Background of Pk (V5) Epitope Tag (GKPIPNPLLGLDST) antibody
Epitope tags are short peptide sequences that are easily recognized by tag-specific antibodies. Due to their small size, epitope tags do not affect the tagged protein's biochemical properties. Most often sequences encoding the epitope tag are included with target DNA at the time of cloning to produce fusion proteins containing the epitope tag sequence. This allows anti-epitope tag antibodies to serve as universal detection reagents for any tag containing protein produced by recombinant means. This means that anti-epitope tag antibodies are a useful alternative to generating specific antibodies to identify, immunoprecipitate or immunoaffinity purify a recombinant protein.
This product has been used in:Randall RE et al. Isolation and characterization of monoclonal antibodies to simian virus 5 and their use in revealing antigenic differences between human, canine and simian isolates. J Gen Virol 68 ( Pt 11):2769-80 (1987). PubMed PMID: 2445904
Western blot using V5 tag antibody [V5.E3]: Western blot analysis of 293 cells transfected with V5-tagged vector (Lane 2) and untransfected control (Lane 1).
Anti-V5 EPITOPE TAG polyclonal antibody detects V5-tagged recombinant protein by western blot. This antibody was used at 1.0 μg/ml to detect 0.05 μg (lane 2) of full-length recombinant mouse serum albumin containing the V5 epitope tag at the carboxy end. Comparison to MW markers (lane 1) indicates detection of monomeric V5 tagged albumin. A 4-20% gradient gel was used to separate the protein by SDS-PAGE under non-reducing conditions. The protein was transferred to nitrocellulose using standard methods. After blocking the membrane was probed with the primary antibody overnight at 4° C followed by washes and reaction with a 1:10,000 dilution of IRDye(TM)800 conjugated Gt-a-Rabbit IgG [H&L] for 45 min at room temperature. LICOR's Odyssey® Infrared Imaging System was used to scan and process the image. Other detection systems will yield similarresults.
Figure 2. Lanes: 1:non-transfected cells; 2:V5-tagged empty plasmid; 3:protein A (V5-tagged); 4:protein B (V5-tagged); 5:protein C (V5-tagged); 6:protein D (V5-tagged); Protocol and data courtesy of Dr. Gustavo Gutierrez, Burnham Institute for Medical Research.
Samples: 200, 100, or 50 ng of E. coli whole cell lysate expressing a multi-tag fusion protein. Antibodies: Affinity-purified, HRP-conjugated, goat anti-V5 antibody GTX30569 used for WB at 0.04 µg/ml (1:25,000). Detection: Chemiluminescence with an exposure time of 10 seconds.
Western Blot: V5 Antibody [NB600-381] - Western blot analysis of V5-tagged Nicastrin, using NB600-381. Samples: NP-40 whole cell lysate (10 ug) from wild-type HEK293 cells (lane 1) or HEK293 cells expressing V5 tagged Nicastrin (lane 2). NB600-381 was used at 0.2 mcg/ml (1:5000).