Improvement of immunodetection of the transcription factor C/EBP homologous protein by western blot
Accumulation of misfolded proteins within the endoplasmic reticulum (ER) induces an unfolded protein response (UPR) that either restores homeostasis or triggers apoptosis in case of adaptation failure. The three activated branches of UPR lead to IRE1-, PERK- and ATF6- dependent transcriptional induc...
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| Veröffentlicht in: | Analytical biochemistry 2020-07, Vol.601, p.113775-113775, Article 113775 |
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| container_title | Analytical biochemistry |
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| creator | Turpin, Jonathan Frumence, Etienne El Safadi, Daed Meilhac, Olivier Krejbich-Trotot, Pascale Viranaïcken, Wildriss |
| description | Accumulation of misfolded proteins within the endoplasmic reticulum (ER) induces an unfolded protein response (UPR) that either restores homeostasis or triggers apoptosis in case of adaptation failure. The three activated branches of UPR lead to IRE1-, PERK- and ATF6- dependent transcriptional induction of the gene encoding the transcription factor C/EBP homologous protein (CHOP) which plays an important role in apoptosis induction. In conventional immunoblotting conditions, detection of CHOP is a difficult task. Using a fixation step, we have optimized the detection of CHOP and this method provides a valuable tool to decipher CHOP involvement in UPR.
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•Low retention of CHOP protein on nitrocellulose membrane during western blot.•Glutaraldehyde (GA) fixation increase reactivity to CHOP in western blot.•GA fixation decrease background.•GA fixation decrease non specific bands. |
| doi_str_mv | 10.1016/j.ab.2020.113775 |
| format | Article |
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[Display omitted]
•Low retention of CHOP protein on nitrocellulose membrane during western blot.•Glutaraldehyde (GA) fixation increase reactivity to CHOP in western blot.•GA fixation decrease background.•GA fixation decrease non specific bands.</description><identifier>ISSN: 0003-2697</identifier><identifier>EISSN: 1096-0309</identifier><identifier>DOI: 10.1016/j.ab.2020.113775</identifier><identifier>PMID: 32416096</identifier><language>eng</language><publisher>United States: Elsevier Inc</publisher><subject>A549 Cells ; Blotting, Western ; CHOP ; Endoplasmic Reticulum ; Endoplasmic Reticulum - chemistry ; Humans ; Immunodetection ; Life Sciences ; Microbiology and Parasitology ; Nitrocellulose ; Transcription Factor CHOP ; Transcription Factor CHOP - analysis ; Unfolded Protein Response ; Western blot</subject><ispartof>Analytical biochemistry, 2020-07, Vol.601, p.113775-113775, Article 113775</ispartof><rights>2020 Elsevier Inc.</rights><rights>Copyright © 2020 Elsevier Inc. All rights reserved.</rights><rights>Attribution - NonCommercial</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c492t-ef711af23c77749470b625f09f838f4ca15773382a79de04095ef90c41e83c753</citedby><cites>FETCH-LOGICAL-c492t-ef711af23c77749470b625f09f838f4ca15773382a79de04095ef90c41e83c753</cites><orcidid>0000-0002-0915-8635 ; 0000-0003-2667-345X ; 0000-0001-7329-1716 ; 0000-0002-3740-7539 ; 0000-0002-7734-7737</orcidid></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktohtml>$$Uhttps://www.sciencedirect.com/science/article/pii/S0003269720303079$$EHTML$$P50$$Gelsevier$$H</linktohtml><link.rule.ids>230,314,776,780,881,3537,27901,27902,65306</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/32416096$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink><backlink>$$Uhttps://hal.univ-reunion.fr/hal-03120150$$DView record in HAL$$Hfree_for_read</backlink></links><search><creatorcontrib>Turpin, Jonathan</creatorcontrib><creatorcontrib>Frumence, Etienne</creatorcontrib><creatorcontrib>El Safadi, Daed</creatorcontrib><creatorcontrib>Meilhac, Olivier</creatorcontrib><creatorcontrib>Krejbich-Trotot, Pascale</creatorcontrib><creatorcontrib>Viranaïcken, Wildriss</creatorcontrib><title>Improvement of immunodetection of the transcription factor C/EBP homologous protein by western blot</title><title>Analytical biochemistry</title><addtitle>Anal Biochem</addtitle><description>Accumulation of misfolded proteins within the endoplasmic reticulum (ER) induces an unfolded protein response (UPR) that either restores homeostasis or triggers apoptosis in case of adaptation failure. The three activated branches of UPR lead to IRE1-, PERK- and ATF6- dependent transcriptional induction of the gene encoding the transcription factor C/EBP homologous protein (CHOP) which plays an important role in apoptosis induction. In conventional immunoblotting conditions, detection of CHOP is a difficult task. Using a fixation step, we have optimized the detection of CHOP and this method provides a valuable tool to decipher CHOP involvement in UPR.
