Monoclonal Antibody to RASSF5 / NORE1 (119

AM32697PU-N OriGene Technologies Inc. OriGene EU

Acris Antibodies GmbH 9620 Medical Center Drive, Ste 200 Schillerstr. 5 Rockville, MD 20850 32052 Herford UNITED STATES GERMANY Phone: +1-888-267-4436 Phone: +49-5221-34606-0 Fax: +1-301-340-8606 Fax: +49-5221-34606-11 [email protected] [email protected]

Monoclonal Antibody to RASSF5 / NORE1 (119-414) - Purified Alternate names: New ras effector 1, RAPL, Ras association domain-containing protein 5, Regulator for cell adhesion and polarization enriched in lymphoid tissue Catalog No.: AM32697PU-N Quantity: 0.1 mg Background: This gene is a member of the Ras association domain family. It functions as a tumor suppressor, and is inactivated in a variety of cancers. The encoded protein localizes to centrosomes and microtubules, and associates with the GTP-activated forms of Ras, Rap1, and several other Ras-like small GTPases. The protein regulates lymphocyte adhesion and suppresses cell growth in response to activated Rap1 or Ras. Multiple transcript variants encoding different isoforms have been found for this gene. Uniprot ID: Q8WWW0 NCBI: NP_872604.1 GeneID: 83593 Host / Isotype: Mouse / IgG1 Recommended SM10P (for use in human samples), AM03095PU-N Isotype Controls: Clone: 10F10 Immunogen: 6His-tagged fusion protein corresponding to residues 119-414 of Human Nore1A. Format: State: Liquid purified IgG fraction Purification: Protein G Chromatography Buffer System: 0.1M Tris-glycine, pH 7.4, 0.15M NaCl Preservatives: 0.05% Sodium Azide Stabilizers: 30% Glycerol Applications: Immunoblot Analysis: 0.5-2.0 μg/ml of this lot detected Nore1A in RIPA lysates from HaCaT and HeLa cells. Quality Assurance: Routinely evaluated by immunoblot on RIPA lysates from HaCaT and HeLa cells. Not recommended for Immunoprecipitation. Other applications not tested. Optimal dilutions are dependent on conditions and should be determined by the user. Molecular Weight: 45 and 47 kDa Specificity: Recognizes Nore1A, Mr 45 and 47kDa. Multiple bands may be observed due to posttranslational modifications. Species Reactivity: Tested: Human.

For research and in vitro use only. Not for diagnostic or therapeutic work. OG/20130626 Material Safety Datasheets are available at www.acris-antibodies.com or on request. Acris Antibodies is now part of the OriGene family. Learn more at www.origene.com 1 / 2 AM32697PU-N: Monoclonal Antibody to RASSF5 / NORE1 (119-414) - Purified Storage: Store undiluted (in aliquots) at -20°C. Avoid repeated freezing and thawing. Shelf life: one year from despatch. General Readings: 1. Feig LA, Buchsbaum RJ. Cell signaling: life or death decisions of ras proteins. Curr Biol. 2002 Apr 2;12(7):R259-61. PubMed PMID: 11937045. 2. Tommasi S, Dammann R, Jin SG, Zhang XF, Avruch J, Pfeifer GP. RASSF3 and NORE1: identification and cloning of two human homologues of the putative tumor suppressor gene RASSF1. Oncogene. 2002 Apr 18;21(17):2713-20. PubMed PMID: 11965544. 3. Cox AD, Der CJ. The dark side of Ras: regulation of apoptosis. Oncogene. 2003 Dec 8;22(56):8999-9006. PubMed PMID: 14663478. Protocols: Immunoblot Protocol 1. Perform SDS-polyacrylamide gel electrophoresis (SDS-PAGE) on a cell lysate sample (cell lysis buffer: 50mM Tris-HCl, pH 7.4; 1% NP-40; 0.25% sodium deoxycholate; 150mM NaCl; 1mM EDTA; 1mM PMSF; 1μg/ml each aprotinin, leupeptin, pepstatin; 1mM Na3VO4, 1mM NaF) and transfer the proteins to nitrocellulose. Wash the blotted nitrocellulose twice with water. 2. Block the blotted nitrocellulose in freshly prepared 5% nonfat dry milk in TBS with 0.05% Tween®-20 (TBST-MLK) for 20 minutes at room temperature with constant agitation. 3. Incubate the nitrocellulose with 0.5-2.0μg/ml of anti-Nore1A, clone 10F10, diluted in freshly prepared TBST-MLK overnight with agitation at 4°C. 4. Wash the nitrocellulose twice with water. 5. Incubate the nitrocellulose in the secondary reagent of choice (a goat anti-mouse HRP conjugated IgG, 1:5000 dilution was used) in TBST-MLK for 1.5 hours with agitation at room temperature. 6. Wash the nitrocellulose twice with water. 7. Wash the nitrocellulose in TBS-0.05% Tween®-20 for 3-5 minutes. 8. Rinse the nitrocellulose in 4-5 changes of water. 9. Use detection method of choice (Visualizer™ Western Blot Detection Kit was used). Pictures: Immunoblot Analysis HaCaT cell lysate was resolved by electrophoresis, transferred to nitrocellulose and probed with anti-Nore1A, clone 10F10 (0.5 μg/ml). Proteins were visualized using a Goat anti-Mouse secondary antibody conjugated to HRP and the Visualizer™ Western Blot Detection Kit. Arrow indicates Nore1A (~45/47kDa).