Preparation Techniques - methods</topic><topic>Imaging, Three-Dimensional - methods</topic><topic>Immunohistochemistry</topic><topic>Methodology</topic><topic>Methods</topic><topic>Mice</topic><topic>Microscopy, Confocal</topic><topic>Neuroimaging - methods</topic><topic>Neurons</topic><topic>Neurophysiology</topic><topic>Physiological aspects</topic><topic>Viral antibodies</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Poguzhelskaya, Ekaterina</creatorcontrib><creatorcontrib>Artamonov, Dmitry</creatorcontrib><creatorcontrib>Bolshakova, Anastasia</creatorcontrib><creatorcontrib>Vlasova, Olga</creatorcontrib><creatorcontrib>Bezprozvanny, Ilya</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>ProQuest Central (Corporate)</collection><collection>Neurosciences Abstracts</collection><collection>Health & Medical Collection</collection><collection>ProQuest Central (purchase pre-March 2016)</collection><collection>Medical Database (Alumni Edition)</collection><collection>Hospital Premium Collection</collection><collection>Hospital Premium Collection (Alumni Edition)</collection><collection>ProQuest Central (Alumni) (purchase pre-March 2016)</collection><collection>ProQuest Central (Alumni Edition)</collection><collection>ProQuest Central UK/Ireland</collection><collection>ProQuest Central Essentials</collection><collection>ProQuest Central</collection><collection>ProQuest One Community College</collection><collection>ProQuest Central Korea</collection><collection>Health Research Premium Collection</collection><collection>Health Research Premium Collection (Alumni)</collection><collection>ProQuest Health & Medical Complete (Alumni)</collection><collection>Health & Medical Collection (Alumni Edition)</collection><collection>Medical Database</collection><collection>Publicly Available Content Database</collection><collection>ProQuest One Academic Eastern Edition (DO NOT USE)</collection><collection>ProQuest One Academic</collection><collection>ProQuest One Academic UKI Edition</collection><collection>ProQuest Central China</collection><collection>MEDLINE - Academic</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>Molecular neurodegeneration</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Poguzhelskaya, Ekaterina</au><au>Artamonov, Dmitry</au><au>Bolshakova, Anastasia</au><au>Vlasova, Olga</au><au>Bezprozvanny, Ilya</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Simplified method to perform CLARITY imaging</atitle><jtitle>Molecular neurodegeneration</jtitle><addtitle>Mol Neurodegener</addtitle><date>2014-05-26</date><risdate>2014</risdate><volume>9</volume><issue>1</issue><spage>19</spage><epage>19</epage><pages>19-19</pages><artnum>19</artnum><issn>1750-1326</issn><eissn>1750-1326</eissn><abstract>Imaging methods are used widely to understand structure of brain and other biological objects. However, sample penetration by light microscopy is limited due to light scattering by the tissue. A number of methods have been recently developed to solve this problem. In one approach (SeeDB) simple procedure for clarifying brain samples for imaging was described. However, this method is not compatible with immunostaining approach as SeeDB-prepared tissue is not permeable to the antibodies. Another technique for clearing brain tissue (CLARITY) was optimized for immunochemistry, but this method technically much more demanding than SeeDB.
Here we report optimized protocol for imaging of brain samples (CLARITY2). We have simplified and shortened the original protocol. Following hydrogel fixation, we cut brain tissue to 1-1.5 mm thick coronal slices. This additional step enabled us to accelerate and simplify clearing, staining and imaging steps when compared to the original protocol. We validated the modified protocol in imaging experiments with brains from line M Thy1-GFP mouse and in immunostaining experiments with antibodies against postsynaptic protein PSD-95 and striatal-specific protein DARPP32.
The original CLARITY protocol was optimized and simplified. Application of the modified CLARITY2 protocol could be useful for a broad range of scientists working in neurobiology and developmental biology.</abstract><cop>England</cop><pub>BioMed Central Ltd</pub><pmid>24885504</pmid><doi>10.1186/1750-1326-9-19</doi><tpages>1</tpages><oa>free_for_read</oa></addata></record> |
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| subjects | Animals Antibodies Brain Histocytological Preparation Techniques - methods Imaging, Three-Dimensional - methods Immunohistochemistry Methodology Methods Mice Microscopy, Confocal Neuroimaging - methods Neurons Neurophysiology Physiological aspects Viral antibodies |
| title | Simplified method to perform CLARITY imaging |
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