RIPAb+ EED - RIP Validated Antibody and Primer Set

Polycomb group proteins are important for maintaining transcriptional silencing. One conserved PcG complex, PRC2, is composed of several proteins including the histone methyltransferase EZH2, the WD-repeat protein EED (Embryonic ectoderm development), and the Zn-finger protein Suz12. Transcriptional repression mediated by EED involves histone deacetylation, while the EZH2 methylates histone H3 on lysine 27. EED protein is present in four isoforms. These EED isoforms selectively associate with distinct EZH2-containing complexes, resulting in differential targeting of their associated methyltransferase activity. These complexes play a role in Hox gene silencing, X-inactivation, germline development, stem cell pluripotency and cancer metastasis.

Not tested in other species.

This antibody recognizes mouse EED, (~30-65 kDa).

Recombinant fusion protein of full length murine EED tagged with MBP.

Immunoprecipitation from RIP lysate:
Representative lot data.
RIP lysate from HeLa cells (~2 X 10E7 cell equivalents per IP) was subjected to immunoprecipitation using 5 µg of either a normal mouse IgG, (Cat. #CS200621), or 5 µg of Anti-EED antibody (Cat. # CS204372). Ten percent of the precipitated proteins (lane 1: mouse IgG, lane 3: EED) and HeLa whole cell lysate (lane 2) were resolved by electrophoresis, transferred to nitrocellulose and probed with Anti-EED antibody (Cat. # CS204372, 1:5000). Proteins were visualized using One-Step IP-Western kit (GenScript Cat. # L00232).
Arrow indicates EED.
Western Blot Analysis:
Representative lot data.
A431 cell lysate was resolved by electrophoresis, transferred to nitrocellulose and probed with Anti-EED (1:5,000 dilution). Proteins were visualized using a goat anti-mouse IgG conjugated to HRP and a chemiluminescence detection system.
Arrow indicates EED (~50 kDa).

Research Category
Epigenetics & Nuclear Function

Apoptosis & Cancer

Research Sub Category
Chromatin Biology

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

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

RNA Binding Protein Immunoprecipitation: Representative lot data.
RIP Lysate prepared from HeLa cells (2 X 10E7 cell equivalents per IP) were subjected to immunoprecipitation using 5 µg of either a normal mouse IgG or 5 µg of Anti-EED antibody and the Magna RIP^® RNA-Binding Protein Immunoprecipitation Kit (Cat. # 17-700).
Successful immunoprecipitation of EED-associated RNA was verified by qPCR using RIP Primers lincSFPQ.
Please refer to the Magna RIP (Cat. # 17-700) or EZ-Magna RIP (Cat. # 17-701) protocol for experimental details.

Ranges from 30-65 kDa (5-6 isoforms of EED are detected).

Protein G Purified

Anti-EED (Mouse Monoclonal). One vial containing 50 μg of protein G purified antibody in 65 μL of 0.1 M Tris-Glycine (pH7.4) 150 mM NaCl, containing 0.05% azide before the addition of 30% glycerol. 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, lincSFPQ.
One vial containing 75 μL of 5 μM of each primer specific for the human lincRNA SFPQ. Store at -20°C.
FOR: TCG TAC TGT TAG GCC CTT GG
REV: AAC CTT GCA TGA AGA GCA CC

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: Variabillity in freezer temperatures below -20°C may cause glycerol containing solutions to become frozen during storage.

Control
Includes negative control normal mouse IgG antibody and control primers specific for the cDNA of human lincRNA SFPQ.

Concentration: Please refer to the Certificate of Analysis for the lot-specific concentration.

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

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

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.