C10orf2 Antikörper (Middle Region)

Produktnummer ABIN2775251

Dieser unabhängig validierte Kaninchen Polyklonal Antikörper detektiert spezifisch C10orf2 in WB. Es zeigt Reaktivität gegenüber Proben von Human, Maus, Ratte, Rind (Kuh), Meerschweinchen, Pferd, Hund, Kaninchen und Zebrafisch (Danio rerio). Es wurde in 2+ Publikationen zitiert.

Kurzübersicht für C10orf2 Antikörper (Middle Region) (ABIN2775251)

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

Reaktivität: Human, Maus, Ratte, Rind (Kuh), Meerschweinchen, Pferd, Hund, Kaninchen, Zebrafisch (Danio rerio)

Wirt: Kaninchen

Klonalität: Polyklonal

Konjugat: Dieser C10orf2 Antikörper ist unkonjugiert

Applikation: Western Blotting (WB)

Produktdetails anti-C10orf2 Antikörper

Bindungsspezifität: Middle Region

Verwendungszweck: TWNK Antibody - middle region

Sequenz: 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%

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

Aufreinigung: Affinity Purified

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

Anwendungsinformationen

Applikationshinweise: Optimal working dilution should be determined by the investigator.

Kommentare:

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

Beschränkungen: Nur für Forschungszwecke einsetzbar

Independent Validation (WB)

Validierung #100038 (Western Blotting)

by: Mitochondrial Biology, University of Eastern Finland

No.: #100038

Datum: 08.08.2016

Antigen: PEO1 antibody - middle region

Chargennummer: QC6322

Validierte Anwendung: Western Blotting

Positivkontrolle: HeLa cells, HeLa cells overexpressing PEO1

Negativkontrolle: rho0 HeLa cells

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

Validierungsbilder

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.

Protokoll

Primärantikörper: ABIN2775251

Sekundärantikörper: 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.

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

Independent Validation (ICC)

Validierung #100039 (Immunocytochemistry)

by: Mitochondrial Biology, University of Eastern Finland

No.: #100039

Datum: 08.08.2016

Antigen: PEO1 antibody - middle region

Chargennummer: QC6322

Validierte Anwendung: Immunocytochemistry

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

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

Validierungsbilder

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.

Protokoll

Primärantikörper: ABIN2775251

Sekundärantikörper: 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.

Handhabung

Format: Liquid

Konzentration: 0.5 mg/mL

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

Konservierungsmittel: Sodium azide

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

Handhabung: prevent freeze-thaw cycles

Lagerung: 4 °C,-20 °C

Informationen zur Lagerung: 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.

Referenzen für anti-C10orf2 Antikörper (ABIN2775251)

Wang, Xu, Alontaga, Chen, Liu: "Impaired p32 regulation caused by the lymphoma-prone RECQ4 mutation drives mitochondrial dysfunction." in: 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." in: Human molecular genetics, Vol. 19, Issue 13, pp. 2695-705, (2010) (PubMed).

Details zu C10orf2

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

Andere Bezeichnung: TWNK

Hintergrund:

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

Molekulargewicht: 77 kDa

Gen-ID: 56652

NCBI Accession: NP_068602

UniProt: Q96RR1