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Western blot analysis of V5-tagged proteins was performed by loading 15ug of 293T cell lysates, from cells transfected with a V5-Wnt3a or V5-Vstm2 construct for the indicated number of hours, per well onto an SDS-PAGE gel. Proteins were transferred to a PVDF membrane and blocked with 5% milk in PBST for 1 hour at room temperature. The membrane was probed with a V5 Epitope Tag monoclonal antibody (Product # MA5-15253) at a dilution of 1:1000 overnight at 4°C, washed in PBST, and probed with an HRP-conjugated anti-mouse IgG secondary antibody at a dilution of 1:5000 for 1 hour at room temperature. Detection was performed using ECL substrate. Data courtesy of the Innovators Program.
|Tested species reactivity||Tag|
|Published species reactivity||Human, Not Applicable|
|Host / Isotype||Mouse / IgG1|
|Immunogen||GKPIPNPLLGLDST (V5) synthetic peptide conjugated to KLH|
|Storage buffer||PBS, pH 7.2|
|Storage Conditions||Store at 4°C short term. For long term storage, store at -20°C, avoiding freeze/thaw cycles.|
|Tested Applications||Dilution *|
|ELISA (ELISA)||Assay Dependent|
|Immunofluorescence (IF)||1:500 - 1:2000|
|Immunoprecipitation (IP)||Assay Dependent|
|Western Blot (WB)||1:1000 - 1:3000|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
|Western Blot (WB)||See 3 publications below|
MA5-15253 recognizes V5 (GKPIPNPLLGLDST) tagged fusion proteins.
MA5-15253 has successfully been used in immunoprecipitation, ELISA, Western blotting, and immunofluorescence applications.
MA5-15253 may be stored at 4°C short term. Add 0.05% sodium azide if desired. For long term storage, store at -20°C, avoiding freeze/thaw cycles.
Epitope tags provide a method to localize gene products in a variety of cell types, study the topology of proteins and protein complexes, identify associated proteins, and characterize newly identified, low abundance or poorly immunogenic proteins when protein specific antibodies are not available. The V5 epitope tag is derived from a small epitope (Pk) present on the P and V proteins of the paramyxovirus of simian virus (SV5). The V5 tag is usually used with all 14 amino acids (GKPIPNPLLGLDST), although it has also been used with a shorter 9 amino acid sequence (IPNPLLGLD).
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.
Cyclophilin-D: a resident regulator of mitochondrial gene expression.
MA5-15253 was used in ChIP assay and western blot to determine if cyclophilin-D regulates mitochondrial gene expression
|Radhakrishnan J,Bazarek S,Chandran B,Gazmuri RJ||FASEB journal : official publication of the Federation of American Societies for Experimental Biology (29:2734)||2015|
E2F1-Mediated Induction of NFYB Attenuates Apoptosis via Joint Regulation of a Pro-Survival Transcriptional Program.
MA5-15253 was used in western blot to identify the NF-YB transcription factor as a novel direct E2F1 target
|Jiang X,Nevins JR,Shats I,Chi JT||PloS one (10:null)||2015|
Mutations in SPINT2 cause a syndromic form of congenital sodium diarrhea.
MA5-15253 was used in western blot to identify the genetic basis of congenital sodium diarrhea
|Heinz-Erian P,Müller T,Krabichler B,Schranz M,Becker C,Rüschendorf F,Nürnberg P,Rossier B,Vujic M,Booth IW,Holmberg C,Wijmenga C,Grigelioniene G,Kneepkens CM,Rosipal S,Mistrik M,Kappler M,Michaud L,Dóczy LC,Siu VM,Krantz M,Zoller H,Utermann G,Janecke AR||American journal of human genetics (84:188)||2009|