製品の概要

  • 製品名
    Total OXPHOS Rodent WB Antibody Cocktail
  • アッセイタイプ
    Quantitative
  • 種交差性
    交差種: Mouse, Rat, Cow, Human, Cynomolgus monkey
  • 製品の概要

    Total OXPHOS Rodent WB Antibody Cocktail ab110413 is an optimized cocktail of high quality antibodies for analyzing relative levels of OXPHOS complexes in rat or mouse mitochondria by western blot. 


    This OXPHOS cocktail contains 5 mouse mAbs, one each against CI subunit NDUFB8 (ab110242), CII-30kDa (ab14714), CIII-Core protein 2 (ab14745) CIV subunit I (ab14705) and CV alpha subunit (ab14748) as an optimized premixed cocktail. The kit is suitable for Western Blotting analysis of the relative levels of the 5 OXPHOS complexes in mitochondrial preparations from mouse, rat, human, or bovine sources.


    Altered levels of assembly can arise from mutations in individual subunits, mutations in assembly factors for the complex(es), mtDNA depletion or as a result of physiological and or pathological changes e.g. hormone treatment, exercise, diet or oxidative stress.


     The mAbs in the cocktail were chosen because they are each against a subunit that is labile when its complex is not assembled. Also, the different subunits are easily resolved in SDS-PAGE (see protocols).


    Note: Mouse tissue samples can easily be contaminated with antibodies from the animal's blood. To avoid background bands, use ab110413 with an anti-mouse secondary against native antibodies. Also, COXI is a highly hydrophobic protein and appears as a broad band at ~35 kDa (not at its true molecular weight at 57 kDa). It is very sensitive to heating. Therefore, the samples, including the positive control, should not be heated over 50°C before loaded on the gel.


    The Western blot cocktail is supplied at a concentration of 1.5 mg/mL.


    We recommend using a high pH CAPS / PVDF transfer protocol when using this antibody for Western blot.

  • 特記事項

    Related products

    Review the mitochondrial assay guide, or the full metabolism assay guide to learn about more assays for metabolites, metabolic enzymes, mitochondrial function, and oxidative stress, and also how to assay metabolic function in live cells using your plate reader.

  • アプリケーション
    適用あり: WBmore details

製品の特性

アプリケーション

Our Abpromise guarantee covers the use of ab110413 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

アプリケーション Abreviews 特記事項
WB Use at an assay dependent concentration.

The antibody cocktail (1.5 mg/mL) should be diluted 250x to a final working concentration of 6.0 µg/mL for Western blotting.

We recommend using PBS with 1% milk as the antibody diluent.

Store the antibody cocktail at 4°C and the control sample at -80°C.

We recommend using a high pH CAPS / PVDF transfer protocol when using this antibody for Western blot. Please contact us for more details.

画像

  • Figure 1. Rat liver mitochondria labeled with ab110413 (MS604). The sample in lane 1 was kept at room temperature, whereas the remaining three samples were heated to 37°C, 50°C, and 100°C, respectively. This blot shows that boiling of samples leads to a decrease in signal due to aggregation of proteins therefore heating samples at or close to boiling is not recommended.

参考文献

This product has been referenced in:
  • Petrick HL & Holloway GP High intensity exercise inhibits carnitine palmitoyltransferase-I sensitivity to L-carnitine. Biochem J N/A:N/A (2019). Read more (PubMed: 30635360) »
  • Pereira SS  et al. MAPK/ERK pathway inhibition is a promising treatment target for adrenocortical tumors. J Cell Biochem 120:894-906 (2019). Read more (PubMed: 30256438) »
See all 432 Publications for this product

レビューと Q&A

1-10 of 41 Abreviews or Q&A

Oxphos Rodent antibody works on Bovine liver

Excellent Excellent 5/5 (Ease of Use)
Abreviews
Bovine liver 20ug of lysates were loaded on to 4-15% gradient gel.
1:4000 dilution of primary antibody used.
Works great!

Julie Kim

Verified customer

投稿 Apr 19 2017

Abreviews
Application: WB
Sample species: Mouse
1) Sample species: Mouse
2) Type: Tissue lysate (Gastrocnemius, skeletal muscle)
3) Amount of protein used: 50ug
4) Specification: Skeletal Muscle
5) Reduce/denaturing condition: SDS sample buffer, no boiling. 4-12% gel (Bis/Tris), MOPS
6) Blocking buffer/concentration/time/temperature:Odyssey Blocking Buffer, Antibody dil. in 4%BSA-TBST/4hrs RT
7) Antibody dilution: 1:1000
8) Incubation time/temperature: 4hrs/RT
9) Diluent: 4% BSA-TBST
10) Secondary antibody information:LICOR/Goat-anti-mouse/1:20,000
11) Detection method: LICOR Odyssey

Dr. Alice Pollard

Verified customer

投稿 Apr 08 2015

Answer

The temperature shown on the WB image is the degrees at which the lysate was denatured prior to loading on the gel (5 min only). We show this as most customers boil the samples before loading. For these antibodies, boiling will actually be counterproductive as the signal disappears as the temperature is increased.

