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Anti-Complex I Immunocapture antibody [18G12BC2] (ab109798)

Submit a review Q&A (8)References (16)
Immunoprecipitation - Anti-Complex I Immunocapture antibody [18G12BC2] (ab109798)
  • Immunocytochemistry/ Immunofluorescence - Anti-Complex I Immunocapture antibody [18G12BC2] (ab109798)
  • Flow Cytometry - Anti-Complex I Immunocapture antibody [18G12BC2] (ab109798)
  • Flow Cytometry - Anti-Complex I Immunocapture antibody [18G12BC2] (ab109798)

Key features and details

  • Mouse monoclonal [18G12BC2] to Complex I Immunocapture
  • Suitable for: ICC/IF, Flow Cyt, IP
  • Reacts with: Mouse, Rat, Cow, Human
  • Isotype: IgG2b

Overview

  • Product name

    Anti-Complex I Immunocapture antibody [18G12BC2]

  • Description

    Mouse monoclonal [18G12BC2] to Complex I Immunocapture

  • Host species

    Mouse

  • Tested applications

    Suitable for: ICC/IF, Flow Cyt, IPmore details

  • Species reactivity

    Reacts with: Mouse, Rat, Cow, Human

  • Immunogen

    Full length protein. This information is proprietary to Abcam and/or its suppliers.

  • Positive control

    • Cow heart tissue lysate - mitochondrial extract (ab110338) can be used as a positive control in WB. fibroblasts, HL-60 cells, tissue mitochondria

  • General notes

    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

    Product was previously marketed under the MitoSciences sub-brand.

Properties

  • Form

    Liquid

  • Storage instructions

    Shipped at 4°C. Store at +4°C. Do Not Freeze.

  • Storage buffer

    pH: 7.5
    Preservative: 0.02% Sodium azide
    Constituent: 99% HEPES buffered saline
  • Concentration information loading...

  • Purity

    IgG fraction

  • Purification notes

    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.

  • Clonality

    Monoclonal

  • Clone number

    18G12BC2

  • Isotype

    IgG2b

  • Light chain type

    kappa

  • Research areas

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab109798 in the following tested applications.

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

Application Abreviews Notes
ICC/IF
Use at an assay dependent concentration.
Flow Cyt
Use at an assay dependent concentration.
IP
Use a concentration of 0.1 - 1 mg/ml.

100 µg mAb can capture at least 25 µg complex I from 1 mg solubilized bovine heart mitochondria.
Notes
ICC/IF
Use at an assay dependent concentration.
Flow Cyt
Use at an assay dependent concentration.
IP
Use a concentration of 0.1 - 1 mg/ml.

100 µg mAb can capture at least 25 µg complex I from 1 mg solubilized bovine heart mitochondria.

Target

  • Relevance

    Complex I, or NADH ubiquinone oxidoreductase, is a large protein complex of 950,000 Da molecular weight made up by 45 to 46 different subunits. A total of seven of the subunits of the complex are encoded by mitochondrial DNA, while the remainder subunits are nuclear encoded, which are translated in the cytosol and translocated into the organelle for assembly at the inner membrane. The enzyme complex catalyses electron entry from NADH via a flavin (FMN) and several non-heme iron centers. Complex I is sensitive to a wide range of inhibitors, many of which are pesticides or other common environmental toxins, such as rotenone. Complex I dysfunction is a common cause of genetic OXPHOS defects. Altered functioning of this complex is also thought to contribute to several neurological disorders including Parkinson’s disease and schizophrenia. Also, there is evidence of Complex I involvement in diabetes.

Images

  • Immunoprecipitation - Anti-Complex I Immunocapture antibody [18G12BC2] (ab109798)
    Immunoprecipitation - Anti-Complex I Immunocapture antibody [18G12BC2] (ab109798)

    Complex I was immunopurified from mitochondria isolated from human heart (HHM), cow/bovine heart (BHM), mouse heart (MHM) and mouse brain (MBM).  The lanes were stained with Coomassie Brilliant Blue R. Bands were excised from the gel and proteolytically digested for mass spectrometry analysis. For the immuno-isolation, 50 μg of mAb (18G12BC2 ab109798) was bound to 5 μl of swollen protein G agarose beads according to protocol described here.

