UQCRC1 Antibody

What is UQCRC1 and what is its physiological function?

UQCRC1 is a core protein component of ubiquinol-cytochrome c reductase complex (complex III or cytochrome b-c1 complex) within the mitochondrial respiratory chain. The protein consists of 480 amino acids with a predicted molecular weight of approximately 52.6-53 kDa. Functionally, UQCRC1 contributes to electron transport and ATP generation within the mitochondrial inner membrane. More specifically, it may mediate the formation of the complex between cytochromes c and c1, playing a critical role in oxidative phosphorylation. The bc1 complex it belongs to contains 11 subunits, with UQCRC1 being one of the two core proteins essential for proper functioning of the respiratory chain .

What types of UQCRC1 antibodies are available for research?

UQCRC1 antibodies are available in several formats to accommodate various research applications. These include:

• Host Species: Mouse and rabbit-derived antibodies are most common

• Clonality: Both monoclonal (e.g., clone O91G2, OTI1A3, 1G2) and polyclonal antibodies

• Target Regions: Antibodies targeting different regions of the protein, including:

  • N-terminal region (AA 1-63, AA 141-170)

  • Middle section (AA 46-305)

  • C-terminal regions

  • Full-length protein (AA 1-480)

• Applications: Antibodies optimized for Western blotting, immunocytochemistry, immunohistochemistry, ELISA, and other applications

• Reactivity: Primarily human-reactive, with some cross-reactivity to mouse and rat

How do I select the appropriate UQCRC1 antibody for my specific research question?

Selection should be guided by your experimental goals and techniques. For localization studies in human samples, consider antibodies validated for immunohistochemistry with human reactivity. For protein expression quantification, Western blot-validated antibodies are essential. When studying protein interactions, consider antibodies recognizing specific domains not involved in those interactions. Monoclonal antibodies offer consistency between lots but may be sensitive to epitope masking, while polyclonal antibodies provide robust detection but with potential batch variation. Review the immunogen information to ensure the antibody recognizes relevant protein regions of interest. For example, if studying N-terminal processing, select antibodies targeting this region such as those recognizing AA 1-63 or AA 141-170. Cross-reactivity information is crucial for comparative studies across species .

What are the optimal conditions for using UQCRC1 antibodies in Western blotting applications?

For Western blotting applications with UQCRC1 antibodies, optimal conditions include using 0.1-1.0 μg/ml antibody concentration as a starting point, then titrating as needed. When preparing samples, ensure complete mitochondrial protein extraction using specialized buffers containing mild detergents like CHAPS or digitonin rather than harsh detergents that may disrupt protein complexes. For gel separation, use 10-12% SDS-PAGE gels to properly resolve the ~53 kDa UQCRC1 protein. Transfer conditions should be optimized for mitochondrial proteins, typically using PVDF membranes with 100-120V transfer for 60-90 minutes in cold conditions. Blocking should employ 5% non-fat milk or BSA in TBST for 1 hour at room temperature. For detection, HRP-conjugated secondary antibodies followed by ECL detection provide good sensitivity, with exposure times of 30 seconds to 5 minutes typically sufficient for visualization .

How should UQCRC1 antibodies be used for immunocytochemistry and immunohistochemistry applications?

For immunocytochemistry (ICC) and immunohistochemistry (IHC) applications with UQCRC1 antibodies, begin with a concentration range of 0.5-5.0 μg/ml as recommended by manufacturers. Fixation protocols should preserve mitochondrial structure—4% paraformaldehyde for 15-20 minutes is generally effective. For tissue sections, antigen retrieval is crucial; citrate buffer (pH 6.0) heat-induced epitope retrieval works well for most UQCRC1 epitopes. When performing permeabilization (essential for accessing mitochondrial proteins), use 0.1-0.2% Triton X-100 for 10 minutes, being careful not to over-permeabilize. Blocking should employ 5-10% normal serum from the secondary antibody host species with 1% BSA for 1 hour. For visualization, fluorescent secondary antibodies work well for co-localization studies with other mitochondrial markers. Counterstaining with DAPI helps visualize nuclei, while MitoTracker can confirm mitochondrial localization. Validation controls should include UQCRC1 knockdown or knockout samples when possible .

What protocols are recommended for measuring UQCRC1 expression changes during disease progression?

