C10orf2 anticorps (Middle Region)

N° du produit ABIN2775251

Cet anticorps Lapin Polyclonal validé indépendamment détecte spécifiquement C10orf2 dans WB. Il présente une réactivité avec des échantillons de Humain, Souris, Rat, Boeuf (Vache), Cobaye, Cheval, Chien, Lapin et Poisson zèbre (Danio rerio). Il a été cité dans 2+ publications.

Aperçu rapide pour C10orf2 anticorps (Middle Region) (ABIN2775251)

Antigène: C10orf2 (C10ORF2) (Chromosome 10 Open Reading Frame 2 (C10ORF2))

Reactivité: Humain, Souris, Rat, Boeuf (Vache), Cobaye, Cheval, Chien, Lapin, Poisson zèbre (Danio rerio)

Hôte: Lapin

Clonalité: Polyclonal

Conjugué: Cet anticorp C10orf2 est non-conjugé

Application: Western Blotting (WB)

Détail du produit anti-C10orf2 anticorps

Épitope: Middle Region

Fonction: TWNK Antibody - middle region

Séquence: GVFRKFATDN NCHVTLVIHP RKEDDDKELQ TASIFGSAKA SQEADNVLIL

Homologie: Cow: 100%, Dog: 100%, Guinea Pig: 100%, Horse: 100%, Human: 100%, Mouse: 100%, Rabbit: 100%, Rat: 100%, Zebrafish: 93%

Attributs du produit: This is a rabbit polyclonal antibody against PEO1. It was validated on Western Blot using a cell lysate as a positive control.

Purification: Affinity Purified

Immunogène: The immunogen is a synthetic peptide directed towards the middle region of human PEO1

Information d'application

Indications d'application: Optimal working dilution should be determined by the investigator.

Commentaires:

We also sell a specific blocking peptide that can be used in combination with this antibody. You can find the blocking peptide under AAP36483-100UG

Restrictions: For Research Use only

Independent Validation (WB)

Validation #100038 (Western Blotting)

by: Mitochondrial Biology, University of Eastern Finland

No.: #100038

Date: 08.08.2016

Antigène: PEO1 antibody - middle region

Numéro du lot: QC6322

Application validée: Western Blotting

Contrôle positif: HeLa cells, HeLa cells overexpressing PEO1

Contrôle négative: rho0 HeLa cells

Conclusion: Passed, regarding sensitivity and specificity. The antibody detects an antigen at the expected apparent molecular weight of its antigen.

Validation Images

75µg total protein of HeLa cells overexpressing PEO1 (lane 1), HeLa cells (lane 2), and rho0 Hela cells (lane 3) were separated on an denaturing 8% polyacrylamid gel. Bands were revealed as described in the protocol sections with ABIN2775251 diluted 1:1000.

Protocole

Anticorps primaire: ABIN2775251

Anticorps secondaire: Goat-anti rabbit IgG (H+L), HRP-linked (Invitrogen, A-16104)

Full Protocol:

• Cell lysates were propared from HeLa cells, HeLa cells overexpressing PEO1, and rho⁰ HeLa cells.
• 75µg total protein or each sample were separated on a denaturing 8% SDS-PAGE gel (Laemmli 1970).
• Immunoblot onto supported nitrocellulose membrane (Protran, GE Healthcare No. 1060005, 0.2µm) (Towbin et al., 1979).
• Blocking of the membrane in 3% skim milk in TBST (50mM Tris-HCl, pH 7.4, 150mM NaCl, 0.1% Tween 20) for 1h at room temperature.
• Incubation with primary antibody ABIN2775251 diluted 1:1000 in 3% BSA in TBST at 4°C overnight.
• Washing in TBST for 3x 5min.
• Incubation with goat-anti rabbit IgG (H+L), HRP-linked (Invitrogen, A-16104) diluted 1:15000 in 3% BSA in TBST for 1h at room temperature.
• Washing in TBST for 5x 10min.
• Chemiluminescence detection with luminol solution (250µg/ml Na-Luminol, 0,01% H₂O₂, 11µg/ml p-hydroxy-coumaric acid in 100mM Tris pH 8.5) and image capture on Kodak ECL film.

Notes: PEO1 runs usually at 95-100 kDa, although the theoretical size is 72 kDa, often it is also visible as dimer.

Independent Validation (ICC)

Validation #100039 (Immunocytochemistry)

by: Mitochondrial Biology, University of Eastern Finland

No.: #100039

Date: 08.08.2016

Antigène: PEO1 antibody - middle region

Numéro du lot: QC6322

Application validée: Immunocytochemistry

Contrôle positif: Mouse-anti-ds DNA antibody (clone 35I9)

Conclusion: Passed, regarding sensitivity and specificity. The antibody shows a staining pattern consistent with the expected mitochondrial localization of PEO1.

