MTCU1 Antibody

What is MTCO1 and why is it important in mitochondrial research?

MTCO1 (also known as MT-CO1, COX1, COXI, or COI) is cytochrome c oxidase subunit I, one of three mitochondrial DNA (mtDNA) encoded subunits of respiratory Complex IV. This protein spans 513 amino acid residues and belongs to the Heme-copper respiratory oxidase family . MTCO1 is critically important in mitochondrial research because:

• It forms a central component of the cytochrome c oxidase (Complex IV), the final enzyme in the electron transport chain

• It contains the active site (binuclear center) that catalyzes the reduction of oxygen to water

• It binds two heme groups (a and a3) and copper B (CuB), which are essential for electron transfer

• As an mtDNA-encoded protein, it serves as a marker for mitochondrial integrity and function

What applications are MTCO1 antibodies typically used for?

MTCO1 antibodies are utilized in multiple research applications including:

• Western blotting (WB): For protein detection and quantification

• Immunohistochemistry (IHC) and immunohistochemistry on formalin-fixed paraffin-embedded samples (IHC-P): For tissue localization studies

• Immunocytochemistry/Immunofluorescence (ICC/IF): For cellular localization

• Flow cytometry: For quantitative analysis of cell populations

• ELISA: For protein quantification in solution

The application-specific dilutions vary by antibody and manufacturer, but typical working dilutions range from 1:100 to 1:1000 for most applications .

How should I select the appropriate MTCO1 antibody for my specific experiment?

When selecting a MTCO1 antibody, consider these methodological factors:

• Species reactivity: Confirm the antibody reacts with your species of interest. Some antibodies show cross-reactivity with human, mouse, and rat samples, while others may be species-specific .

• Clonality selection:

  • Monoclonal antibodies (e.g., clone 1D6E1A8, 5D11-1C9) offer consistent batch-to-batch reproducibility and high specificity

  • Polyclonal antibodies may provide higher sensitivity by recognizing multiple epitopes

• Application compatibility: Verify the antibody has been validated for your specific application (WB, IHC, ICC, etc.) .

• Epitope location: Consider antibodies targeting different regions (N-terminal vs. C-terminal) as accessibility may vary depending on experimental conditions .

• Conjugation needs: Determine if you need an unconjugated antibody or one conjugated to fluorophores (like Alexa Fluor 488) or enzymes .

What controls should I include when using MTCO1 antibodies in mitochondrial research?

A methodologically sound experimental design should include:

• Positive control: Tissue or cell lysates known to express MTCO1 (e.g., HeLa cells, heart tissue) .

• Negative control:

  • Cells depleted of mitochondria or with mtDNA mutations affecting MTCO1

  • Isotype control antibody (e.g., mouse IgG2a for monoclonal antibodies) to assess non-specific binding

• Loading controls:

  • For Western blots: Nuclear-encoded mitochondrial proteins (e.g., HSP60) or total protein stains

  • For immunofluorescence: Co-staining with other mitochondrial markers (e.g., TOMM20)

• Validation approach: Comparing results using two different antibodies targeting different epitopes of MTCO1 .

How can MTCO1 antibodies be used to investigate mitochondrial disorders?

MTCO1 antibodies can be utilized in several advanced methodologies for investigating mitochondrial disorders:

• Quantitative analysis of mitochondrial content: Using flow cytometry with MTCO1-specific antibodies to assess mitochondrial mass in patient samples compared to controls .

• Detection of mtDNA mutations: Combined immunohistochemistry and genetic analysis to correlate MTCO1 expression levels with specific mtDNA mutations, especially in conditions like LHON (Leber Hereditary Optic Neuropathy) .

• Single-cell analysis protocol:

  • Fix cells with 100% methanol (5 min)

  • Permeabilize with 0.1% Triton X-100 (5 min)

  • Block with 1% BSA/10% normal serum/0.3M glycine in 0.1% PBS-Tween (1 hour)

  • Incubate with primary MTCO1 antibody overnight at 4°C (typically 1:1000 dilution)

  • Apply appropriate secondary antibody and counterstain

• Tissue-specific analysis: Compare MTCO1 expression across multiple tissues to identify tissue-specific mitochondrial defects, particularly in diseases with heterogeneous presentation .