To measure UQCRC1 expression changes during disease progression, a multi-technique approach is recommended. For quantitative protein analysis, Western blotting with carefully selected loading controls (preferably other mitochondrial proteins of different complexes like VDAC or ATP5A) should be performed. For tissue studies, immunohistochemistry with standardized scoring systems (H-score or percent positive cells) enables comparison across disease stages. For example, studies in pancreatic cancer demonstrated a gradual increase in UQCRC1 expression during progression from pancreatic intraepithelial neoplasias (PanIN) to PDAC, with elevated expression observed in 72.3% of PDAC cases that correlated with poor prognosis. RT-qPCR can complement protein studies by measuring mRNA expression changes, while digital spatial profiling allows region-specific quantification within heterogeneous tissues. Longitudinal sampling when possible (e.g., in animal models or sequential biopsies) provides the most accurate assessment of temporal changes during disease progression .

How does UQCRC1 expression correlate with cancer progression and what methodologies are used to study this relationship?

UQCRC1 expression shows significant correlation with cancer progression in multiple malignancies. In pancreatic ductal adenocarcinoma (PDAC), researchers have documented a gradual increase in UQCRC1 expression from early pancreatic intraepithelial neoplasias (PanIN) stages to advanced PDAC in KPC mouse models. Methodologically, this correlation is established through a combination of techniques. Immunohistochemistry staining of tissue microarrays with anti-UQCRC1 antibodies allows quantitative scoring across tumor stages, revealing elevated expression in 72.3% of PDAC cases with strong correlation to poor prognosis. Real-time cell analysis (RTCA) and colony formation assays with UQCRC1-overexpressing cells demonstrate increased growth rates. For mechanistic understanding, extracellular flux analysis using Seahorse technology reveals UQCRC1's role in enhancing mitochondrial oxidative phosphorylation and ATP production. RNA-Seq analysis of UQCRC1-modulated cells provides insights into downstream pathway activation. Validating these findings through in vivo subcutaneous and orthotopic mouse models with UQCRC1 knockdown/overexpression confirms the causal relationship between UQCRC1 expression and cancer progression .

What functional roles does UQCRC1 play in cancer cell metabolism and how can this be experimentally assessed?

UQCRC1 serves as a critical regulator of cancer cell metabolism through several mechanisms that can be experimentally assessed. Its primary role involves enhancing mitochondrial oxidative phosphorylation (OXPHOS) and ATP production, which can be quantified using Seahorse XF analyzers measuring oxygen consumption rate (OCR) in cells with UQCRC1 overexpression or knockdown. The resulting increased ATP production is measurable using luminescence-based ATP assays. Importantly, research shows this ATP is released into the extracellular space via pannexin 1 channels—a process assessable through extracellular ATP measurements with luciferin-luciferase assays. The released ATP functions as an autocrine/paracrine signaling molecule activating the ATP/P2Y2-RTK/AKT axis, detectable through phosphorylation status of these pathway components using phospho-specific antibodies in Western blots. This signaling cascade ultimately promotes cancer cell proliferation, measurable through proliferation assays including RTCA and colony formation. The functional significance can be validated by blocking experiments targeting either UQCRC1 expression (RNA interference) or ATP release pathways, which effectively inhibit cancer growth both in vitro and in vivo, as demonstrated in PDAC models .

Beyond cancer, what other pathological conditions involve UQCRC1 dysregulation?

Beyond its well-documented role in cancer, UQCRC1 dysregulation appears in several neurological, metabolic, and developmental disorders. In schizophrenia, altered UQCRC1 expression correlates with mitochondrial dysfunction in brain tissue, potentially contributing to the energy metabolism abnormalities observed in this condition. Methods to assess this include post-mortem brain tissue analysis using UQCRC1 antibodies for Western blotting and immunohistochemistry. In Rett syndrome, a severe neurodevelopmental disorder, UQCRC1 abnormalities may contribute to the mitochondrial dysfunction characteristic of the disease. Similarly, in inherited insulin resistance, UQCRC1 dysregulation has been linked to metabolic abnormalities. Research approaches to study these connections include patient-derived fibroblast or induced pluripotent stem cell (iPSC) models, where UQCRC1 expression and function can be assessed through a combination of protein quantification, mitochondrial function assays, and metabolic profiling. Animal models with UQCRC1 manipulation provide additional insights into the causal relationships between its dysregulation and pathological phenotypes in these diverse conditions .

How can UQCRC1 antibodies be utilized in the study of mitochondrial complex assembly and function?