Validation Images

ABIN2775251 was used at a 1:200 dilution to visualize PEO1 (green, secondary antibody AF488 conjugate). A dsDNA antibody (red, secondary antibody AF594 conjugate) and a DAPI counterstain (blue) were used as reference.

Protocole

Anticorps primaire: ABIN2775251

Anticorps secondaire: Goat-anti-Rabbit IgG (H+L) Secondary Antibody, Alexa Fluor 488 conjugate (Invitrogen, A-11034, Lot 702323)

Full Protocol:

• HeLa cells were grown in DMEM high glucose + 10% FBS on acid-cleaned glass coverslips to 50% confluency.
• Cells were fixed in 4% para-formaldehyde in growth medium for 25min at RT, then washed once with growth medium and twice with PBS.
• Permeabilization was achieved by incubation with 0.5% Triton X-100, 10% fetal bovine serum in PBS for 15min.
• Primary antibodies:
• • ABIN2775251 diluted 1:200 in PBS with 0.1% Triton X-100 , 10% fetal bovine serum.
  • Co-staining of mitochondrial DNA with monoclonal antibody mouse-anti-dsDNA clone 35I9 DNA (Abcam ab27156, Lot GR27035-10) diluted 1:400 in PBS with 0.1% Triton X-100, 10% fetal bovine serum.
• Incubation for 1h at RT.
• Wash 3x 5min with PBS.
• Secondary antibodies:
• • Goat anti-Rabbit IgG (H+L) Secondary Antibody, Alexa Fluor 488 conjugate (Invitrogen, A-11034, Lot 702323) diluted 1:1000 in PBS with 0,1% Triton X-100 , 10% fetal bovine serum, 1µM DAPI.
  • Goat anti-Mouse IgG (H+L) Secondary Antibody, Alexa Fluor 594 conjugate (Invitrogen, A-11032, Lot 621333).
• Incubation for 1h at RT.
• Wash 3x 5min with PBS.
• The coverslips were mounted with Immu-mount mounting medium (Thermo Fisher, 9990402) and imaged with a 630 x magnification on a Zeiss Axioplan 2 fluorescent microscope using an Axiocam camera and Axiovision 4.8 software.
• For visualization of PEO1 the excitation wavelength was 488nm and emission wavelength >535nm, exposure time 53ms. DNA was visualized by excitation at 580nm and emission at >590nm, exposure time 410ms.

Stockage

Format: Liquid

Concentration: 0.5 mg/mL

Buffer: Purified antibody supplied in 1x PBS buffer with 0.09 % (w/v) sodium azide and 2 % sucrose.

Agent conservateur: Sodium azide

Précaution d'utilisation: This product contains Sodium azide: a POISONOUS AND HAZARDOUS SUBSTANCE which should be handled by trained staff only.

Conseil sur la manipulation: prevent freeze-thaw cycles

Stock: 4 °C,-20 °C

Stockage commentaire: For short term use, store at 2-8C up to 1 week. For long term storage, store at -20 °C in small aliquots to prevent freeze-thaw cycles.

Références pour anticorps anti-C10orf2 (ABIN2775251)

Wang, Xu, Alontaga, Chen, Liu: "Impaired p32 regulation caused by the lymphoma-prone RECQ4 mutation drives mitochondrial dysfunction." dans: Cell reports, Vol. 7, Issue 3, pp. 848-58, (2014) (PubMed).

Ylikallio, Tyynismaa, Tsutsui, Ide, Suomalainen: "High mitochondrial DNA copy number has detrimental effects in mice." dans: Human molecular genetics, Vol. 19, Issue 13, pp. 2695-705, (2010) (PubMed).

Détails sur C10orf2

Antigène: C10orf2 (C10ORF2) (Chromosome 10 Open Reading Frame 2 (C10ORF2))

Autre désignation: TWNK

Sujet:

Background Information: This gene encodes a hexameric DNA helicase which unwinds short stretches of double-stranded DNA in the 5' to 3' direction and, along with mitochondrial single-stranded DNA binding protein and mtDNA polymerase gamma, is thought to play a key role in mtDNA replication. The protein localizes to the mitochondrial matrix and mitochondrial nucleoids. Mutations in this gene cause infantile onset spinocerebellar ataxia (IOSCA) and progressive external ophthalmoplegia (PEO) and are also associated with several mitochondrial depletion syndromes. Alternative splicing results in multiple transcript variants encoding distinct isoforms.

Gene Name: twinkle mtDNA helicase

Alternative Symbols: PEO, PEO1, SCA8, ATXN8, IOSCA, PEOA3, SANDO, TWINL, MTDPS7, PRLTS5, C10orf2

Protein Name: twinkle protein, mitochondrial

Poids moléculaire: 77 kDa

ID gène: 56652

NCBI Accession: NP_068602

UniProt: Q96RR1