Customer Review For ENT1 Polyclonal Antibody (STJ96396)

Target information

Equilibrative nucleoside transporter 1 (ENT1) is a protein that in humans is encoded by the SLC29A1 gene. Multiple alternatively spliced variants, encoding the same protein, have been found for this gene.

This gene is a member of the equilibrative nucleoside transporter family. The gene encodes a transmembrane glycoprotein that localizes to the plasma and mitochondrial membranes and mediates the cellular uptake of nucleosides from the surrounding medium. The protein is categorized as an equilibrative (as opposed to concentrative) transporter that is sensitive to inhibition by nitrobenzylmercaptopurine ribonucleoside (NBMPR). Nucleoside transporters are required for nucleotide synthesis in cells that lack de novo nucleoside synthesis pathways, and are also necessary for the uptake of cytotoxic nucleosides used for cancer and viral chemotherapies.

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2. WHAT WAS USED? Primary antibody: STJ96396. ENT1 Polyclonal Antibody. 100ul Provider: St John’s Laboratory Dilution ratio: 1:300 Application: Western Blot. Materials for validation: Rh30 cell line, Rh41 cell line, HT-29 cell line, Colo320DM cell line, Hela cell line, NB1643 cell line Treatment of materials: When cells were cultured to 70%- 80% plating density, lysis buffer with protease inhibitors and 4X loading buffer were added for 5 min water bath at 100?C. Image: Lane1: Rh30; Lane 2: Rh41; Lane 3: HT-29; Land 4: Colo320DM; Lane 5: Hela, Lane 6: NB1643.
3. 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 (dry transfer), 7 min. Blocking: 1X TBST with 5% skimmed milk powder for 1 hr. Primary antibody probing: primary antibody was added to the solution of 1X TBST, 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 1:3000 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).
4. 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 (dry transfer), 7 min. No. Antigen Loading amount Primary antibody Primary antibody dilution ratio Secondary antibody dilution ratio Target band KD Visualization time 1 Rho30 lysate 50ug STJ9639 ENT1 antibody 1:300 1:3000 ~55KD 2 min 2 Rh41lysate 50ug ~55KD 2 min 3 HT-29 lysate 50ug ~55KD 2 min 4 Colo320DM 50ug ~55KD 2 min 5 HeLa lysate 50ug ~55KD 2 min 6 NB1643 lysate 50ug ~55KD 2 min
5. WHAT DID THE CUSTOMER THINK? Testing results were provided by Tingting Wang from Nationwide Children¡¯s Hospital. They stated that ¡°In all cell lines, there were target bands detected by STJ96396 ENT1 antibody with different levels.¡±
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