Anti-ATP5A antibody [15H4C4] - Mitochondrial Marker

• 产品名称

  Anti-ATP5A抗体[15H4C4] - Mitochondrial Marker
  参阅全部 ATP5A 一抗

• 描述

  小鼠单克隆抗体[15H4C4] to ATP5A - Mitochondrial Marker

• 宿主

  Mouse

• 经测试应用

  适用于: WB, IHC-P, ICC/IF, Flow Cytmore details

• 种属反应性

  与反应: Mouse, Rat, Cow, Human, Drosophila melanogaster
  预测可用于: Pig

• 免疫原

  Tissue, cells or virus. This information is considered to be commercially sensitive.

• 阳性对照

• WB: Isolated mitochondria from human, cow, rat and mouse heart. Human liver tissue lysate. HepG2 whole cell lysate. ICC/IF: HeLa, MCF7 and MDA-MB-231 cells. IHC-P: Human heart tissue. Flow Cyt: HepG2 cells.

• 常规说明

  
  

  This antibody clone is manufactured by Abcam. If you require a custom buffer formulation or conjugation for your experiments, please contact orders@abcam.com.

  
  

  The Life Science industry has been in the grips of a reproducibility crisis for a number of years. Abcam is leading the way in addressing this with our range of recombinant monoclonal antibodies and knockout edited cell lines for gold-standard validation. Please check that this product meets your needs before purchasing.

  If you have any questions, special requirements or concerns, please send us an inquiry and/or contact our Support team ahead of purchase. Recommended alternatives for this product can be found below, along with publications, customer reviews and Q&As

• 形式

  Liquid

• 存放说明

  Shipped at 4°C. Store at +4°C.

• 存储溶液

  pH: 7.5
  Preservative: 0.02% Sodium azide
  Constituent: HEPES buffered saline

• 纯度

  IgG fraction

• 纯化说明

  Near homogeneity as judged by SDS-PAGE. The antibody was produced in vitro using hybridomas grown in serum-free medium, and then purified by biochemical fractionation.

• 克隆

  单克隆

• 克隆编号

  15H4C4

• 同种型

  IgG2b

• 轻链类型

  kappa

The Abpromise guarantee

Abpromise™承诺保证使用ab14748于以下的经测试应用

“应用说明”部分 下显示的仅为推荐的起始稀释度；实际最佳的稀释度/浓度应由使用者检定。

• 数据库链接

  hide

• SwissProt: P35381 Drosophila melanogaster

• SwissProt: P25705 Human

• SwissProt: Q03265 Mouse

• SwissProt: P80021 Pig

• SwissProt: P15999 Rat

• Unigene: 7300 Drosophila melanogaster

• Unigene: 298280 Human

• Unigene: 276137 Mouse

• Unigene: 474505 Mouse

• Unigene: 40255 Rat

• Entrez Gene: 282578 Cow

• Entrez Gene: 37617 Drosophila melanogaster

• Entrez Gene: 498 Human

• Entrez Gene: 11946 Mouse

• Entrez Gene: 100157880 Pig

• Entrez Gene: 65262 Rat

• Omim: 164360 Human

• SwissProt: P19483 Cow

• 别名

• ATP synthase subunit alpha mitochondrial antibody

• ATP synthase, H+ transporting, mitochondrial F1 complex, alpha subunit 1, cardiac muscle antibody

• ATP synthase, H+ transporting, mitochondrial F1 complex, alpha subunit, 1 antibody

• ATP synthase, H+ transporting, mitochondrial F1 complex, alpha subunit, isoform 1, cardiac muscle antibody

• ATP synthase, H+ transporting, mitochondrial F1 complex, alpha subunit, isoform 2, non-cardiac muscle-like 2 antibody

• ATP sythase (F1 ATPase) alpha subunit antibody

• ATP5A antibody

• Atp5a1 antibody

• ATP5AL2 antibody

• ATPA_HUMAN antibody

• ATPM antibody

• Epididymis secretory sperm binding protein Li 123m antibody

• hATP1 antibody

• HEL-S-123m antibody

• MC5DN4 antibody

• mitochondrial antibody

• Mitochondrial ATP synthetase antibody

• Mitochondrial ATP synthetase oligomycin resistant antibody

• Modifier of Min 2 antibody

• Modifier of Min 2 mouse homolog antibody

• Modifier of Min 2, mouse, homolog of antibody

• MOM2 antibody

• OMR antibody

• ORM antibody

• OTTHUMP00000163475 antibody

• ATP synthase alpha chain antibody

• ATP synthase alpha chain, mitochondrial antibody

• ATP synthase subunit alpha antibody

• 

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ATP5A antibody [15H4C4] - Mitochondrial Marker (ab14748)

  Immunohistochemical analysis of formalin fixed paraffin embedded human liver labelling ATP5A with ab14748 at a concentration of 0.01µg/ml.

