Western blot analysis of V5 was performed by loading 25ug of 293t cells transfected with 12Tag onto an SDS polyacrylamide gel. Proteins were transferred to a PVDF membrane and blocked at 4°C overnight. The membrane was probed with a V5 Epitope Tag polyclonal antibody (Product # PA1-993) at a dilution of 1:2000 overnight at 4°C, washed in TBST, and probed with an HRP-conjugated secondary antibody for 1 hr at room temperature in the dark. Chemiluminescent detection was performed using Pierce ECL Plus Western Blotting Substrate (Product # 32132). Results show a band at ~45 kDa.
|Tested species reactivity||Tag|
|Host / Isotype||Rabbit / IgG|
|Immunogen||Synthetic peptide corresponding to residues G K P I P N P L L G L D S T.|
|Purification||Antigen affinity chromatography|
|Storage buffer||PBS with 1mg/ml BSA|
|Contains||0.05% sodium azide|
|Storage Conditions||-20° C, Avoid Freeze/Thaw Cycles|
|Tested Applications||Dilution *|
|Western Blot (WB)||1:1000-1:5000|
* 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.
PA1-993 detects V5 tagged proteins.
PA1-993 has been used successfully in Western blot procedures. By Western blot this antibody detects the presence of V5-tagged proteins.
The PA1-993 immunogen corresponds to the V5 tag.
Epitope tagging is a powerful and versatile strategy for detecting and purifying proteins expressed by cloned genes. To utilize this feature, protein expression vectors are typically engineered with a nucleotide sequence that encodes the epitope tag. The gene of interest is cloned in-frame relative to the tag and, upon expression, the protein of interest is synthesized as a fusion protein with the epitope tag. Fusion protein detection and/or purification is mediated by highly specific antibodies to the engineered protein, thus eliminating the need for antibodies to proteins from each newly cloned gene. Commonly used epitope tags include glutathione-S-transferase (GST), c-myc, 6-histidine (6X-His), FLAG, green fluorescent protein (GFP), maltose binding protein (MBP), influenza A virus haemagglutinin (HA), b-galactosidase, and GAL4.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.