03-110

RIPAb+ Ago2 - RIP Validated Antibody and Primer Set

ascites fluid, from mouse

• Synonym(s): Argonaute-2, Eukaryotic translation initiation factor 2C, Slicer protein, Piwi domain protein
• UNSPSC Code: 12352203
• NACRES: NA.52
• eCl@ss: 32160702

Properties

• biological source: mouse
• Quality Segment: 100
• antibody form: ascites fluid
• clone: monoclonal
• species reactivity: human
• species reactivity (predicted by homology): bovine
• manufacturer/tradename: RIPAb+, Upstate^®
• technique(s): RIP: suitable, immunoprecipitation (IP): suitable, western blot: suitable
• isotype: IgG1κ
• NCBI accession no.: NP_001158095.1
• UniProt accession no.: Q9UKV8
• shipped in: dry ice

Description

General description

100 kDa

Ago2 (argonaute-2), also known as eukaryotic translation initiation factor 2C (EIF2C2), is an essential component for siRNA-directed RNA interference (RNAi) response. Ago2 is an endonuclease required for the unwinding of siRNA duplex and assembly of siRNA into RNA-induced silencing complexes (RISC). Ago2 interacts with DICER1 through its Piwi domain. This Piwi domain is thought to provide RNA cleavage activity via a mechanism similar to RNase H. Ago2 activity is necessary for embryonic development as well as RNA-mediated gene silencing (RNAi).

Immunogen

KLH-conjugated, synthetic peptide containing the sequence YSGAGPALAPPAPPPPIQG.

Application

RNA Binding Protein Immunoprecipitation (microRNA):
RIP Lysate prepared from HeLa cells (2 X 10⁷ cell equivalents per IP) were subjected to immunoprecipitation using 5 µL of either a normal mouse IgG or 5 µL of Anti-Ago2 antibody and the Magna RIP^® RNA-Binding Protein Immunoprecipitation Kit (Cat. # 17-700).
Successful immunoprecipitation of Ago2-associated microRNA was verified by TaqMan microRNA assay, hsa-miR-23a. (Please see figures).
Please refer to the Magna RIP (Cat. # 17-700) or EZ-Magna RIP (Cat. # 17-701) protocol for experimental details.


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 µL of either a normal mouse IgG or Anti-Ago2 antibody. Precipitated proteins (lane 1: mouse IgG, lane 2: anti-Ago2) and HeLa whole cell lysate (lane 3) were resolved by electrophoresis, transferred to nitrocellulose and probed with anti-Ago2 antibody (1:5,000 dilution).
Proteins were visualized using goat anti-mouse secondary antibody conjugated to HRP and chemiluminescence detection.
(Please see figures).

Research Category
Epigenetics & Nuclear Function

Research Sub Category
RNA Metabolism & Binding Proteins

RNA Binding Protein (RBP)

This RIPAb+ Ago2 -RIP Validated Antibody & Primer Set conveniently includes the antibody & the specific control PCR primers.

Biochem/physiol Actions

Recognizes Ago2.

Packaging

10 assays per set. Recommended use: ~5 μL of antibody per RIP (dependent upon biological context).

Physical form

Anti-Ago2 (Mouse Monoclonal IgG1κ). One vial containing 50 µL of ascites with 0.05% sodium azide. Store at -20°C.

Normal Mouse IgG. One vial containing 125 μg of purified mouse IgG in 125 µL of stoage buffer containing 0.1% sodium azide. Store at -20°C.

RIP Primers, FOS. One vial containing 75 μL of 5 μM of each primer specific for human FOS gene. 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

Preparation Note

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

Analysis Note

RNA Binding Protein Immunoprecipitation:
RIP Lysate prepared from HeLa cells (2 X 10⁷ cell equivalents per IP) were subjected to immunoprecipitation using 5 µL of either a normal mouse IgG or 5 µL of Anti-Ago2 antibody and the Magna RIP^® RNA-Binding Protein Immunoprecipitation Kit (Cat. # 17-700).
Successful immunoprecipitation of Ago2-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.

Control
Includes negative control mouse IgG antibody and control primers specific for human FOS.

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 Information

• Storage Class: 10 - Combustible liquids
• wgk: WGK 1