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Analysis Note

ControlIncludes negative control mouse IgG antibody and control primers specific for mouse Iqsec1.

Application

Western Blot Analysis:
Representative lot data.
Mouse E17 spinal cord lysate (Lane 1), Mouse E16 brain (Lane 2) and NIH/3T3 lysate (Lane 3) were resolved by electrophoresis, transferred to PVDF membrane and probed with anti-Fragile X Mental Retardation Protein (1:500 dilution of a previous lot).
Proteins were visualized using a goat anti-mouse secondary antibody conjugated to HRP and a chemiluminescence detection system.
Arrow indicates Fragile X Mental Retardation Protein (~71 kDa) (Please see figures).


Immunohistochemistry: Frozen and paraffin sections. 1:500-1:5,000 dilution of a previous lot was used.

ELISA: A 1:500-1:5,000 dilution of a previous lot was used in ELISA.

Detection of FMRP on Blood Smears: A 1:500-1:5,000 dilution of a previous lot was used.

Immunocytochemistry: A 1:500-1:5,000 dilution of a previous lot was used on transfected cells. Light fixation (2% PFA, permeabilize with 0.1% triton in block only) (Please see figures).

Optimal working dilutions must be determined by the end user.

This RIPAb+ Fragile X Mental Retardation Protein -RIP Validated Antibody & Primer Set conveniently includes the antibody & the specific control PCR primers.

Research CategoryEpigenetics & Nuclear FunctionNeuroscience

Research Sub CategoryRNA Binding Protein (RBP)RNA Metabolism & Binding ProteinsNeurodegenerative Diseases

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

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.

Immunogen

Fusion protein with a full length FMRP (human).

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: ~5 µL of antibody per RIP (dependent upon biological context).

Physical form

Anti-Fragile X Mental Retardation Protein (Mouse Monoclonal IgG1κ). One vial containing 50 µL of liquid, does not contain any preservative. Store at -20°C.

Normal Mouse IgG.
One vial containing 125 µg of purified mouse IgG in 125 µL of storage buffer containing 0.1% sodium azide. Store at -20°C.
RIP Primers, Iqsec1.

One vial containing 75 µL of 5 µM of each primer specific for mouse Iqsec1.
Store at -20°C.
FOR: GCG TCT AGG ACC TTG TGA GC
REV: GAA GGC TGA CCT CCA TGT TG

Format: Purified

Quality

RNA Binding Protein Immunoprecipitation:
RIP Lysate prepared from NIH/3T3 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-Fragile X Mental Retardation Protein antibody and the Magna RIP^® RNA-Binding Protein Immunoprecipitation Kit (Cat. # 17-700).
Successful immunoprecipitation of Fragile X Mental Retardation Protein-associated RNA was verified by qPCR using RIP Primers Iqsec1 (Please see figures).
Please refer to the Magna RIP™ (Cat. # 17-700) or EZ-Magna RIP™ (Cat. # 17-701) protocol for experimental details.

Specificity

FMRP. Cross reaction with FXR protein may be detected in cases of high expression in mice.

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.

Target description

71 kDa