  The immunostaining was performed on a Ventana DISCOVERY ULTRA (Roche Tissue Diagnostics) instrument with an OptiView DAB IHC Detection Kit. Heat mediated antigen retrieval was conducted for 32min with DISCOVERY cell conditioning solution (CC1) 100°C, pH8.5. ab14748 anti-ATP5A was incubated at 37°C for 16min.

  Sections were counterstained is with Hematoxylin II. Image inset shows absence of staining in secondary antibody only control.

  Customers are encouraged to optimise antigen retrieval conditions, antibody concentration, incubation times and temperature for best results in their own IHC assay workflow (automated and manual)

• 

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ATP5A antibody [15H4C4] - Mitochondrial Marker (ab14748)

  Immunohistochemical analysis of formalin fixed paraffin embedded human liver labelling ATP5A with ab14748 at a concentration of 0.05µg/ml. The immunostaining was performed on a Leica Biosystems BOND® RX instrument with a Bond™ Polymer Refine Detection kit. Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution 2) for 20 mins.

  ab14748 anti-ATP5A antibody [15H4C4] was incubated for 15mins at room temperature. Sections were counterstained with Hematoxylin. Image inset shows absence of staining in secondary antibody only control.

  Customers are encouraged to optimise antigen retrieval conditions, antibody concentration, incubation times and temperature for best results in their own IHC assay workflow (automated and manual)

• 

  Immunocytochemistry/ Immunofluorescence - Anti-ATP5A antibody [15H4C4] - Mitochondrial Marker (ab14748)Vedelek et al PLoS One. 2016 Aug 16;11(8):e0161289. doi: 10.1371/journal.pone.0161289. eCollection 2016. Fig 4. Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/

  Testes of bb8ms mutants show defects in post-meiotic, elongated spermatids.

  (A-B) Spermatids from WT (A) and bb8ms (B) testis both have elongated cysts, but there are large spherical vesicles in the mutant (arrows) by phase contrast microscopy. Scale bars: 100 μm.

  (C, D) Mitochondria of elongated spermatids stained with ATP5α antibody ab14748 in WT (C) and in bb8ms (D) mutants. ATP5α positive staining of the large vesicles in the cysts are indicated by arrow. Scale bars: 50 μm.

  (E, F) JC-1 staining positive large vesicles (arrows) are absent from WT (E), but present in bb8ms elongated cysts (F). Scale bars: 25 μm.

• 

  Western blot - Anti-ATP5A antibody [15H4C4] - Mitochondrial Marker (ab14748)Kayser et al PLoS One. 2016 Jan 29;11(1):e0148219. doi: 10.1371/journal.pone.0148219. eCollection 2016. Fig S2. Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/

  Western blots were probed for HNE-damaged mitochondrial protein from brainstem (A), cerebellum (B) and “rest” brain (R). The sample designation indicates the age group (y for P25-P35, o for P45-P55), the genotype (KO, Ctrl for controls) and a number to distinguish independent samples.

  Panel D is the blot from panel A reprobed for the mitochondrial marker ATPase (ATP5a) using ab14748 to demonstrate that extended sample storage did not degrade sample protein in general. Black lines in the MW lanes are magic marker on the film to indicate the positions of the prestained molecular weight standards on the blot.

  For full image please see paper.

• 

  Western blot - Anti-ATP5A antibody [15H4C4] - Mitochondrial Marker (ab14748)

  All lanes : Anti-ATP5A antibody [15H4C4] - Mitochondrial Marker (ab14748) at 1 µg/ml
  
  Lane 1 : Isolated mitochondria from human heart at 10 µg
  Lane 2 : Isolated mitochondria from bovine heart at 4 µg
  Lane 3 : Isolated mitochondria from rat heart at 10 µg
  Lane 4 : Isolated mitochondria from mouse heart at 10 µg
  Lane 5 : HepG2 (Human liver hepatocellular carcinoma cell line) lysate at 20 µg
  

• 

  Western blot - Anti-ATP5A antibody [15H4C4] - Mitochondrial Marker (ab14748)

  All lanes : Anti-ATP5A antibody [15H4C4] - Mitochondrial Marker (ab14748) at 1 µg/ml
  
  Lane 1 : Human liver tissue lysate
  Lane 2 : HepG2 (Human hepatocellular liver carcinoma cell line) whole cell lysate
  
  Lysates/proteins at 10 µg per lane.
  
