General description

All ChIPAb+ antibodies are individually validated for chromatin precipitation, every lot, every time. Each ChIPAb+ antibody set includes control primers (tested every lot by qPCR) to biologically validate your IP results in a locus-specific context. The qPCR protocol and primer sequences are provided, allowing researchers to validate ChIP protocols when using our antibody in their chromatin context. Each set also includes a negative control antibody to ensure specificity of the ChIP reaction.

The ChIPAb+ REST set includes the REST antibody, a Normal Rabbit IgG, and control primers which amplify a 89 bp region of the human Snap25 promoter. The REST and negative controls are supplied in a scalable "per ChIP" reaction size and can be used to functionally validate the precipitation of REST associated chromatin.

Specificity

Recognizes REST, MW 125kDa predicted, 200 kDa observed.

Immunogen

Epitope: a.a. 801-1097

The REST purified antibody is made against a GST fusion protein corresponding to amino acids 801-1097 of human REST.

Application

Chromatin Immunoprecipitation:
Sonicated chromatin prepared from HeLa cells (4e6 cell equivalents per IP) were subjected to chromatin immunoprecipitation using 4 µg of either Normal Rabbit IgG (Part No. PP64B), or 4 µg Anti-REST (Part No.CS207378) and the Magna ChIP HiSens (Cat. # 17-10460). Successful immunoprecipitation of REST associated DNA fragments was verified by qPCR using ChIP Primers, human Snap25 (Part No. CS207379) as a positive locus, and CDC25 as a negative locus. (Figure 2). Data is presented as percent input of each IP sample relative to input chromatin for each amplicon and ChIP sample as indicated.

Western Blotting Analysis:
Human HEK293 cell lysate (30 ug) was resolved by electrophoresis, transferred to PVDF membrane and probed with the anti-REST (1:1000 dilution). Proteins were visualized using a secondary antibody conjugated to HRP and a chemiluminescence detection system. Data courtesy of Glen Corson laboratory of Gail Mandel, Oregon Health and Sciences University, Portland, OR.
REST has a predicted molecular weight of 125 kDa. Observed size was ~200 kDa.

Research Category
Epigenetics & Nuclear Function

Research Sub Category
Chromatin Biology

Histones

This ChIPAb+ REST -ChIP Validated Antibody & Primer Set conveniently includes the antibody & the specific control PCR primers.

Packaging

25 assays per set. Recommended use: ~2 μg antibody per chromatin immunoprecipitation (dependent upon biological context).

Quality

Target description

120 kDa predicted, 200 kDa observed

Physical form

Anti-REST (Rabbit Polyclonal). One vial containing 100 µL 1.0 mg/mL purified rabbit polyclonal in buffer containing 0.014 M phosphate buffer, pH 7.6, 0.175 M NaCl, 0.07% 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.

ChIP Primers, human Snap25. One vial containing 75 μL of 5 μM of each primer specific for human Snap25 promoter. Store at -20°C.
FOR: AGA CTC CTT TGC AGA CAA TTT CCT
REV: CAA CAC AGA AGA TTT CCA CAA GTA GAC

Format: Purified

Storage and Stability

Stable for 1 year at -20°C from date of receipt. Handling Recommendations: Upon first thaw, 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

Control
Includes normal rabbit IgG and primers specific for human Snap25.

Legal Information

MAGNA CHIP 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.

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