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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|
|Host / Isotype||Mouse / IgG1|
|Immunogen||GKPIPNPLLGLDST (V5) synthetic peptide conjugated to KLH|
|Storage buffer||PBS with proprietary stabilizer|
|Contains||0.02% sodium azide|
|Storage Conditions||4° C|
|Tested Applications||Dilution *|
|ELISA (ELISA)||Assay Dependent|
|Immunoprecipitation (IP)||Assay Dependent|
|Western Blot (WB)||1:500 - 1:1000|
* 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.
MA5-15253-BTIN has successfully been used for Western blot, ELISA, ICC/IF and IP applications.
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.