Home Primary Antibodies Mox1 Antibody [M9G22]

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Mox1 Antibody [M9G22]

Catalog No.: F4321

    Application: Reactivity:
    • F4321-wb
      Lane 1: 293

    Usage Information

    Dilution
    1:100-1:1000
    1:50-1:500
    1:50-1:500
    Application
    WB, IP, IF, ELISA
    Reactivity
    Human, Mouse, Rat
    Source
    Mouse Monoclonal Antibody
    Storage Buffer
    PBS, pH 7.2+50% Glycerol+0.05% BSA+0.01% NaN3
    Storage (from the date of receipt)
    -20°C (avoid freeze-thaw cycles), 2 years
    Predicted MW
    65 kDa, 59 kDa, 22 kDa
    Positive Control 293; RT-4; JC; AT-3; WEHI-231
    Negative Control

    Exprimental Methods

    WB
    Experimental Protocol:
     
    Sample preparation
    1. Tissue: Lyse the tissue sample by adding an appropriate volume of ice-cold RIPA/NP-40 Lysis Buffer (containing Protease Inhibitor Cocktail)，and homogenize the tissue at a low temperature.
    2. Adherent cell: Aspirate the culture medium and wash the cells with ice-cold PBS twice. Lyse the cells by adding an appropriate volume of RIPA/NP-40 Lysis Buffer (containing Protease Inhibitor Cocktail) and put the sample on ice for 5 min.
    3. Suspension cell: Transfer the culture medium to a pre-cooled centrifuge tube. Centrifuge and aspirate the supernatant. Wash the cells with ice-cold PBS twice. Lyse the cells by adding an appropriate volume of RIPA/NP-40 Lysis Buffer (containing Protease Inhibitor Cocktail) and put the sample on ice for 5 min.
    4. Place the lysate into a pre-cooled microcentrifuge tube. Centrifuge at 4°C for 15 min. Collect the supernatant;
    5. Remove a small volume of lysate to determine the protein concentration;
    6. Combine the lysate with protein loading buffer. Boil 20 µL sample under 95-100°C for 5 min. Centrifuge for 5 min after cool down on ice.
     
    Electrophoretic separation
    1. According to the concentration of extracted protein, load appropriate amount of protein sample and marker onto SDS-PAGE gels for electrophoresis. Recommended separating gel (lower gel) concentration: 10%. Reference Table for Selecting SDS-PAGE Separation Gel Concentrations
    2. Power up 80V for 30 minutes. Then the power supply is adjusted (110 V~150 V), the Marker is observed, and the electrophoresis can be stopped when the indicator band of the predyed protein Marker where the protein is located is properly separated. (Note that the current should not be too large when electrophoresis, too large current (more than 150 mA) will cause the temperature to rise, affecting the result of running glue. If high currents cannot be avoided, an ice bath can be used to cool the bath.)
     
    Transfer membrane
    1. Take out the converter, soak the clip and consumables in the pre-cooled converter;
    2. Activate PVDF membrane with methanol for 1 min and rinse with transfer buffer;
    3. Install it in the order of "black edge of clip - sponge - filter paper - filter paper - glue -PVDF membrane - filter paper - filter paper - sponge - white edge of clip";
    4. The protein was electrotransferred to PVDF membrane. ( 0.45 µm PVDF membrane is recommended ) Reference Table for Selecting PVDF Membrane Pore Size Specifications
    Recommended conditions for wet transfer: 200 mA, 120 min.
    ( Note that the transfer conditions can be adjusted according to the protein size. For high-molecular-weight proteins, a higher current and longer transfer time are recommended. However, ensure that the transfer tank remains at a low temperature to prevent gel melting.)
     
    Block
    1. After electrotransfer, wash the film with TBST at room temperature for 5 minutes;
    2. Incubate the film in the blocking solution for 1 hour at room temperature;
    3. Wash the film with TBST for 3 times, 5 minutes each time.
     
