720380

antibody from Invitrogen Antibodies

Targeting: ELF5

Western blot (Supportive data in Antibodypedia)

Immunocytochemistry (Supportive data in Antibodypedia)

Chromatin Immunoprecipitation (Supportive data in Antibodypedia)

Antibody Data

Product number

720380

Provider

Invitrogen Antibodies

Product name

ELF5 Polyclonal Antibody

Antibody type

Polyclonal

Antigen

Synthetic peptide

Description

This antibody is predicted to react with Monkey, Rat, Rabbit

Concentration

0.5 mg/mL

Western blot

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot analysis was performed on whole cell extracts (30 µg lysate) of MCF-7 (Lane 1), MDA-MB-231 (Lane 2), T-47D (Lane 3), U-87 MG (Lane 4), Hep G2 (Lane 5) and tissue extracts of Mouse Kidney (Lane 6), Rat Kidney (Lane 7), Mouse Testis (Lane 8) and Rat Testis (Lane 9). The blots were probed with Anti-ELF5 Rabbit Polyclonal Antibody (Product # 720380, 1-2 µg/mL). An isoform of 31 kDa was observed in all cell lines tested and 16 kDa was observed in all tissues tested. The blots were detected by chemiluminescence using Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.4 µg/mL, 1:5000 dilution). Known quantity of protein samples were electrophoresed using Novex® NuPAGE® 4-12% Bis-Tris gel (Product # NP0321BOX), 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).

Immunocytochemistry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

For immunofluorescence analysis, Ntera2 cells were fixed and permeabilized for detection of endogenous ELF5 using Anti- ELF5 Rabbit polyclonal Antibody (Product # 720380, 2 µg/mL) and labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034, 1:2000). Panel a) shows representative cells that were stained for detection and localization of ELF5 protein (green), Panel b) is stained for nuclei (blue) using SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). Panel c) represents cytoskeletal F-actin staining using Rhodamine Phalloidin (Product # R415, 1:300). Panel d) is a composite image of panels a, b and c clearly demonstrating nuclear localization of ELF5. Panel e) represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.

Chromatin Immunoprecipitation

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Enrichment of endogenous ELF5 protein at specific gene loci using Anti-ELF5 Rabbit Polyclonal Antibody: Chromatin Immunoprecipitation (ChIP) was performed using Anti-ELF5 Rabbit Polyclonal Antibody (Product # 720380, 5 µg) on sheared chromatin from 2 million MDA-MB-231 cells using the MAGnify ChIP system kit (Product # 49-2024). Normal Rabbit IgG (1 µg) was used as a negative IP control. The purified DNA was analyzed by 7500 Fast qPCR system (Product # 4351106) with optimized PCR primer pairs for the promoter of the active SNAI2 region used as positive control target genes, and the region of the inactive, SAT alpha, SAT2 satellite repeat, used as negative control target gene. Data is presented as fold enrichment of the antibody signal versus the negative control IgG using the comparative CT method.