[Display omitted]
•Low retention of CHOP protein on nitrocellulose membrane during western blot.•Glutaraldehyde (GA) fixation increase reactivity to CHOP in western blot.•GA fixation decrease background.•GA fixation decrease non specific bands.</description><subject>A549 Cells</subject><subject>Blotting, Western</subject><subject>CHOP</subject><subject>Endoplasmic Reticulum</subject><subject>Endoplasmic Reticulum - chemistry</subject><subject>Humans</subject><subject>Immunodetection</subject><subject>Life Sciences</subject><subject>Microbiology and Parasitology</subject><subject>Nitrocellulose</subject><subject>Transcription Factor CHOP</subject><subject>Transcription Factor CHOP - analysis</subject><subject>Unfolded Protein Response</subject><subject>Western blot</subject><issn>0003-2697</issn><issn>1096-0309</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2020</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNp1kc1vEzEQxS0EatPSOye0R3rYdPyx65hbiUpbKRIc4Gx5vWPiaHcdbCeo_30dtvTGyfbo996M5xHygcKSAm1vdkvTLRmw8qRcyuYNWVBQbQ0c1FuyAABes1bJc3KR0g6AUtG0Z-ScM0HbAi6IfRz3MRxxxClXwVV-HA9T6DGjzT5Mp1LeYpWjmZKNfv-36IzNIVbrm7sv36ttGMMQfoVDqopTRj9V3VP1B1PGWK5DyO_JO2eGhFcv5yX5-fXux_qh3ny7f1zfbmorFMs1OkmpcYxbKaVQQkLXssaBciu-csIa2kjJ-YoZqXoEAapBp8AKiquiafgluZ59t2bQ--hHE590MF4_3G70qQacMqANHGlhP81smfn3oQyrR58sDoOZsHxFM1EaCFZaFhRm1MaQUkT36k1Bn2LQO206fYpBzzEUyccX90M3Yv8q-Lf3AnyeASz7OHqMOlmPk8Xex7J53Qf_f_dn81OV8A</recordid><startdate>20200715</startdate><enddate>20200715</enddate><creator>Turpin, Jonathan</creator><creator>Frumence, Etienne</creator><creator>El Safadi, Daed</creator><creator>Meilhac, Olivier</creator><creator>Krejbich-Trotot, Pascale</creator><creator>Viranaïcken, Wildriss</creator><general>Elsevier Inc</general><general>Elsevier Masson</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>7X8</scope><scope>1XC</scope><scope>VOOES</scope><orcidid>https://orcid.org/0000-0002-0915-8635</orcidid><orcidid>https://orcid.org/0000-0003-2667-345X</orcidid><orcidid>https://orcid.org/0000-0001-7329-1716</orcidid><orcidid>https://orcid.org/0000-0002-3740-7539</orcidid><orcidid>https://orcid.org/0000-0002-7734-7737</orcidid></search><sort><creationdate>20200715</creationdate><title>Improvement of immunodetection of the transcription factor C/EBP homologous protein by western blot</title><author>Turpin, Jonathan ; Frumence, Etienne ; El Safadi, Daed ; Meilhac, Olivier ; Krejbich-Trotot, Pascale ; Viranaïcken, Wildriss</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c492t-ef711af23c77749470b625f09f838f4ca15773382a79de04095ef90c41e83c753</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2020</creationdate><topic>A549 Cells</topic><topic>Blotting, Western</topic><topic>CHOP</topic><topic>Endoplasmic Reticulum</topic><topic>Endoplasmic Reticulum - chemistry</topic><topic>Humans</topic><topic>Immunodetection</topic><topic>Life Sciences</topic><topic>Microbiology and Parasitology</topic><topic>Nitrocellulose</topic><topic>Transcription Factor CHOP</topic><topic>Transcription Factor CHOP - analysis</topic><topic>Unfolded Protein Response</topic><topic>Western