Sphingosine kinase 2 (SK2) is a ubiquitously expressed lipid kinase that has important, albeit complex and poorly understood, roles in regulating cell survival and cell death. In addition to being able to promote cell cycle arrest and apoptosis under certain conditions, it has recently been shown that SK2 can promote neoplastic transformation and tumorigenesis in vivo. Therefore, well validated and reliable tools are required to study and better understand the true functions of SK2. Here, we compare two commercially available SK2 antibodies: a rabbit polyclonal antibody from Proteintech that recognizes amino acids 266-618 of human SK2a, and a rabbit polyclonal antibody from ECM Biosciences that recognizes amino acids 36-52 of human SK2a. We examine the performance of these antibodies for use in immunoblotting, immunoprecipitation and immunofluorescence staining of endogenous SK2, using human HEK293 and HeLa cell lines, as well as mouse embryonic fibroblasts (MEFs). Furthermore, we assess the specificity of these antibodies to the target protein through the use of siRNA-mediated SK2 knockdown and SK2 knockout ( Sphk2-/-) MEFs. Our results demonstrate that the Proteintech anti-SK2 antibody reproducibly displayed superior sensitivity and selectivity towards SK2 in immunoblot analyses, while the ECM Biosciences anti-SK2 antibody was reproducibly superior for SK2 immunoprecipitation and detection by immunofluorescence staining. Notably, both antibodies produced non-specific bands and staining in the MEFs, which was not observed with the human cell lines. Therefore, we conclude that the Proteintech SK2 antibody is a valuable reagent for use in immunoblot analyses, and the ECM Biosciences SK2 antibody is a useful tool for SK2 immunoprecipitation and immunofluorescence staining, at least in the human cell lines employed in this study.
Pubmed ID: 28105318 RIS Download
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View all literature mentionsCollection of data of protein sequence and functional information. Resource for protein sequence and annotation data. Consortium for preservation of the UniProt databases: UniProt Knowledgebase (UniProtKB), UniProt Reference Clusters (UniRef), and UniProt Archive (UniParc), UniProt Proteomes. Collaboration between European Bioinformatics Institute (EMBL-EBI), SIB Swiss Institute of Bioinformatics and Protein Information Resource. Swiss-Prot is a curated subset of UniProtKB.
View all literature mentionsCommercial instrument and chemical vendor. Developer and manufacturer of specialized technological products for life science research and clinical diagnostics markets.
View all literature mentionsCommercial vendor and service provider of laboratory reagents and antibodies. Supplier of scientific instrumentation, reagents and consumables, and software services.
View all literature mentionsProteintech Europe Ltd is an ISO 9001:2008 certified company
View all literature mentionsMus musculus with name C57BL/6J from IMSR.
View all literature mentionsCell line HEK293 is a Transformed cell line with a species of origin Homo sapiens (Human)
View all literature mentionsCell line HeLa is a Cancer cell line with a species of origin Homo sapiens
View all literature mentionsThis polyclonal targets
View all literature mentionsThis polyclonal targets IgG (H+L)
View all literature mentionsThis polyclonal secondary targets IgG (H+L)
View all literature mentionsThis polyclonal secondary targets IgG (H+L)
View all literature mentionsThis monoclonal targets alpha Tubulin antibody [DM1A] - Loading Control
View all literature mentionsThis polyclonal targets SPHK2
View all literature mentionsThis polyclonal targets IgG (H+L)
View all literature mentionsThis polyclonal secondary targets IgG (H+L)
View all literature mentionsThis polyclonal secondary targets IgG (H+L)
View all literature mentionsThis monoclonal targets alpha Tubulin antibody [DM1A] - Loading Control
View all literature mentionsThis polyclonal targets SPHK2
View all literature mentionsThis polyclonal targets
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