Whole cell extracts (15 µg total protein) from HEK293 cells either non-transfected or transfected with an expression vector for a His-tagged protein were resolved on a 4-12% Bis-Tris gel, transferred to a PVDF membrane and then probed with HRP anti-His Tag (clone J099B12). Direct-Blot ™ HRP anti-GAPDH (Cat. No. 607904) was used as a loading control. Western-Ready™ ECL Substrate Premium Kit (Cat. No. 426318) was used as a detection agent. Lane M: Molecular weight marker
Whole cell extracts (15 µg total protein) from HEK293 cells either non-transfected or transfected with an expression vector for a His-tagged protein were resolved on a 4-12% Bis-Tris gel, transferred to a PVDF membrane and then probed with HRP anti-His Tag (clone J099B12). Direct-Blot ™ HRP anti-GAPDH (Cat. No. 607904) was used as a loading control. Western-Ready™ ECL Substrate Premium Kit (Cat. No. 426318) was used as a detection agent. Lane M: Molecular weight marker
ELISA assay was performed by coating a 96-well plate with 100 µL/well of a His-tagged recombinant protein (100 ng/mL). The coated His-tagged recombinant protein was detected with HRP anti-His Tag Antibody (Clone J099B12) followed by TMB High Sensitivity Substrate Solution (Cat. No. 421501). Absorbance was measured at 450 nm.
This antibody was raised against linear 8XHis tag. It reacts with His tag protein, either C, N, or internal His6 and up repeats. It can be used for Western blotting to detect linear His repeat (His6 or more).
This antibody is provided in 50% glycerol in aqueous buffered solution with preservatives.
Preparation
The antibody was purified by affinity chromatography and conjugated with HRP under optimal conditions.
Concentration
0.5 mg/mL
Storage & Handling
Upon receipt, the antibody solution should be stored undiluted at -20°C, and protected from prolonged exposure to light. Avoid exposure to sodium azide.
Each lot of this antibody is quality control tested by western blotting. For Western blotting, the suggested use of this reagent is 0.1 - 1.0 µg per mL. For direct ELISA, a concentration range of 0.001 - 0.1 µg per mL is recommended. Avoid using diluent solutions containing sodium azide. Sodium azide is an inhibitor of horseradish peroxidase. It is recommended that the end-user titrate each lot of reagents for optimal performance.
Application References
(PubMed link indicates BioLegend citation)
Burritt JB, et al. 1995. J. Biol. Chem. 270:16974.
Taylor RM, et al. 2007. Methods Mol. Biol. 412:429.
Product Citations
Hay CE, et al. 2020. J Pharmacol Exp Ther. 16:374. PubMed
Bojadzic D, et al. 2021. Front Pharmacol. 11:600372. PubMed
Konstantoulea K, et al. 2022. EMBO J. 41:e108591. PubMed
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