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Analysis Note

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

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 Sub CategoryRNA Binding Protein (RBP)

Research CategoryEpigenetics & Nuclear Function

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

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.

General description

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.

Immunogen

KLH-conjugated synthetic linear peptide corresponding to Upf1.

Legal Information

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

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

Packaging

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

Physical form

Affinity purified

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

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.

Specificity

This antibody detects Upf1.

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.

Target description

~125 kDa