UQCRC1 antibodies serve as valuable tools for studying mitochondrial complex III assembly and function through multiple advanced techniques. For complex assembly analysis, blue native PAGE (BN-PAGE) combined with UQCRC1 antibodies in subsequent Western blots allows visualization of intact complex III (~450 kDa) versus free UQCRC1 subunits (~53 kDa). Co-immunoprecipitation experiments using UQCRC1 antibodies can identify interaction partners and assembly intermediates—the bc1 complex contains 11 subunits including 3 respiratory subunits, 2 core proteins (including UQCRC1), and 6 low-molecular weight proteins. For functional studies, immuno-capture of complex III with UQCRC1 antibodies coupled to activity assays measuring electron transfer from ubiquinol to cytochrome c provides insights into functional integrity. Proximity ligation assays (PLA) using UQCRC1 antibodies paired with antibodies against other complex III components or interacting complexes enable in situ visualization of protein proximity. For dynamic assembly studies, pulse-chase experiments with metabolic labeling followed by UQCRC1 immunoprecipitation track the incorporation rate of newly synthesized protein into mature complexes. Super-resolution microscopy with fluorophore-conjugated UQCRC1 antibodies provides nanoscale visualization of complex distribution within mitochondria .

What are common technical challenges when working with UQCRC1 antibodies and how can they be overcome?

Researchers working with UQCRC1 antibodies frequently encounter several technical challenges that can be systematically addressed. Mitochondrial membrane protein extraction inefficiency can be overcome by using specialized mitochondrial isolation buffers containing digitonin or mild detergents like CHAPS, followed by sonication in the presence of protease inhibitors. Epitope masking—particularly problematic for antibodies targeting regions involved in complex formation—can be addressed by testing multiple antibodies recognizing different epitopes or employing denaturing conditions for Western blotting while using milder conditions for native complex studies. Cross-reactivity with other mitochondrial proteins may occur; this requires thorough validation using UQCRC1 knockout/knockdown controls and pre-absorption with recombinant proteins when necessary. Inconsistent immunohistochemistry staining can result from variable fixation affecting mitochondrial structure integrity; standardized fixation protocols (4% paraformaldehyde, 10-15 minutes) and careful optimization of antigen retrieval methods (citrate buffer pH 6.0 or Tris-EDTA pH 9.0) significantly improve consistency. The high abundance of mitochondria in certain tissues may create background issues; this can be addressed by careful titration of primary antibody concentrations (starting at 0.1-1.0 μg/ml) and using monoclonal antibodies when specific detection is critical .

How can UQCRC1 antibodies be integrated into multi-parameter analyses of mitochondrial function?

Integration of UQCRC1 antibodies into multi-parameter mitochondrial function analyses provides comprehensive insights into respiratory chain dynamics. For multiplexed immunofluorescence, combine fluorophore-conjugated UQCRC1 antibodies with markers for other respiratory complexes, mitochondrial membrane potential indicators like TMRM, and superoxide sensors like MitoSOX, ensuring compatible fluorophore selection to avoid spectral overlap. Flow cytometry applications require careful permeabilization protocols (0.1% saponin works well) to access mitochondrial epitopes while preserving functional parameters; this approach enables correlation of UQCRC1 levels with mitochondrial mass, membrane potential, and ROS production at the single-cell level. Mass cytometry (CyTOF) with metal-conjugated UQCRC1 antibodies allows simultaneous assessment of dozens of parameters without fluorescence limitations. For functional correlation, combine immunocytochemistry with live-cell microscopy by first measuring functional parameters (OCR, ATP production) then fixing and staining the same cells. Spatial proteomics approaches using multiplexed ion beam imaging (MIBI) or CO-Detection by indEXing (CODEX) with UQCRC1 antibodies enable visualization of its distribution relative to other mitochondrial and cellular components in intact tissue architecture .

What methodological approaches enable the study of UQCRC1 post-translational modifications?

