702742

antibody from Invitrogen Antibodies

Targeting: HK1

Western blot (Supportive data in Antibodypedia)

Antibody Data

Product number

702742

Provider

Invitrogen Antibodies

Product name

HK1 Recombinant Rabbit Monoclonal Antibody (18H16L7)

Antibody type

Monoclonal

Antigen

Synthetic peptide

Description

This antibody is predicted to react with Monkey, Horse, Dog, Bovine

Antibody clone number

18H16L7

Concentration

0.5 mg/mL

Western blot

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Knockdown of Hexokinase 1 was achieved by transfecting A549 cells with Hexokinase 1 specific validated siRNA (Silencer® select Product # s6558). Western blot analysis (Fig a) was performed using whole cell extracts (1% SDS) from Hexokinase 1 knockdown cells (Lane 3), non-specific scrambled siRNA transfected cells (Lane 2) and untransfected cells (Lane 1). The blots were probed with Anti-Hexokinase 1 Recombinant Rabbit Monoclonal Antibody (Product # 702742, 1-3 µg/mL) and Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.25 µg/mL, 1:4000 dilution). Densitometric analysis of this Western blot is shown in histogram (Fig b). Loss of signal upon siRNA mediated knock down confirms that antibody is specific to Hexokinase 1.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot analysis was performed on Whole cell extracts (60 µg lysate) of PC-3 (Lane 1), K-562 (Lane 2), U-87 MG (Lane 3), MCF-7 (Lane 4), A-431 (Lane 5) PANC-1 (Lane 6) and A549 (Lane 7). The blots were probed with Anti-Hexokinase 1 Recombinant Rabbit Monoclonal Antibody (Product # 702742, 2.5 µg/mL) and detected by chemiluminescence using Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.25 µg/mL, 1:4000 dilution). A 102 kDa band corresponding to Hexokinase 1 was observed across the cell lines tested. Known quantity of protein samples were electrophoresed using Novex®NuPAGE®4-12% Bis-Tris gel (Product # NP0322BOX), XCell SureLock™ Electrophoresis System (Product # EI0002) and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary Antibody following blocking with 5% skimmed milk. Chemiluminescent detection was performed using Pierce™ ECL Western blotting Substrate (Product # 32106).