Comparative study of commercially available and homemade anti-VAMP7 antibodies using CRISPR/Cas9-depleted HeLa cells and VAMP7 knockout mice [version 2; referees: 2 approved]
VAMP7 (vesicle-associated membrane protein) belongs to the intracellular membrane fusion SNARE (Soluble N-ethylmaleimide-sensitive factor attachment protein receptors) protein family. In this study, we used CRISPR/Cas9 genome editing technology to generate VAMP7 knockout (KO) human HeLa cells and mo...
| Main Authors: | , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
F1000 Research Ltd
2019-02-01
|
| Series: | F1000Research |
| Online Access: | https://f1000research.com/articles/7-1649/v2 |
| _version_ | 1818959600090611712 |
|---|---|
| author | Agathe Verraes Beatrice Cholley Thierry Galli Sebastien Nola |
| author_facet | Agathe Verraes Beatrice Cholley Thierry Galli Sebastien Nola |
| author_sort | Agathe Verraes |
| collection | DOAJ |
| description | VAMP7 (vesicle-associated membrane protein) belongs to the intracellular membrane fusion SNARE (Soluble N-ethylmaleimide-sensitive factor attachment protein receptors) protein family. In this study, we used CRISPR/Cas9 genome editing technology to generate VAMP7 knockout (KO) human HeLa cells and mouse KO brain extracts in order to test the specificity and the background of a set of commercially available and homemade anti-VAMP7 antibodies. We propose a simple profiling method to analyze western blotting and use visual scoring for immunocytochemistry staining to determine the extent of the antibodies’ specificity. Thus, we were able to rank the performance of a set of available antibodies and further showed an optimized procedure for VAMP7 immunoprecipitation, which we validated using wild-type and KO mouse brain extracts. |
| first_indexed | 2024-12-20T11:44:13Z |
| format | Article |
| id | doaj.art-13650a3d73954407a49ae306dc8c3c83 |
| institution | Directory Open Access Journal |
| issn | 2046-1402 |
| language | English |
| last_indexed | 2024-12-20T11:44:13Z |
| publishDate | 2019-02-01 |
| publisher | F1000 Research Ltd |
| record_format | Article |
| series | F1000Research |
| spelling | doaj.art-13650a3d73954407a49ae306dc8c3c832022-12-21T19:41:54ZengF1000 Research LtdF1000Research2046-14022019-02-01710.12688/f1000research.15707.219822Comparative study of commercially available and homemade anti-VAMP7 antibodies using CRISPR/Cas9-depleted HeLa cells and VAMP7 knockout mice [version 2; referees: 2 approved]Agathe Verraes0Beatrice Cholley1Thierry Galli2Sebastien Nola3Membrane Traffic in Health and Disease, INSERM ERL U950, Univ Paris Diderot, Sorbonne Paris Cité, Institut Jacques Monod, CNRS UMR 7592, Paris, France, 75013, FranceMembrane Traffic in Healthy & Diseased Brain, Institute of Psychiatry and Neuroscience of Paris, INSERM U1266, Université Paris Descartes, Sorbonne Paris-Cité, 75014, FranceMembrane Traffic in Health and Disease, INSERM ERL U950, Univ Paris Diderot, Sorbonne Paris Cité, Institut Jacques Monod, CNRS UMR 7592, Paris, France, 75013, FranceMembrane Traffic in Health and Disease, INSERM ERL U950, Univ Paris Diderot, Sorbonne Paris Cité, Institut Jacques Monod, CNRS UMR 7592, Paris, France, 75013, FranceVAMP7 (vesicle-associated membrane protein) belongs to the intracellular membrane fusion SNARE (Soluble N-ethylmaleimide-sensitive factor attachment protein receptors) protein family. In this study, we used CRISPR/Cas9 genome editing technology to generate VAMP7 knockout (KO) human HeLa cells and mouse KO brain extracts in order to test the specificity and the background of a set of commercially available and homemade anti-VAMP7 antibodies. We propose a simple profiling method to analyze western blotting and use visual scoring for immunocytochemistry staining to determine the extent of the antibodies’ specificity. Thus, we were able to rank the performance of a set of available antibodies and further showed an optimized procedure for VAMP7 immunoprecipitation, which we validated using wild-type and KO mouse brain extracts.https://f1000research.com/articles/7-1649/v2 |
| spellingShingle | Agathe Verraes Beatrice Cholley Thierry Galli Sebastien Nola Comparative study of commercially available and homemade anti-VAMP7 antibodies using CRISPR/Cas9-depleted HeLa cells and VAMP7 knockout mice [version 2; referees: 2 approved] F1000Research |
| title | Comparative study of commercially available and homemade anti-VAMP7 antibodies using CRISPR/Cas9-depleted HeLa cells and VAMP7 knockout mice [version 2; referees: 2 approved] |
| title_full | Comparative study of commercially available and homemade anti-VAMP7 antibodies using CRISPR/Cas9-depleted HeLa cells and VAMP7 knockout mice [version 2; referees: 2 approved] |
| title_fullStr | Comparative study of commercially available and homemade anti-VAMP7 antibodies using CRISPR/Cas9-depleted HeLa cells and VAMP7 knockout mice [version 2; referees: 2 approved] |
| title_full_unstemmed | Comparative study of commercially available and homemade anti-VAMP7 antibodies using CRISPR/Cas9-depleted HeLa cells and VAMP7 knockout mice [version 2; referees: 2 approved] |
| title_short | Comparative study of commercially available and homemade anti-VAMP7 antibodies using CRISPR/Cas9-depleted HeLa cells and VAMP7 knockout mice [version 2; referees: 2 approved] |
| title_sort | comparative study of commercially available and homemade anti vamp7 antibodies using crispr cas9 depleted hela cells and vamp7 knockout mice version 2 referees 2 approved |
| url | https://f1000research.com/articles/7-1649/v2 |
| work_keys_str_mv | AT agatheverraes comparativestudyofcommerciallyavailableandhomemadeantivamp7antibodiesusingcrisprcas9depletedhelacellsandvamp7knockoutmiceversion2referees2approved AT beatricecholley comparativestudyofcommerciallyavailableandhomemadeantivamp7antibodiesusingcrisprcas9depletedhelacellsandvamp7knockoutmiceversion2referees2approved AT thierrygalli comparativestudyofcommerciallyavailableandhomemadeantivamp7antibodiesusingcrisprcas9depletedhelacellsandvamp7knockoutmiceversion2referees2approved AT sebastiennola comparativestudyofcommerciallyavailableandhomemadeantivamp7antibodiesusingcrisprcas9depletedhelacellsandvamp7knockoutmiceversion2referees2approved |