Studying UQCRC1 post-translational modifications requires specialized methodological approaches. For comprehensive modification profiling, immunoprecipitate UQCRC1 using validated antibodies followed by mass spectrometry analysis—this approach has identified phosphorylation, acetylation, and ubiquitination sites on UQCRC1. To study specific modifications, use modification-specific antibodies (e.g., anti-phospho-UQCRC1) in Western blotting or develop them if not commercially available. Alternatively, employ antibodies against common modifications (phospho-Ser/Thr/Tyr, acetyl-Lys) after UQCRC1 immunoprecipitation. For modification site identification, combine site-directed mutagenesis of predicted modification sites with functional assays to determine their impact on protein function or complex assembly. To assess modification dynamics during cellular processes, use pulse-chase labeling with modification-specific tracers followed by UQCRC1 immunoprecipitation. For spatial distribution of modified UQCRC1, perform proximity ligation assays using pairs of antibodies against UQCRC1 and specific modifications. Modification-specific functional impacts can be assessed by comparing activity of immunoprecipitated native complexes before and after treatment with modification-removing enzymes (phosphatases, deacetylases, etc.) .

How can UQCRC1 be therapeutically targeted in cancer and what methods are used to evaluate efficacy?

Therapeutic targeting of UQCRC1 in cancer can be approached through several strategies with specific methodologies for efficacy evaluation. RNA interference represents a direct approach, where siRNA or shRNA against UQCRC1 delivered via lipid nanoparticles or viral vectors has shown efficacy in inhibiting PDAC growth in both in vitro and in vivo models. Efficacy is evaluated through tumor growth measurements, immunohistochemical analysis of proliferation markers, and survival assessments in animal models. Downstream pathway inhibition offers an alternative approach by targeting the ATP/P2Y2-RTK/AKT signaling axis activated by UQCRC1-mediated ATP production. Small molecule inhibitors of P2Y2 receptors or AKT can be evaluated using phospho-protein analysis of pathway components, proliferation assays, and xenograft models. ATP release blockade represents a particularly promising strategy, as research has shown that blocking the pannexin 1 channel (through which ATP is released) effectively inhibits PDAC growth. This approach can be assessed through extracellular ATP measurements, cell proliferation assays, and in vivo tumor models. Combination strategies targeting UQCRC1 alongside standard chemotherapeutics require careful evaluation through combination index calculations and in vivo efficacy studies with clinically relevant endpoints .

What are emerging techniques for studying UQCRC1 interactions with the mitochondrial respiratory chain?

Emerging techniques for studying UQCRC1 interactions with the mitochondrial respiratory chain provide unprecedented insights into its functional relationships. Cryo-electron microscopy now achieves near-atomic resolution of respiratory complexes, revealing detailed interaction interfaces between UQCRC1 and other components. This approach requires purified complexes and specialized sample preparation but yields structural information impossible to obtain through other methods. Cross-linking mass spectrometry (XL-MS) employs chemical cross-linkers that covalently link interacting proteins, followed by mass spectrometry to identify interaction sites; when applied to isolated mitochondria, this technique maps the UQCRC1 interaction network within its native environment. Proximity-dependent labeling approaches like BioID or APEX2 involve fusing these enzymes to UQCRC1, allowing biotinylation of proximal proteins that can be identified by mass spectrometry, providing a dynamic view of the UQCRC1 interactome. Single-molecule techniques including FRET (Förster Resonance Energy Transfer) with fluorescently labeled components enable real-time visualization of interactions and conformational changes during electron transport. Integrative structural biology approaches combining multiple data sources (cryo-EM, XL-MS, computational modeling) generate comprehensive models of UQCRC1's role within the respiratory chain architecture .

How can UQCRC1 antibodies contribute to biomarker development and precision medicine approaches?

UQCRC1 antibodies offer significant potential for biomarker development and precision medicine applications, particularly in cancer. For diagnostic applications, immunohistochemistry using UQCRC1 antibodies on tissue microarrays enables stratification of cancer patients—this approach has already demonstrated clinical relevance with 72.3% of PDAC cases showing elevated UQCRC1 expression correlating with poor prognosis. Multiplex immunofluorescence panels incorporating UQCRC1 alongside other markers provide more comprehensive tumor characterization. For liquid biopsy development, UQCRC1 antibodies can detect the protein in circulating tumor cells or extracellular vesicles isolated from patient blood samples, potentially enabling non-invasive monitoring. In predictive biomarker applications, correlation of UQCRC1 expression levels determined by immunohistochemistry with treatment responses helps identify patient subgroups likely to benefit from specific therapies. Companion diagnostic development using standardized UQCRC1 immunoassays could eventually guide patient selection for therapies targeting mitochondrial metabolism. For monitoring treatment response, sequential tissue or liquid biopsies analyzed for UQCRC1 expression provide insights into therapeutic efficacy, particularly for treatments targeting mitochondrial function or downstream pathways activated by UQCRC1-mediated ATP production .