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  • 03-191 - RIPAb+ Upf1 - RIP Validated Antibody and Primer Set

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03-191 Sigma-Aldrich

RIPAb+ Upf1 - RIP Validated Antibody and Primer Set

from rabbit

Synonym: ATP-dependent helicase RENT1, Nonsense mRNA reducing factor 1, UP Frameshift 1, UPF1 regulator of nonsense transcripts homolog (yeast), Up-frameshift suppressor 1 homolog, delta helicase, regulator of nonsense transcripts 1, up-frameshift mutation 1 homo

  •  eCl@ss 32160702

  •  NACRES NA.32

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Properties

Related Categories Antibodies, Antibody Kits, Immunoprecipitation Kits More...
Quality Level   100
biological source   rabbit
clone   polyclonal
species reactivity   human, canine, mouse, bovine, human, rat
species reactivity (predicted by homology)   rat, canine, mouse, bovine
mfr. no.   RIPAb+
  Upstate®
application(s)   RIP: suitable
  immunoprecipitation (IP): suitable
  western blot: suitable
NCBI accession no.   NP_002902.2
UniProt accession no.   Q92900
shipped in   dry ice

Description

General description

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.
Upf1 is part of a post-splicing multiprotein complex involved in both mRNA nuclear export and mRNA surveillance. Upf1 is located only in the cytoplasm. When translation ends, it interacts with the protein that is a functional homolog of yeast Upf2p to trigger mRNA decapping.

Upf1 is part of a post-splicing multiprotein complex involved in both mRNA nuclear export and mRNA surveillance. mRNA surveillance detects exported mRNAs with truncated open reading frames and initiates nonsense-mediated mRNA decay (NMD). When translation ends upstream from the last exon-exon junction, this triggers NMD to degrade mRNAs containing premature stop codons. Upf1 is located only in the cytoplasm. When translation ends, it interacts with the protein that is a functional homolog of yeast Upf2p to trigger mRNA decapping.

Specificity

This antibody detects Upf1.

Immunogen

KLH-conjugated synthetic linear peptide corresponding to Upf1.

Application

Immunoprecipitation from RIP lysate:
Representative lot data.
RIP lysate from HeLa cells (~2 X 10E7 cell equivalents per IP) was subjected to immunoprecipitation using either 7 µL of a normal rabbit IgG or 7 µL of Anti-Upf1 antibody.
Ten percent of the precipitated proteins (lane 1: rabbit IgG, lane 2: Upf1) were resolved by electrophoresis, transferred to PVDF and probed with anti-Upf1 antibody (1:2000).
Proteins were visualized using One-Step IP-Western kit (GenScript Cat. # L00232)
Arrow indicates Upf1 (~125 kDa) (Please see figures).

Western Blot Analysis:
Representative lot data.
U2OS cell lysate was resolved by electrophoresis, transferred to PVDF membrane and probed with anti-Upf1 (0.5 μg/mL).
Proteins were visualized using a donkey anti-rabbit secondary antibody conjugated to HRP and a chemiluminescence detection system.
Arrow indicates Upf1 (~125 kDa) (Please see figures).

Research Category
Epigenetics & Nuclear Function

Research Sub Category
RNA Binding Protein (RBP)

This RIPAb+ Upf1 -RIP Validated Antibody & Primer Set conveniently includes the antibody & the specific control PCR primers.

Packaging

10 assays per set. Recommended use: ~7 μL of antibody per RIP (dependent upon biological context).

Quality

RNA Binding Protein Immunoprecipitation:
RIP Lysate prepared from HeLa cells (2 X 10E7 cell equivalents per IP) were subjected to immunoprecipitation using either 7 µL of a normal rabbit IgG or 7 µL Anti-Upf1 antibody and the Magna RIP® RNA-Binding Protein Immunoprecipitation Kit (Cat. # 17-700).
Successful immunoprecipitation of Upf1-associated RNA was verified by qPCR using RIP Primers ARF1 (Please see figures).
Please refer to the Magna RIP (Cat. # 17-700) or EZ-Magna RIP (Cat. # 17-701) protocol for experimental details.

Target description

~125 kDa

Physical form

Affinity Purfied

Anti-Upf1 (Rabbit Polyclonal). One vial containing 70 μL of purified rabbit polyclonal antibody in buffer containing 0.1 M Tris-Glycine (pH 7.4), 150 mM NaCl with 0.05% sodium azide and 30% glycerol. Store at -20°C.

Normal Rabbit IgG. One vial containing 125 µg of rabbit IgG in 125 µL of storage buffer containing 0.05% sodium azide. Store at -20°C.

RIP Primers, ARF1. One vial containing 75 μL of 5 μM of each primer specific for human ARF1.
Store at -20°C.
FOR: GAC CAC GAT CCT CTA CAA GC
REV: TCC CAC ACA GTG AAG CTG ATG

Format: Purified

Storage and Stability

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.

Analysis Note

Control
Includes negative control normal rabbit IgG antibody and control primers specific for the cDNA of human ARF1.

Legal Information

MAGNA RIP is a registered trademark of Merck KGaA, Darmstadt, Germany

UPSTATE is a registered trademark of Merck KGaA, Darmstadt, Germany

Disclaimer

Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.

Safety & Documentation

Safety Information

Safety Information for this product is unavailable at this time.
Protocols & Articles

Articles

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Peer-Reviewed Papers
15

References

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