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Key Specifications Table
| Species Reactivity | Key Applications |
|---|---|
| H, M, R | RIP, IP, WB |
| Description | |
|---|---|
| Catalogue Number | 03-111 |
| Brand Family | Upstate |
| Trade Name |
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| Description | RIPAb+ AUF1 - RIP Validated Antibody and Primer Set |
| Overview | RIPAb+ antibodies are evaluated using the RNA Binding Protein Immunoprecipitation (RIP) assay. Each RIPAb+ antibody set includes a negative control antibody to ensure specificity of the RIP reaction and is verified for the co-immunoprecipitation of RNA associated specifically with the immunoprecipitated RNA binding protein of interest. Where appropriate, the RIPAb+ set also includes quantitative RT-PCR control primers (RIP Primers) to biologically validate your IP results by successfully co-precipitating the specific RNA targets, such as messenger RNAs. The qPCR protocol and primer sequences are provided, allowing researchers to validate RIP protocols when using the antibody in their experimental context. If a target specific assay is not provided, the RIPAb+ kit is validated using an automated microfluidics-based assay by enrichment of detectable RNA over control immunoprecipitation. |
| Alternate Names |
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| Background Information | A+U Rich RNA Binding Factor (AUF1) is comprised of four isoforms of 37, 40, 42, and 45 kDa. Although the role of each isoform has yet to be fully characterized, a direct correlation has been observed between each AUF1 isoform’s binding affinity and its RNA destabilizing activity toward different AREs (A + U rich element in the 3’ untranslated region), with the isoforms p37 and p42 being the most effective. AUF1 is a bcl2 mRNA binding protein and potentially all its isoforms are able to form complexes with the bcl2 ARE. ARE mediated bcl2 mRNA downregulation during apoptosis involves AUF1 and suggest different roles for its four isoforms. |
| Product Information | |
|---|---|
| Format | Purified |
| Control |
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| Presentation | Anti-AUF1 (Rabbit Polyclonal). One vial containing 50 μg of protein A purified rabbit IgG in 50 μL of 70% storage buffer (0.1M Tris-glycine, pH 7.4, 0.15M NaCl, 0.05% sodium azide) and 30% glycerol. Store at -20°C. Normal Rabbit IgG. One vial containing 125 μg Rabbit IgG in 125 μL storage buffer containing 0.05% sodium azide. Store at -20°C. RIP Primers, FOS. One vial containing 75 μL of 5 μM of each primer specific for human FOS. Store at -20°C. FOR: GAG AGC TGG TAG TTA GTA GCA TGT TGA REV: AAT TCC AAT AAT GAA CCC AAT AGA TTA GTT A |
| Quality Level | MQ100 |
| Applications | |
|---|---|
| Application | This RIPAb+ AUF1 -RIP Validated Antibody & Primer Set conveniently includes the antibody & the specific control PCR primers. |
| Key Applications |
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| Application Notes | Immunoprecipitation from RIP lysate: Representative lot data. RIP lysate from HeLa cells (2 X 106 cell equivalents per IP) was subjected to immunoprecipitation using 0.5 µg of either a normal rabbit IgG or Anti-AUF1 antibody. Precipitated proteins (Lane 1: rabbit IgG, Lane 2: anti-AUF1) and HeLa whole cell lysate (Lane 3) were resolved by electrophoresis, transferred to nitrocellulose and probed with anti-AUF1 antibody (1.0 µg/mL). Proteins were visualized using One-Step™ IP-Western kit (GenScript Cat. # L00231) (Please see figures). Western Blot Analysis: 0.01-1 µg/mL of a previous lot detected AUF1 in RIPA lysates from HeLa nuclear extract and A431 cells. |
| Biological Information | |
|---|---|
| Immunogen | Human AUF1 purified by Ni2+ affinity column. |
| Host | Rabbit |
| Specificity | AUF1 isoforms at 37, 40, 42 and 45 kDa. |
| Species Reactivity |
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| Species Reactivity Note | Human. Predicted cross-reactivity with mouse and rat based on sequence homology. |
| Antibody Type | Polyclonal Antibody |
| Entrez Gene Number |
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| Entrez Gene Summary | This gene belongs to the subfamily of ubiquitously expressed heterogeneous nuclear ribonucleoproteins (hnRNPs). The hnRNPs are nucleic acid binding proteins and they complex with heterogeneous nuclear RNA (hnRNA). These proteins are associated with pre-mRNAs in the nucleus and appear to influence pre-mRNA processing and other aspects of mRNA metabolism and transport. While all of the hnRNPs are present in the nucleus, some seem to shuttle between the nucleus and the cytoplasm. The hnRNP proteins have distinct nucleic acid binding properties. The protein encoded by this gene has two repeats of quasi-RRM domains that bind to RNAs. It localizes to both the nucleus and the cytoplasm. This protein is implicated in the regulation of mRNA stability. Alternative splicing of this gene results in four transcript variants. |
| Gene Symbol |
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| UniProt Number |
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| UniProt Summary | Function: Binds with high affinity to RNA molecules that contain AU-rich elements (AREs) found within the 3-UTR of many proto-oncogenes and cytokine mRNAs. Also binds to double- and single-stranded DNA sequences in a specific manner and functions a transcription factor. Each of the RNA-binding domains specifically can bind solely to a single-stranded non-monotonous 5-UUAG-3 sequence and also weaker to the single-stranded 5-TTAGGG-3 telomeric DNA repeat. Binds RNA oligonucleotides with 5-UUAGGG-3 repeats more tightly than the telomeric single-stranded DNA 5-TTAGGG-3 repeats. Binding of RRM1 to DNA inhibits the formation of DNA quadruplex structure which may play a role in telomere elongation. May be involved in translationally coupled mRNA turnover. Implicated with other RNA-binding proteins in the cytoplasmic deadenylation/translational and decay interplay of the FOS mRNA mediated by the major coding-region determinant of instability (mCRD) domain. Subunit structure: Identified in a mRNP granule complex, at least composed of ACTB, ACTN4, DHX9, ERG, HNRNPA1, HNRNPA2B1, HNRNPAB, HNRNPD, HNRNPL, HNRNPR, HNRNPU, HSPA1, HSPA8, IGF2BP1, ILF2, ILF3, NCBP1, NCL, PABPC1, PABPC4, PABPN1, RPLP0, RPS3, RPS3A, RPS4X, RPS8, RPS9, SYNCRIP, TROVE2, YBX1 and untranslated mRNAs. Part of a complex associated with the FOS mCRD domain and consisting of PABPC1, PAIP1, CSDE1/UNR and SYNCRIP. Interacts with IGF2BP2. Subcellular location: Nucleus. Cytoplasm. Note: Localized in cytoplasmic mRNP granules containing untranslated mRNAs. Component of ribonucleosomes. Post-translational modification: Arg-345 is dimethylated, probably to asymmetric dimethylarginine. Sequence similarities: Contains 2 RRM (RNA recognition motif) domains. |
| Molecular Weight | 37, 40, 42 and 45 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 5 µg of Anti-AUF1 antibody and the Magna RIP™ RNA-Binding Protein Immunoprecipitation Kit (Cat. # 17-700). Successful immunoprecipitation of AUF1-associated RNA was verified by qPCR using RIP Primers FOS (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. Handling Recommendations: Upon receipt, and prior to removing the cap, centrifuge the vial and gently mix the solution. Aliquot into microcentrifuge tubes and store at -20°C. Avoid repeated freeze/thaw cycles, which may damage IgG and affect product performance. 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). |