Customer Review for His-tag Polyclonal Antibody (STJ90106)


Target information

A polyhistidine-tag is an amino acid motif in proteins that consists of at least six histidine (His) residues, often at the N- or C-terminus of the protein. It is also known as hexa histidine-tag, 6xHis-tag, His6 tag and by the trademarked name His-tag. Polyhistidine-tags are often used for affinity purification of polyhistidine-tagged recombinant proteins expressed in Escherichia coli and other prokaryotic expression systems.

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1. HOW DID OUR ANTIBODY PERFORM? CUSTOMER REVIEW: His-tag Polyclonal Antibody (STJ90106)
2. WHAT WAS USED? Primary antibody: Product Catalog No: STJ90106. His-tag polyclonal antibody. 100ul Provider: St John’s Laboratory Dilution ratio: 1:1000 Secondary antibody: HRP goat anti-rabbit (provided by BPI) Reference antibody: Actin (provided by BPI)Figure 1. Lane1: HeLa; Lane 2: Jurkat
3. WHAT WAS THE PROTOCOL? Antibody: STJ90106. His-tag antibody Method for validation: Western Blot Materials for validation: HeLa cell line, Jurkat cell line Treatment of materials: When cells were cultured to 70%-80% plating density, lysis buffer with protease inhibitors (Sigma) and equal volume of 2X loading buffer were added for 10 min water bath at 100?C.Figure 1. Lane1: HeLa; Lane 2: Jurkat
4. WHAT WAS THE PROTOCOL? Gel electrophoresis: 10% polyacrylamide gel, constant voltage 160 V, 60 min. Transfer: 0.45 nm PVDF membrane, 1 piece of gel (wet transfer), constant voltage 100 V 60 min. Blocking: 1X TBST with 5% skimmed milk powder for 2 hr. Primary antibody probing: primary antibody was added to the solution of 1X TBST with 5% skimmed milk powder, 4?C for overnight. Membrane wash: 1X TBST wash for 3 times. Secondary antibody probing: secondary antibody was added to the solution of 1X TBST with 5% skimmed milk powder with 1:10000 dilution ratio, for 1.5 hr. Membrane wash: 1X TBST wash for 3 times. Visualization: ECL visualization for 5 min (the visualization time was determined by actual result. It was ensured that the visualization time was identical for the same antigen). Figure 1. Lane1: HeLa; Lane 2: Jurkat
5. WHAT WAS THE PROTOCOL? Gel electrophoresis information£º10% polyacrylamide gel, constant voltage 160 V, 60 min. Transfer information£º0.45 nm PVDF membrane, 1 piece of gel (wet transfer), constant voltage 100 V 60 min. No. Antigen Loading amount Primary antibody Primary antibody dilution ratio Secondary antibody dilution ratio Target band KD Visualization time 1 HeLa lysate 10 ug STJ90106. His-tag antibody 1:1000 1:10000 ~20KD 20S 2 Jurkat lysate 10 ug 1:10000 ~20KD 20S Figure 1. Lane1: HeLa; Lane 2: Jurkat
6. WHAT DID THE CUSTOMER THINK? Testing results were provided by BPI laboratory. They stated that ¡°In HeLa and Jurkat cell line, there were target bands detected by STJ90106 His-tag Antibody¡±.
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