What is the significance of MTCO1 expression patterns in different organs and age groups?

Research has revealed significant patterns in MTCO1 expression across tissues and age groups:

• Tissue-specific expression dynamics:

  • Non-immune organs (heart, lung, liver, kidneys, intestines) show age-dependent changes in MTCO1 expression

  • Immune organs (lymph nodes, spleen, thymus) display distinct patterns with lymph nodes showing increased expression in older subjects while spleen and thymus show decreased expression

• Metabolic implications: Changes in MTCO1 expression correlate with oxidative stress markers, such as malondialdehyde (MDA) levels, suggesting functional consequences of altered mitochondrial respiratory chain activity .

• Age-related differences: Younger subjects often show higher MTCO1 expression in non-immune organs compared to older subjects, pointing to potential age-related decline in mitochondrial function .

• Disease model variations: In models like the MRL/lpr lupus model, MTCO1 expression patterns differ significantly from controls, suggesting disease-specific mitochondrial adaptations .

What are the most common pitfalls when using MTCO1 antibodies and how can they be addressed?

How can I optimize protein extraction for MTCO1 detection?

For optimal MTCO1 detection, follow this methodological approach:

• Mitochondrial enrichment:

  • Homogenize tissues in isotonic buffer (250mM sucrose, 10mM Tris-HCl, 1mM EDTA, pH 7.4)

  • Perform differential centrifugation (600g to remove nuclei, 7,000g to pellet mitochondria)

  • For increased purity, use density gradient centrifugation

• Membrane protein solubilization:

  • Use RIPA buffer supplemented with 1-2% mild detergent (digitonin or n-dodecyl β-D-maltoside)

  • Include protease inhibitors and perform extraction at 4°C

  • Avoid excessive sonication which may damage mitochondrial complexes

• Sample handling:

  • Process samples quickly to minimize degradation

  • If using frozen samples, avoid repeated freeze-thaw cycles

  • For Western blotting, do not heat samples above 70°C to prevent aggregation of membrane proteins

How do I interpret changes in MTCO1 expression in relation to mitochondrial function?

When analyzing MTCO1 expression data:

• Expression level contextualization:

  • Increased MTCO1 often indicates mitochondrial biogenesis or compensatory upregulation

  • Decreased MTCO1 may suggest impaired mitochondrial content, mtDNA depletion, or specific Complex IV defects

  • Compare results to other mitochondrial markers (nuclear-encoded subunits, mitochondrial mass indicators)

• Functional correlation analysis:

  • Correlate MTCO1 levels with markers of oxidative stress (e.g., MDA levels)

  • Assess relationship between MTCO1 expression and respiratory chain enzyme activities

  • Consider the ratio of mtDNA-encoded to nuclear-encoded subunits as an indicator of mitochondrial genetics integrity

• Tissue-specific interpretation:

  • Brain tissue typically shows strong correlation between low MTCO1 mRNA expression and high oxidative stress

  • Immune tissues demonstrate unique patterns that may reflect tissue-specific mitochondrial adaptations

What are the latest advanced techniques incorporating MTCO1 antibodies in mitochondrial research?

Recent methodological advances include:

• Multiplexed imaging:

  • Combining MTCO1 antibodies with other mitochondrial markers using spectral imaging

  • Super-resolution microscopy applications for sub-mitochondrial localization

  • Multi-epitope ligand cartography for in situ protein interaction studies

• Single-cell proteomics:

  • Mass cytometry (CyTOF) integrating MTCO1 antibodies for high-dimensional analysis

  • Correlation of mitochondrial parameters with cellular phenotypes at single-cell resolution

• Proximity labeling techniques:

  • BioID or APEX2 fusions to identify proteins interacting with MTCO1 in living cells

  • Spatial mapping of the MTCO1 interactome during mitochondrial complex assembly

• Live-cell applications:

  • Development of cell-permeable antibody fragments for tracking MTCO1 in living systems

  • Antibody-guided targeting of therapeutic compounds to mitochondria