  • Immunocytochemistry/ Immunofluorescence - Anti-Complex I Immunocapture antibody [18G12BC2] (ab109798)
    Immunocytochemistry/ Immunofluorescence - Anti-Complex I Immunocapture antibody [18G12BC2] (ab109798)
    Immunocytochemistry image of ab109798 stained fibroblasts cells. The cells were paraformaldehyde fixed (4%, 20 minutes) and Triton X-100 permeabilized (0.1%, 15 minutes). The cells were incubated with the antibody (ab109798, 1 µg/mL) for 2 hours at room temperature or over night at 4°C. The secondary antibody was (green) Alexa Fluor® 4884 goat anti-mouse IgG (H+L) at a 1/1000 dilution for 1 hour. 10% Goat serum was used as the blocking agent for all blocking steps. The target protein locates to the mitochondria.
  • Flow Cytometry - Anti-Complex I Immunocapture antibody [18G12BC2] (ab109798)
    Flow Cytometry - Anti-Complex I Immunocapture antibody [18G12BC2] (ab109798)
    HL-60 cells were stained with 1 µg/mL Complex I antibody ab109798 (blue) or an equal amount of an isotype control antibody (red) and analyzed by flow cytometry.
  • Flow Cytometry - Anti-Complex I Immunocapture antibody [18G12BC2] (ab109798)
    Flow Cytometry - Anti-Complex I Immunocapture antibody [18G12BC2] (ab109798)

    Overlay histogram showing HepG2 cells stained with ab109798 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab109798, 2µg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG2b [PLPV219] (ab91366, 2µg/1x106 cells ) used under the same conditions. Acquisition of >5,000 events was performed.

References (16)

Publishing research using ab109798? Please let us know so that we can cite the reference in this datasheet.

ab109798 has been referenced in 16 publications.

  • Amoedo ND  et al. Targeting the mitochondrial trifunctional protein restrains tumor growth in oxidative lung carcinomas. J Clin Invest 131:N/A (2021). PubMed: 33393495
  • Bogenhagen DF & Haley JD Pulse-chase SILAC-based analyses reveal selective oversynthesis and rapid turnover of mitochondrial protein components of respiratory complexes. J Biol Chem 295:2544-2554 (2020). PubMed: 31974161
  • Abuaita BH  et al. Mitochondria-Derived Vesicles Deliver Antimicrobial Reactive Oxygen Species to Control Phagosome-Localized Staphylococcus aureus. Cell Host Microbe 24:625-636.e5 (2018). PubMed: 30449314
  • Inoue N  et al. Knockdown of the mitochondria-localized protein p13 protects against experimental parkinsonism. EMBO Rep 19:N/A (2018). PubMed: 29371327
  • Zhang Y  et al. Lysine desuccinylase SIRT5 binds to cardiolipin and regulates the electron transport chain. J Biol Chem 292:10239-10249 (2017). PubMed: 28458255
View all Publications for this product

Customer reviews and Q&As

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Submit a review Submit a question

1-8 of 8 Abreviews or Q&A

Question

How was the blot on the ab109798 datasheet stained?

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Answer

The image is not of a blot but of a gel stained for total protein. The annotations refer to subunits of mouse brain mitochondria Complex I, identified by mass spec as described in the reference at this link: http://www.mcponline.org/content/4/1/84.full.pdf+html .

Schilling B et al. Rapid purification and mass spectrometric characterization of mitochondrial NADH dehydrogenase (Complex I) from rodent brain and a dopaminergic neuronal cell line. Mol Cell Proteomics 4:84-96 (2005).

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Question

I wonder if you could answer some questions regarding your inmunocapture antibody for mitochondrial complex I (ab109798). I will be interested in using it for other uses besides inmunoprecipitation (western blotting).

Would it be possible to know which subunit inside the complex is recognizing?

Do you know if it has been tested for its use in BN-PAGE?

Many thanks for your help,

Best regards,

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Answer

This antibody is an immunocapture antibody which can pull down the whole complex I from a given sample. It works also as a ICC and Flow antibody. It's a weak WB antibody, light up a 8kda subunit of complex I but we have difficulty to identify which subunit it exactly targets on. It has not been tested for its use in BN-PAGE. For BN-PAGE we highly recommend to use ab110412.

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Question

What is the immunogen used to produce this antibody?

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Answer

whole (45 subunit, 900 kDa) active bovine Complex I - this was shown with mass spec.

It is excellent at binding native, active complex I but has proven difficult to identify the exact subunit to which it binds.

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Question

Is the S1 subunit in the western blot image on the webpage, known by any other names?

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Answer

Thank you for contacting Abcam.

I have attached a paper that looks at the structure of complex 1 and Table 1 has a complete list of the subunits and I believe (but I am no expert in this field) that S1 is also known as NDUFS1.