blot</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Turpin, Jonathan</creatorcontrib><creatorcontrib>Frumence, Etienne</creatorcontrib><creatorcontrib>El Safadi, Daed</creatorcontrib><creatorcontrib>Meilhac, Olivier</creatorcontrib><creatorcontrib>Krejbich-Trotot, Pascale</creatorcontrib><creatorcontrib>Viranaïcken, Wildriss</creatorcontrib><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><collection>Hyper Article en Ligne (HAL)</collection><collection>Hyper Article en Ligne (HAL) (Open Access)</collection><jtitle>Analytical biochemistry</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Turpin, Jonathan</au><au>Frumence, Etienne</au><au>El Safadi, Daed</au><au>Meilhac, Olivier</au><au>Krejbich-Trotot, Pascale</au><au>Viranaïcken, Wildriss</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Improvement of immunodetection of the transcription factor C/EBP homologous protein by western blot</atitle><jtitle>Analytical biochemistry</jtitle><addtitle>Anal Biochem</addtitle><date>2020-07-15</date><risdate>2020</risdate><volume>601</volume><spage>113775</spage><epage>113775</epage><pages>113775-113775</pages><artnum>113775</artnum><issn>0003-2697</issn><eissn>1096-0309</eissn><abstract>Accumulation of misfolded proteins within the endoplasmic reticulum (ER) induces an unfolded protein response (UPR) that either restores homeostasis or triggers apoptosis in case of adaptation failure. The three activated branches of UPR lead to IRE1-, PERK- and ATF6- dependent transcriptional induction of the gene encoding the transcription factor C/EBP homologous protein (CHOP) which plays an important role in apoptosis induction. In conventional immunoblotting conditions, detection of CHOP is a difficult task. Using a fixation step, we have optimized the detection of CHOP and this method provides a valuable tool to decipher CHOP involvement in UPR.
[Display omitted]
•Low retention of CHOP protein on nitrocellulose membrane during western blot.•Glutaraldehyde (GA) fixation increase reactivity to CHOP in western blot.•GA fixation decrease background.•GA fixation decrease non specific bands.</abstract><cop>United States</cop><pub>Elsevier Inc</pub><pmid>32416096</pmid><doi>10.1016/j.ab.2020.113775</doi><tpages>1</tpages><orcidid>https://orcid.org/0000-0002-0915-8635</orcidid><orcidid>https://orcid.org/0000-0003-2667-345X</orcidid><orcidid>https://orcid.org/0000-0001-7329-1716</orcidid><orcidid>https://orcid.org/0000-0002-3740-7539</orcidid><orcidid>https://orcid.org/0000-0002-7734-7737</orcidid><oa>free_for_read</oa></addata></record> |
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| subjects | A549 Cells Blotting, Western CHOP Endoplasmic Reticulum Endoplasmic Reticulum - chemistry Humans Immunodetection Life Sciences Microbiology and Parasitology Nitrocellulose Transcription Factor CHOP Transcription Factor CHOP - analysis Unfolded Protein Response Western blot |
| title | Improvement of immunodetection of the transcription factor C/EBP homologous protein by western blot |
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