    Antibody incubation
    1. Use 5% skim milk powder to prepare the primary antibody working liquid (recommended dilution ratio for primary antibody 1:100), gently shake and incubate with the film at 4°C overnight;
    2. Wash the film with TBST 3 times, 5 minutes each time;
    3. Add the secondary antibody to the blocking solution and incubate with the film gently at room temperature for 1 hour;
    4. After incubation, wash the film with TBST 3 times for 5 minutes each time.
     
    Antibody staining
    1. Add the prepared ECL luminescent substrate (or select other color developing substrate according to the second antibody) and mix evenly;
    2. Incubate with the film for 1 minute, remove excess substrate (keep the film moist), wrap with plastic film, and expose in the imaging system.
    IF
    Experimental Protocol：
     
    Specimen Preparation 
    1. Aspirate liquid, then cover cells to a depth of 2–3 mm with 4% Paraformaldehyde diluted in 1X PBS.
    NOTE: Paraformaldehyde is toxic, use only in a fume hood.
    2. Fix cells for 15 min at room temperature.
    3. Aspirate fixative, rinse three times in 1X PBS for 5 min each.
    4. Proceed with Immunostaining.
     
    Immunostaining
    1. Add theblocking buffer and incubate for 60 min at RT.
    2. Prepare primary antibody diluent in antibody dilution buffer as recommended .
    3. Aspirate blocking solution, apply diluted primary antibody.
    4. Incubate overnight at 4°C.
    5. Rinse three times in 1X PBS for 5 min each.
    6. Incubate specimens in fluorochrome-conjugated secondary antibody diluted in antibody dilution buffer for 1–2 hr at room temperature in the dark.
    7. Rinse three times in 1X PBS for 5 min each.
    8. Mount slides usingmounting medium with DAPI and cover with coverslips.
    9. For best results, allow mountant to cure overnight at room temperature. For long-term storage, store slides flat at 23°C protected from light.
     

    Datasheet & SDS

    Biological Description

    Specificity

    Mox1 Antibody [M9G22] detects endogenous levels of total Mox1 protein.
    Subcellular Location
    Cell junction, Cell membrane, Cell projection, Membrane
    Uniprot ID
    Q9Y5S8
    Clone
    M9G22
    Synonym(s)
    NADPH oxidase 1, NOX-1, Mitogenic oxidase 1 (MOX-1), NADH/NADPH mitogenic oxidase subunit P65-MOX, NOH-1, MOX1, NOH1.
    Background
    The Mox gene family, comprising Mox1 and Mox2, forms a distinct subgroup of non-clustered, divergent antennapedia-like homeobox genes expressed across various mesodermal tissues. Both Mox and Pax family genes are involved in overlapping developmental processes, exhibiting strong spatial and temporal co-expression during embryogenesis. Mox1 and Mox2 show high expression in developing somites, particularly within the sclerotome, where they are co-expressed with Pax1. Additionally, Mox2 and Pax3 are both present in migrating myoblasts. Functionally, Mox1 is essential for axial skeleton formation, while Mox2 plays a key role in limb muscle development. Beyond development, Mox1 is involved in the regulation of reactive oxygen species (ROS), particularly in non-phagocytic cells, and shares structural similarity with the glycoprotein gp91^phox, a core subunit of the NADPH oxidase complex. This complex generates superoxide, a reactive molecule central to numerous signaling pathways and immune responses. Mox1 expression is most abundant in the colon, with lower levels in the uterus and prostate, and is upregulated in vascular smooth muscle cells in response to platelet-derived growth factor stimulation. Mox1 contributes to the modulation of NADPH oxidase activity in tissues where ROS production is critical for signaling and adaptation to hypoxic conditions. The distinct tissue distribution of Mox1 compared to gp91phox further points to potential functional divergence, with Mox1 acting as a specialized regulator of oxidative stress responses.
    References
    • https://pubmed.ncbi.nlm.nih.gov/1363541/#&dopt=Abstract
    • https://pubmed.ncbi.nlm.nih.gov/11423130/

    Tech Support

    Answers to questions you may have can be found in the inhibitor handling instructions. Topics include how to prepare stock solutions, how to store inhibitors, and issues that need special attention for cell-based assays and animal experiments.

    Handling Instructions

    Tel: +1-832-582-8158 Ext:3
    If you have any other enquiries, please leave a message.

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