  Secondary
  All lanes : Goat polyclonal to Mouse IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilution
  
  Observed band size: 55 kDawhy is the actual band size different from the predicted?
  Additional bands at: 36 kDa. We are unsure as to the identity of these extra bands.
  
  

• 

  Immunocytochemistry/ Immunofluorescence - Anti-ATP5A antibody [15H4C4] - Mitochondrial Marker (ab14748)

  ICC/IF image of ab14748 stained HeLa (Human epithelial cell line from cervix adenocarcinoma) cells.

  The cells were fixed in 100% methanol (5 minutes), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1 hour. The cells were then incubated overnight at +4°C with ab14748 at 1 µg/ml (shown in red) and ab6160 (Rat monoclonal to Tubulin) at 1 µg/ml (shown in green).

  This was followed by an incubation at room temperature for 1 hour with ab150119, Goat Anti-Mouse IgG H&L (Alexa Fluor® 647) preadsorbed, at 0.5 µg/ml (shown in red) and ab150165, Goat Anti-Rat IgG H&L (Alexa Fluor® 488) preadsorbed, at 0.5 µg/ml (shown in green).

  Nuclear DNA was labeled with DAPI (shown in blue).

  Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

  This product also gave a positive signal under the same testing conditions in HeLa cells fixed with 4% formaldehyde (10 minutes).

• 

  Immunocytochemistry/ Immunofluorescence - Anti-ATP5A antibody [15H4C4] - Mitochondrial Marker (ab14748)This image is courtesy of an anonymous Abreview

  ab14748 staining ATP5A in MDA-MB-231 cells by ICC/IF (Immunocytochemistry/immunofluorescence).

  Cells were fixed with formaldehyde, permeabilized with 1% Triton X-100 and blocked with 10% BSA for 1 hour at 21°C. Samples were incubated with primary antibody (1/100 in BSA + 0.02% Tween 20) for 1 hour at 21°C. A DyLight® 550-conjugated goat anti-mouse IgG polyclonal  (1/500) was used as the secondary antibody.

  See Abreview

• 

  Immunocytochemistry/ Immunofluorescence - Anti-ATP5A antibody [15H4C4] - Mitochondrial Marker (ab14748)

  ICC/IF image of ab14748 stained MCF7 (Human breast adenocarcinoma cell line) cells.

  The cells were fixed in 4% formaldehyde (10 minutes) and then incubated in 1% BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1 hour to permeabilize the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab14748, 10 µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-mouse IgG (H+L) used at a 1/1000 dilution for 1 hour. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1 hour. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43 µM.
  
  

• 

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ATP5A antibody [15H4C4] - Mitochondrial Marker (ab14748)

  ab14748 (1 µg/ml) staining ATP5A in human heart (left ventricle), using an automated system (DAKO Autostainer Plus). Using this protocol there is mitochondrial staining of cardiomyocytes.
  Sections were rehydrated and antigen retrieved with the Dako 3 in 1 AR buffer citrate pH 6.1 in a DAKO PT link. Slides were peroxidase blocked in 3% H₂O₂ in methanol for 10 minutes. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 minutes and detected with Dako envision flex amplification kit for 30 minutes. Colorimetric detection was completed with Diaminobenzidine for 5 minutes.

  Slides were counterstained with hematoxylin and coverslipped under DePeX. Please note that, for manual staining, optimization of primary antibody concentration and incubation time is recommended. Signal amplification may be required.

• 

  Flow Cytometry - Anti-ATP5A antibody [15H4C4] - Mitochondrial Marker (ab14748)

  Overlay histogram showing HepG2 (Human liver hepatocellular carcinoma cell line) cells stained with ab14748 (red line).

  The cells were fixed with 4% paraformaldehyde (10 minutes) and then permeabilized with 0.1% PBS-Tween for 20 minutes. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab14748, 1 µg/1x106 cells) for 30 minutes at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 minutes at 22ºC. Isotype control antibody (black line) was mouse IgG2b [PLPV219] (ab91366, 2 µg/1x106 cells) used under the same conditions.

  Acquisition of >5,000 events was performed.

  This antibody gave a positive signal in HepG2 cells fixed with 80% methanol (5 minutes)/permeabilized in 0.1% PBS-Tween used under the same conditions.