NIR Mitochondrial Membrane Potential Assay Kit (Microplate) (ab112150)

Reviews (1)Q&A (1)
Functional Studies - NIR Mitochondrial Membrane Potential Assay Kit (Microplate) (ab112150)

    Key features and details

    • Assay type: Direct
    • Detection method: Fluorescent
    • Platform: Microplate reader
    • Sample type: Adherent cells, Suspension cells

    Overview

    • Product name

      NIR Mitochondrial Membrane Potential Assay Kit (Microplate)
      See all Mitochondrial Membrane Potential kits

    • Detection method

      Fluorescent

    • Sample type

      Adherent cells, Suspension cells

    • Assay type

      Direct

    • Product overview

      ab112150 is designed to detect cell apoptosis by measuring the loss of the mitochondrial membrane potential. The collapse of mitochondrial membrane potential coincides with the opening of the mitochondrial permeability transition pores, leading to the release of cytochrome C into the cytosol, which in turn triggers other downstream events in the apoptotic cascade.


      ab112150 NIR Mitochondria Membrane Potential Assay Kit provides all the essential components with an optimized assay method. This fluorometric assay uses our proprietary cationic MitoNIR Dye for the detection of the mitochondrial membrane potential change in cells. In normal cells, the red fluorescence intensity is increased when NIR Dye is accumulated in the mitochondria. However, in apoptotic cells, NIR stain intensity is decreased following the collapse of MMP. Cells stained with NIR Dye can be monitored fluorimetrically at Em = 660-680 nm with excitation at Ex = 620-640 nm.


      ab112150 can be used for screening apoptosis activators and inhibitors. The assay can be performed in a convenient 96-well and 384-well fluorescence microtiter-plate format.

    • Notes

      Related assays

      Review the cell health assay guide to learn about kits to perform a cell viability assay, cytotoxicity assay and cell proliferation assay. 

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

    • Platform

      Microplate reader

    Properties

    Associated products

    Images

    • Functional Studies - NIR Mitochondrial Membrane Potential Assay Kit (Microplate) (ab112150)
      Functional Studies - NIR Mitochondrial Membrane Potential Assay Kit (Microplate) (ab112150)
      The decrease in NIR fluorescence with the addition of FCCP in HeLa cells.
      HeLa cells were dye loaded with NIR alone, or in the presence of 20 µM FCCP for 15 minutes. The fluorescence intensity of NIR was measured 30 minutes after adding Assay buffer B (Component C) with a microplate reader at Ex/Em = 640/680 nm (cut off 665 nm).

    References (0)

    Publishing research using ab112150? Please let us know so that we can cite the reference in this datasheet.

    ab112150 has not yet been referenced specifically in any publications.

    Customer reviews and Q&As

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    1-2 of 2 Abreviews or Q&A

    NIR Mitochondrial Membrane Potential Assay Kit (Microplate) (ab112150)

     Good Average 3/5 (Ease of Use)
    Abreviews
    This kit it is very simple to perform, just few steps and done. However I encounter so many troubles to get the right signal.
    Seeding density, I would suggest go in the high range suggested, either 96w or 384w plate format. I used FCCP and CCCP to test kit and both compounds work very nice, there are more efficacious if you do stimulation in PBS. On the reads I used Cytation 3 and I have to play so much to get right reads!...In my hands did not work auto gain, I have to play with it and set it at 150 gain.
    In my third run I could see the clear effect in a dose-response manner in two cell lines tested. Worth using it.
    However I tested JC-10 in paralel and in one run I got similar results with the not so easy NIR kit.

    Anabel de la Garza

    Verified customer

    Submitted Dec 24 2019

    Question

    What we are interested in is whether the assays are sensitive enough for our application.


    Will the abpromise mean that we are able to obtain a refund on the product if it is not sensitive enough?



    Many Thanks

    Read More
    Answer

    Thank you for your reply.

    The only data for sensitivity we have at the moment for ab112150 can be found on the datasheet as graph. I also include the actual numbers here:

    NIR fluorescence, four replicates.

    Control FCCP (20 uM)




    1777.557
    14157.28

    3074.621
    17427.79

    2893.949
    19235.95

    2681.359
    16799.75






    ab109905

    This assay was not designed to measure the activity of BHM, but rather the effect of a compound on a robust activity of BHM (which is provided with the kit). Ifyou want to usewhole mitochondria,the amount of mitochondria will have to be optimized, so that before you test compounds, you can measure a robust activity. The kit should work with any species, however the quality of the mitos will be very important.

    Are you asking forthe sensitivity of a particular drug on the activity of complex III (II+III)? This of course is compound dependant, but we can provide the IC50 for some examples we have run in house.





    Are you asking for the sensitivity of the assay at detecting BHM? The assay does not have an antibody. This is an in-solution assay which measures complex II+III in a 96 well plate. The assay was designed to use a robust signal of BHM so that compounds can be tested. Also, the assay does not have a “control protein complex II+III” to be able to produce the exact numbers I understand you wnat to see. We provide the BHM sample (which contains all the complexes of the electron transport chain). When customers run their assay, they should run the assay in the presence of compound and diluents. The diluent will typically be DMSO, which does not affect the assay at concentrations of 1 – 2%.





    Do youwant to know the raw signal in mOD/min in the presence of DMSO?. We can provide this as a reference guide, but not for ab109905, because the raw number is dependent on the BHM batch preparation and the substrates batch preparation. We can however giveout raw data from the QC of a particular batch, which is performed without DMSO.



    I hope this information is helpful and wish you good luck with your research.

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