Read More

OXPHOS for Rat endothelial cells

Good Good 4/5 (Ease of Use)
Abreviews
Application: Western blot
Sample: Rat Cell lysate - whole cell (Rat microvascular endothelial cells)- triplicates in lanes 2,3,4.
All blue marker in lane 1.
Loading amount: 20 µg
Specification: Rat microvascular endothelial cells
Gel Running Conditions: Reduced Denaturing (12% gel)
Blocking step: Milk as blocking agent for 24 hours · Concentration: 5 % · Temperature: 4°C
Primary antibody conditions:
Incubation time:
1 hour Temperature: 23°C Diluent: TBS+Tween (0.1%)
Secondary antibody
Name: Non-Abcam antibody was used: Goat Anti-Mouse IgG (H+L)-HRP Conjugate
Conjugation: HRP polymer
Dilution:1/5000
Detection method: ECL
Exposure: 1 minute and 0 seconds
Bands: Specific: 50, 40, 30, 25, 18 Kda

Dr. Dhanush Haspula

Verified customer

投稿 May 07 2019

Abreviews
The molecular weight marker is at 50kDA, 37kDA, 25kDa and 20kDA. In rails 1 to 7 and 9 to 14 are different Rabbit samples (1-3 control rabbit samples) and in rail 8 is the Rat Heart mitochondria WB control. The samples were kept at room temperature (not heated) and a PVDF membrane was used for transfer.

Daniel Jerico

Verified customer

投稿 Apr 12 2019

Total OXPHOS Western Blot C2C12 (mouse)

Excellent Excellent 5/5 (Ease of Use)
Abreviews
Cell line: C2C12 (myoblasts)
Protein loaded 25 ug
Primary antibody dilution: 1:1000
Secondary: 1:10000 (HRP), goat anti-mouse IgG-HRP: sc-2005

Abcam user community

Verified customer

投稿 Nov 21 2018

OXPHOS Antibody on Pig Skeletal Muscle

Average Excellent 5/5 (Ease of Use)
Abreviews
Used Pig Skeletal Muscle preps. Loaded 30ug of protein for pig skeletal muscle samples, and 5uL of the Rat Heart Positive Control onto 5-12% gradient gel. Transferred onto nitrocellulose membrane. Blocked with 1XTBST with 5% nonfat dry milk. Antibody was diluted 1:1000 in 1xTBST. Secondary Goat anti Mouse Secondary Antibody (green). After imaging, only picked up 4 bands total in pig muscle (2 very faint).

Abcam user community

Verified customer

投稿 Oct 02 2018

Answer

The antibodies detect different subunits of cytochrome c oxidase, which is also known as Complex IV, and which is abbreviated as CIV on the ab110413 datasheet (Total OXPHOS Rodent WB Antibody Cocktail).

The CIV (Complex IV) antibody in the panel ab110413 detects Compex IV subunit 1, also called COXI and MT-CO1, at ˜40kDa. This antibody is available separately as ab14705. The COX IV mitochondria loading control antibody ab16056 detects Complex IV subunit 4, also known as COXIV, at about 15kDa. Here are links to the UniProt records for the human proteins.

MT-CO1 https://www.uniprot.org/uniprot/P00395
COXIV https://www.uniprot.org/uniprot/P13073

Read More
Abreviews
Worked great in Western Blot. Clear bands that were easy to interpret.

Abcam user community

Verified customer

投稿 Feb 14 2018

Cell lysate: mouse cardiomyocytes

Excellent Excellent 5/5 (Ease of Use)
Abreviews
Blocking step: Milk as blocking agent for 1 hour · Concentration: 5 % · Temperature: RT°C

Dilution: 1/100

Incubation time: 13 hours Temperature: 4°C Diluent: 1x TBS with 0.1% Blocking

Secondary antibody
Name: anti-mouse Conjugation: HRP
Dilution:1/200

Detection method: ECL chemi method

Abcam user community

Verified customer

投稿 Nov 01 2017

1-10 of 41 Abreviews or Q&A

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