Please let me know if there is anything else I can help you with.

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Question

Is this antibody is suitable to capture the complex I in NSL melamide(non sialated alkaloid)treated mouse heart tissue lysates?

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Answer

Thank you very much for your phone call.
I have had discussion with my colleague in lab. They have suggested that the antibody was tested only with lysates and cells as presented on the datasheet. No other tested data is available for this product.
I am sorry we are unable to provide further details. The antibody is fully characterized and is guaranteed to for IP, ICC, In-Cell ELISA, Flow Cyt irrespective of protocols used.
I hope this information will be helpful. Should you have any other inquiry please do not hesitate to ask.

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Question

Thank you very much for your warmhearted reply.  Cat. No. of  antibody I bought from your company is MS101c. I have seen a paper published by Sashi Nadanaciva which use  MS101c as an antibody to capture complex I from bovine heart mitochondria for measurement of complex I activity. Our aim is to detect complex I activity in blood cells from different patients and hope  to find those with complex I gene dificiency. I suspect complex I activity captured by MS101c is rotenone-insensitive. Could you please tell me why complex I has two different activities, namely rotenone-sensitive and rotenone-insensitive.  By the way, please tell me whether I can use MS101c as antibody to detect those patients with complex I dificiency.  I am looking forward to your reply, thanks!    Best regards,

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Answer

Thank you for contacting us. I am not sure which publication you are looking at however I have the following publication from Sashi Nadanaciva; http://www.mitosciences.com/PDF/MitoSciences_OXPHOS_Screen_Paper.pdf In this article MS101c is used as a Complex I capture antibody; which helps binding the Complex I to ELISA plate. Now check careful the second paragraph on page 3; "Complex I activity was measured by adding an assay solution containing 25mM KH2PO4 pH 7.2, 5mM MgCl2,1.2% (w/v) bovine serum albumin (BSA), 0.15mM Coenzyme Q1 and 0.26mM NADH. The oxidation of NADH was monitored by measuring its decrease in absorbance at 340 nm in a Spectra Max Plus384 plate reader (Molecular Devices) in kinetic mode at 30 °C for 90min. The rate was linear during this period" Now check the following Kit datasheet https://www.abcam.com/Complex-I-Enzyme-Activity-Microplate-Assay-Kit-ab109721-protocols.html ab109798 alone can’t be used to detect the enzyme activity; it will only bind the complex to the ELISA plate and then you will need further reagents which get reduced by complex 1 for activity measurement. I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

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Question

Hi, I have bought Complex I Immunocapture monoclonal antibody from your company for assay of complex I enzyme activity. I want to know whether or not complex I activity captured by this antibody is inhibited by rotenone? I am looking forward your reply.Thanks!  Sincerely yours,                                                            

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Answer

Thank you for contacting us. I suppose you have a question about ab109798; Anti-Complex I Immunocapture antibody. This antibody detects the presence of Complex I in cell sample; this unfortunately is not suitable for assaying the enzymatic activity. You need to select other products kits e.g. ab109721, ab109720 or other. Rotenone is an Electron Transport Chain inhibitor; it does not affect the biogenesis of mitochondrial complexes so this antibody will not be able to find difference between the Rotenone treated and Rotenone untreated cells.   For more information please click the following links http://www.mitosciences.com/mitobiogenesis-in-cell-elisa-kit-colorimetric.html http://www.ruf.rice.edu/~bioslabs/studies/mitochondria/mitopoisons.html I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

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Question

Dear technical team, Hope this e-mail finds you well! Kindly advise the suitable antibodies for below specificities as desired by one of our customer : Target :  Mitochondrial Complex I (encoded by mitochondrial DNA & not nuclear DNA)               Mitochondrial Complex III  (encoded by mitochondrial DNA & not nuclear DNA)Mitochondrial Complex IV  (encoded by mitochondrial DNA & not nuclear DNA) Mitochondrial Complex V   (encoded by mitochondrial DNA & not nuclear DNA)  Applications : IHC & WB Looking forward towards your valued response on the above. With Best Regards,  

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Answer

Thank you for contacting us. Please click these links to find the suitable antibodies. Type the MS code in Abcam search box to get the Abcam catalogue number. Complex I http://www.mitosciences.com/complex_i.html Complex IV http://www.mitosciences.com/complex_iv.html Complex III http://www.mitosciences.com/complex_iii.html Complex II http://www.mitosciences.com/complex_ii.html We also have Immuno capture kits available https://www.abcam.com/index.html?pageconfig=resource&rid=13917 I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

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