M. Biddle, Charles Alende, Maryam Fotouhi, Carolyn Jones, Riham Ayoubi, K. Southern, C. Laflamme, H. Virk
{"title":"用于 Western 印迹、免疫沉淀、免疫荧光和流式细胞术的 Synaptotagmin-1(Uniprot ID P21579)高效抗体选择指南","authors":"M. Biddle, Charles Alende, Maryam Fotouhi, Carolyn Jones, Riham Ayoubi, K. Southern, C. Laflamme, H. Virk","doi":"10.12688/f1000research.154034.1","DOIUrl":null,"url":null,"abstract":"Synaptotagmin-1 is a synaptic vesicle transmembrane protein that senses calcium influx via its tandem C2-domains, triggering synchronous neurotransmitter release. Disruption to SYT1 is associated with neurodevelopmental disorders, highlighting the importance of identifying high-quality research reagents to enhance understanding of Synaptotagmin-1 in health and disease. Here we have characterized thirteen Synaptotagmin-1 commercial antibodies for western blot, immunoprecipitation, immunofluorescence and flow cytometry using a standardized experimental protocol based on comparing read-outs in knockout cell lines and isogenic parental controls. These studies are part of a larger, collaborative initiative seeking to address antibody reproducibility issues by characterizing commercially available antibodies for human proteins and publishing the results openly as a resource for the scientific community. While use of antibodies and protocols vary between laboratories, we encourage readers to use this report as a guide to select the most appropriate antibodies for their specific needs.","PeriodicalId":504605,"journal":{"name":"F1000Research","volume":null,"pages":null},"PeriodicalIF":0.0000,"publicationDate":"2024-07-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"A guide to selecting high-performing antibodies for Synaptotagmin-1 (Uniprot ID P21579) for use in western blot, immunoprecipitation, immunofluorescence and flow cytometry\",\"authors\":\"M. Biddle, Charles Alende, Maryam Fotouhi, Carolyn Jones, Riham Ayoubi, K. Southern, C. Laflamme, H. Virk\",\"doi\":\"10.12688/f1000research.154034.1\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"Synaptotagmin-1 is a synaptic vesicle transmembrane protein that senses calcium influx via its tandem C2-domains, triggering synchronous neurotransmitter release. Disruption to SYT1 is associated with neurodevelopmental disorders, highlighting the importance of identifying high-quality research reagents to enhance understanding of Synaptotagmin-1 in health and disease. Here we have characterized thirteen Synaptotagmin-1 commercial antibodies for western blot, immunoprecipitation, immunofluorescence and flow cytometry using a standardized experimental protocol based on comparing read-outs in knockout cell lines and isogenic parental controls. These studies are part of a larger, collaborative initiative seeking to address antibody reproducibility issues by characterizing commercially available antibodies for human proteins and publishing the results openly as a resource for the scientific community. While use of antibodies and protocols vary between laboratories, we encourage readers to use this report as a guide to select the most appropriate antibodies for their specific needs.\",\"PeriodicalId\":504605,\"journal\":{\"name\":\"F1000Research\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":0.0000,\"publicationDate\":\"2024-07-19\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"F1000Research\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"https://doi.org/10.12688/f1000research.154034.1\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"\",\"JCRName\":\"\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"F1000Research","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.12688/f1000research.154034.1","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
引用次数: 0
摘要
SYT1是一种突触小泡跨膜蛋白,通过其串联的C2-结构域感知钙离子的流入,从而触发神经递质的同步释放。SYT1 的破坏与神经发育障碍有关,这凸显了鉴定高质量研究试剂的重要性,以加深对突触表蛋白-1 在健康和疾病中的作用的了解。在此,我们采用标准化实验方案,比较了基因敲除细胞系和同源亲本对照的读出结果,鉴定了 13 种 Synaptotagmin-1 商用抗体,用于 Western 印迹、免疫沉淀、免疫荧光和流式细胞术。这些研究是一项大型合作计划的一部分,该计划旨在通过鉴定市售的人类蛋白质抗体来解决抗体的可重复性问题,并将结果公开发表,作为科学界的资源。虽然不同实验室使用的抗体和方案各不相同,但我们鼓励读者以本报告为指南,根据自己的具体需要选择最合适的抗体。
A guide to selecting high-performing antibodies for Synaptotagmin-1 (Uniprot ID P21579) for use in western blot, immunoprecipitation, immunofluorescence and flow cytometry
Synaptotagmin-1 is a synaptic vesicle transmembrane protein that senses calcium influx via its tandem C2-domains, triggering synchronous neurotransmitter release. Disruption to SYT1 is associated with neurodevelopmental disorders, highlighting the importance of identifying high-quality research reagents to enhance understanding of Synaptotagmin-1 in health and disease. Here we have characterized thirteen Synaptotagmin-1 commercial antibodies for western blot, immunoprecipitation, immunofluorescence and flow cytometry using a standardized experimental protocol based on comparing read-outs in knockout cell lines and isogenic parental controls. These studies are part of a larger, collaborative initiative seeking to address antibody reproducibility issues by characterizing commercially available antibodies for human proteins and publishing the results openly as a resource for the scientific community. While use of antibodies and protocols vary between laboratories, we encourage readers to use this report as a guide to select the most appropriate antibodies for their specific needs.