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Key Specifications Table
| Species Reactivity | Key Applications |
|---|---|
| H, M, R | RIP, IP, WB, ICC, IHC |
| Description | |
|---|---|
| Catalogue Number | 03-102 |
| Brand Family | Upstate |
| Trade Name |
|
| Description | RIPAb+ HuR - RIP Validated Antibody and Primer Set |
| Overview | RIPAb+ antibodies are evaluated using the RNA Binding Protein Immunoprecipitation (RIP) assay. Each RIPAb+ antibody set includes quantitative RT-PCR control primers (RIP Primers) to biologically validate your IP results by successfully coprecipitating a specific RNA target (where such a specific target is known). The qPCR protocol and primer sequences are provided, allowing researchers to validate RIP protocols when using the antibody in their experimental context. Each set also includes a negative control antibody to ensure specificity of the RIP reaction. The RIPAb+ HuR set includes the HuR antibody, a negative control antibody (purified Rabbit IgG), and positive qPCR primers which amplify a 100 bp fragment of the human beta actin (ACTB) cDNA. The HuR and negative control antibodies are supplied in a scalable "per RIP" reaction size and can be used to functionally validate the precipitation HuR-associated RNAs. |
| Alternate Names |
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| Background Information | Hu-antigen R (HuB), ELAV-like protein 1, (ELAV1) is a transcription factor belonging to the ELAV family of RNP proteins. It binds avidly to the Au-rich element in cFos and interleukin-3 mRNAs. |
| Product Information | |
|---|---|
| Format | Purified |
| Control |
|
| Presentation | Anti-HuR (rabbit polyclonal). One vial containing 50 μg of purified rabbit polyclonal in 100 μL of buffer containing PBS in 0.05% sodium azide with 50% Glycerol. Liquid at -20°C. Normal Rabbit IgG. One vial containing 125 μg purified rabbit IgG in 125 μL storage buffer containing 0.1% sodium azide. Store at -20°C. RIP Primers, ACTB. One vial containing 75 μL of 5 μM of each primer specific for human beta actin (ACTB). Store at -20°C. FOR: TTG TTA CAG GAA GTC CCT TGC C REV: ATG CTA TCA CCT CCC CTG TGT g |
| Quality Level | MQ100 |
| Applications | |
|---|---|
| Application | This RIPAb+ HuR -RIP Validated Antibody & Primer Set conveniently includes the antibody & the specific control PCR primers. |
| Key Applications |
|
| Application Notes | Immunoprecipitation from RIP lysate: RIP lysate from MCF7 cells (2 X 106 cell equivalents per IP) was subjected to immunoprecipitation using 0.5 μg of either a normal rabbit IgG or Anti-HuR antibody. Precipitated proteins were resolved by electrophoresis, transferred to nitrocellulose and probed with anti-HuR (1.0 μg/mL). Proteins were visualized using One-Step™ IP-Western kit (GenScript Cat. # L00231) (Please see figures). Western Blot Analysis: 3T3/A31 lysate was resolved by electrophoresis, transferred to PVDF membranes and probed with HuR, hu (1:500 dilution). Proteins were visualized using a Donkey anti-Rabbit conjugated to HRP and visualized using a chemiluminescence detection system. (Please see figures). Immunoprecipitation Analysis: 10 ug of this antibody immunoprecipitated HuR from 500 ug of 3T3/A31 RIPA lysate (Please see figures). mmunocytochemistry Analysis: Confocal IF analysis of HeLa, NIH/3T3 using anti-HuR (Red). Actin filaments have been labeled with AlexaFluor® 488 -Phalloidin (Green). Nucleus is stained with DAPI (Blue) (Please see figures). |
| Biological Information | |
|---|---|
| Immunogen | KLH-conjugated, synthetic peptide corresponding to amino acids 1-13 of human HuR. |
| Epitope | a.a. 1-13 of HuR |
| Host | Rabbit |
| Specificity | HuR |
| Species Reactivity |
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| Antibody Type | Polyclonal Antibody |
| Entrez Gene Number |
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| Entrez Gene Summary | 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. |
| Gene Symbol |
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| Purification Method | Protein A Purfied |
| UniProt Number |
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| UniProt Summary | FUNCTION: SwissProt: Q15717 # Binds avidly to the AU-rich element in FOS and IL3/interleukin-3 mRNAs. In the case of the FOS AU-rich element, HUR binds to a core element of 27 nucleotides that contain AUUUA, AUUUUA, and AUUUUUA motifs. SIZE: 326 amino acids; 36092 Da SUBUNIT: Interacts with ANP32A. TISSUE SPECIFICITY: Ubiquitous. PTM: Methylated at Arg-217 by CARM1 in macrophages in response to LPS challenge. SIMILARITY: SwissProt: Q15717 ## Belongs to the RRM elav family. & Contains 3 RRM (RNA recognition motif) domains. |
| Molecular Weight | 35 kDa |
| Product Usage Statements | |
|---|---|
| Quality Assurance | RNA binding protein Immunoprecipitation: RIP Lysate prepared from HeLa cells (2 x 107 cell equivalents per IP) were subjected to immunoprecipitation using 5 µg of either a normal Rabbit IgG or Anti-HuR antibody and the Magna RIP Kit (Cat. # 17-700). Successful immunoprecipitation of HuR-associated RNA was verified by qPCR using RIP Primers, ACTB (Please see figures). Please refer to the Magna RIP™ (Cat. # 17-700) or EZ-Magna RIP™ (Cat. # 17-701) protocol for experimental details. |
| Usage Statement |
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| Storage and Shipping Information | |
|---|---|
| Storage Conditions | Stable for 1 year at -20°C from date of receipt. Note: Variability in freezer temperatures below -20°C may cause glycerol containing solutions to become frozen during storage. |
| Packaging Information | |
|---|---|
| Material Size | 10 assays |
| Material Package | 10 assays per set. Recommended use: ~5 μg of antibody per RIP (dependent upon biological context). |