Invitrogen

HuR Monoclonal Antibody (3A2)

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This Antibody was verified by Relative expression to ensure that the antibody binds to the antigen stated. View Details

View all (23) HuR antibodies

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Datasheet

Questions & Answers

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HuR Antibody in Western Blot (WB) — Western blot analysis was performed on whole cell extracts (30 µg lysate) of HeLa (Lane 1), HCT 116 (Lane 2), Jurkat (Lane 3), MCF7 (Lane 4), Hep G2 (Lane 5), A549 (Lane 6), LNCap (Lane 7), K-562 (Lane 8), MOLT4 (Lane 9), Ramos (Lane 10) and NIH/3T3 (Lane 11). The blot was probed with Anti-HuR Monoclonal Antibody (3A2) (Product # MA1-167, 1:500 dilution) and detected by chemiluminescence using Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A28177, 0.25µg/ml, 1:4000 dilution). A 38 kDa band corresponding to Aconitase was observed across the cell lines tested.

HuR Antibody in Immunofluorescence (IF) — Immunofluorescent analysis of HuR (3A2) (green) in HeLa cells. Cells were fixed in 4% paraformaldhyde, permeabilized with 0.1% Triton X-100, and blocked with 1% Blocker BSA (Product # 37525), each for 15 minutes at room temperature. Cells were stained with HuR (3A2) monoclonal antibody (Product # MA1-167) at a dilution of 1:50 in blocking buffer at 37C for at least one hour. Cells were then washed with PBS and then incubated with a DyLight 488-conjugated goat anti-mouse IgG secondary antibody (Product # 35502) at a dilution of 1:1000 for 1 hour at room temperature. Nuclei (blue) were stained with with DAPI (Product # 46190). Images were taken on a Thermo Scientific ToxInsight Instrument at 20X magnification.

HuR Antibody in Immunoprecipitation (IP) — Immunoprecipitation of HuR was performed using MCF-7 cells. Antigen-antibody complexes were formed by incubating 500 µg of MCF-7 whole cell lysates with 2 µg of HuR (3A2) monoclonal antibody (Product # MA1-167) overnight on a rocking platform at 4C. The immune complexes were captured on 100 µL Protein A/G Agarose (Product # 20421), washed extensively, and eluted with 5X Lane Marker Reducing Sample Buffer (Product # 39000). The IP eluate (right lane) and 20 µg of MCF-7 lysate (left lane) were resolved on a 4-20% Tris-Glycine polyacrylamide gel, transferred to a nitrocellulose membrane, and blocked with 5% Milk in TBST for at least 1 hour. The membrane was probed with a HuR (3A2) monoclonal antibody monoclonal antibody (Product # MA1-167) at a dilution of 1:250 overnight rotating at 4C, washed in TBST, and probed with goat anti-mouse IgG-HRP secondary antibody (Product # 31430) at a dilution of 1:10,000 for at least 30 minutes at room temperature. Chemiluminescent detection was performed using SuperSignal West Pico (Product # 34080).

HuR Antibody in RNA Immunoprecipitation (RIP) — RNA immunoprecipitation (RIP) western of ELAVL1 was performed on K562 cells. Antigen-antibody complexes were formed by incubating approximately 500 µg whole cell lysate with 5 µg of monoclonal ELAVL1 antibody (Product # MA1-167) rotating 60 min at RT. The immune complexes were captured on 625 µg of Dynabeads M-280 sheep anti-mouse IgG (Product # 11202D), washed extensively, and eluted in NuPAGE LDS sample buffer (Product # NP0007). Samples were resolved onto a 4-12% Bis-Tris protein gel (Product # NP0335BOX). Lanes 1 and 3 are input and lanes 2 and 4 are IP. Proteins were transferred to nitrocellulose membrane (Product # IB23001) and blocked in 5% milk. ELAVL1 was detected at approximately 36 kDa using a monoclonal ELAVL1 antibody (Product # MA1-167) at a dilution of 1:2000, followed by incubation with anti-mouse secondary antibody. Chemiluminescent detection was performed using ECL Western Blotting Substrate (Product # 32106). Data courtesy of the Yeo lab as part of the ENCODE project (www.encodeproject.org).

HuR Antibody in Relative expression — Antibody specificity was demonstrated by detection of binding of the target protein to specific RNA. RNA Immunoprecipitation (RIP) was performed using Anti-HuR Monoclonal Antibody (Product # MA1-167) using primer pairs over ACTB mRNA (positive) and 18S rRNA (inactive). {RE}

HuR Antibody in Cell treatment — Altered expression of proteins upon cell treatment demonstrates antibody specificity. Immunofluorescence analysis using HuR Monoclonal Antibody (3A2) (Product # MA1-167), shows translocation from nucleus to cytoplasm in HeLa cell line when treated with Actinomycin D. {TM}

Product Details

Applications

Tested Dilution

Publications

Western Blot (WB)

1:125-1:500

Immunocytochemistry (ICC)

1:25-1:100

Immunofluorescence (IF)

1:25-1:100

Immunoprecipitation (IP)

2 µg/10^6 cells

RNA Immunoprecipitation (RIP)

2 µg/10^6 cells

Product Specifications

Species Reactivity

Human, Mouse, Rat

Host/Isotype

Mouse / IgG1

Class

Monoclonal

Type

Antibody

Clone

3A2

Immunogen

Recombinant human HuR protein

Conjugate

Unconjugated

Form

Liquid

Concentration

0.5 mg/mL

Purification

Protein A

Storage buffer

PBS, pH 7.2-7.4, with 1mg/mL BSA, 30% glycerol

Contains

0.05% sodium azide

Storage conditions

-20° C, Avoid Freeze/Thaw Cycles

RRID

AB_2536870

Target Information

The protein encoded by this gene is a member of the ELAVL protein family. This encoded protein contains 3 RNA-binding domains and binds cis-acting AU-rich elements. It destabilizes mRNAs and thereby regulates gene expression.

For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.

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Bioinformatics

Protein Aliases: ELAV (embryonic lethal, abnormal vision, Drosophila)-like 1 (Hu antigen R); ELAV like 1; Elav-like generic protein; ELAV-like protein 1; embryonic lethal, abnormal vision, drosophila, homolog-like 1; Hu antigen R; HU-antigen A; Hu-antigen R; Human antigen R; HuR; MelG

Gene Aliases: 2410055N02Rik; ELAV1; ELAVL1; Elra; Hua; HUR; MelG; W91709

UniProt ID: (Human) Q15717, (Mouse) P70372, (Rat) B5DF91

Entrez Gene ID: (Human) 1994, (Mouse) 15568, (Rat) 363854

Function(s)

mRNA splicing, via spliceosome multicellular organism development regulation of mRNA stability positive regulation of translation mRNA stabilization negative regulation of gene silencing by miRNA 3'-UTR-mediated mRNA stabilization regulation of stem cell population maintenance regulation of translation

Process(es)