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Western blot analysis of His-tagged GFP was performed by loading 100ng of purified recombinant His-GFP and 7ul of Molecular Weight Protein Ladder per well onto a 4-12% Bis-Tris polyacrylamide gel. Proteins were transferred to a nitrocellulose membrane and blocked with 5% BSA/TBST for at least 1 hour. The membrane was probed with an anti-6X-His polyclonal antibody (Product # PA1983B) at a dilution of 1:250 overnight at 4°C on a rocking platform, washed in TBS-0.1%Tween-20, and probed with a goat anti-rabbit IgG-HRP secondary antibody (Product # 32460) at a dilution of 1:150,000 for at least 1 hour. Chemiluminescent detection was performed using SuperSignal West Dura (Product # 34075).
|Tested species reactivity||Tag|
|Published species reactivity||Yeast, Not Applicable|
|Host / Isotype||Rabbit / IgG|
|Immunogen||Synthetic peptide corresponding to residues H H H H H H.|
|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)||1:250|
* 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 2 publications below|
PA1-983B detects 6x-His Epitope tagged proteins
PA1-983B has been successfully used in Western blot procedures.
The PA1-983B immunogen is a synthetic peptide corresponding to residues H H H H H H.
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. Expression of recombinant proteins in E. coli as a fusion protein with neighboring histidine residues is one of the most popular methods of epitope tags. The affinity of the histidine-tag motif to Ni2+ by chelation is strong and selective enough to enable purification of the protein to homogeneity by affinity chromatography on a Ni2+-NTA adsorbent.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.
Identification of Novel Stress Granule Components That Are Involved in Nuclear Transport.
PA1-983B was used in western blot to study the role of importin-alpha subfamily members as novel stress granule components
|Mahboubi H,Seganathy E,Kong D,Stochaj U||PloS one (8:null)||2016|
Methods for efficient high-throughput screening of protein expression in recombinant Pichia pastoris strains.
PA1-983B was used in western blot to discuss methods for the high-throughput analysis of recombinant protein production by Pichia pastoris
|Camattari A,Weinhandl K,Gudiminchi RK||Methods in molecular biology (Clifton, N.J.) (1152:113)||2014|