SLC25A35 Antibody

What is SLC25A35 and why is it significant for research?

SLC25A35 (Solute Carrier Family 25, Member 35) is a member of the mitochondrial carrier family that functions as a multi-pass membrane protein in the inner mitochondrial membrane . It contains three Solcar repeats, a characteristic feature of this protein family . Recent research has established SLC25A35 as a significant metabolic regulator that enhances fatty acid oxidation (FAO) and mitochondrial biogenesis, making it relevant for understanding mitochondrial metabolism in both normal and disease states . The protein has emerged as a potential oncogene in hepatocellular carcinoma (HCC), where it reprograms mitochondrial metabolism to support cancer cell proliferation and metastasis .

What types of SLC25A35 antibodies are available for research applications?

Several types of SLC25A35 antibodies are available for research, targeting different regions of the protein:

• Middle region-specific antibodies (AA 120-168)

• N-terminal region-specific antibodies

• Specific amino acid sequence targeting antibodies (AA 154-204)

These antibodies are predominantly rabbit polyclonal antibodies and are available in various conjugated and unconjugated forms, including:

What is the predicted reactivity spectrum of SLC25A35 antibodies?

Most commercially available SLC25A35 antibodies demonstrate cross-reactivity across multiple species, though with varying degrees of sequence homology. The predicted reactivity based on sequence conservation includes :

• Human: 100%

• Mouse: 100%

• Rat: 100%

• Dog: 100%

• Horse: 100%

• Pig: 100%

• Rabbit: 100%

• Cow: 86%

• Guinea Pig: 93%

• Zebrafish: 91%

This broad cross-reactivity makes these antibodies versatile tools for comparative studies across different model organisms .

What are the optimal protocols for Western blot analysis using SLC25A35 antibodies?

For Western blot analysis using SLC25A35 antibodies, researchers should consider the following methodological approach:

• Sample preparation: Isolate proteins from cells or tissues using RIPA buffer supplemented with protease inhibitors.

• Protein quantification: Use Bradford or BCA assay to normalize protein loading.

• Gel electrophoresis: Separate 20-50 μg of protein on 10-12% SDS-PAGE gels.

• Transfer: Transfer proteins to PVDF membranes (0.45 μm pore size preferred).

• Blocking: Block membranes in 5% non-fat milk or BSA in TBST for 1 hour at room temperature.

• Primary antibody incubation: Dilute SLC25A35 antibody at 1:500-1:2000 in blocking buffer and incubate overnight at 4°C .

• Secondary antibody: Use appropriate HRP-conjugated secondary antibody (typically anti-rabbit IgG) at 1:5000-1:10000 dilution.

• Detection: Develop using ECL substrate and visualize using a digital imaging system.

The middle region targeting antibody (ABIN5517415) has been validated specifically for Western blot applications, making it a reliable choice for protein expression analysis .

How should SLC25A35 antibodies be stored and handled to maintain optimal activity?

Proper storage and handling of SLC25A35 antibodies is critical for maintaining their activity and specificity:

• Storage temperature: Store at -20°C for up to 1 year from receipt date .

• Avoid freeze-thaw cycles: Aliquot antibodies upon first thaw to minimize repeated freeze-thaw cycles .

• Reconstitution: For lyophilized antibodies, reconstitute in distilled water to a final concentration of 1 mg/mL .

• Storage buffer: Some antibodies are provided in PBS containing 50% glycerol, 0.5% BSA, and 0.02% sodium azide to enhance stability .

• Working dilution preparation: Prepare working dilutions on the day of the experiment and discard after use.

• Transportation: Transport on ice when moving between storage and experimental areas.

Following these guidelines will help maintain antibody stability and experimental reproducibility .

How can specificity of SLC25A35 antibodies be validated in experimental settings?

Validating the specificity of SLC25A35 antibodies is crucial for reliable experimental outcomes. Researchers should consider implementing the following validation approaches:

• Positive controls: Use cell lines or tissues known to express SLC25A35, such as HCC cell lines (SNU-368, HLE, SNU739, SNU-354, HLF, HUH-7) .

• Negative controls:

  • Isotype controls to detect non-specific binding

  • SLC25A35 knockout or knockdown samples using siRNA

• Peptide competition assay: Pre-incubate antibody with the immunizing peptide before application to samples.

• Multiple antibody verification: Use different antibodies targeting distinct epitopes of SLC25A35 to confirm observations.

• Cross-reactivity testing: Evaluate potential cross-reactivity with other SLC25 family members through sequence alignment analysis.

• Immunoprecipitation followed by mass spectrometry: To confirm the identity of the protein being recognized.

The antibody targeting amino acids 154-204 has been confirmed to detect endogenous levels of SLC25A35 in human and mouse samples , providing a validated option for research applications.

How can SLC25A35 antibodies be utilized to investigate mitochondrial metabolism reprogramming in cancer?

Recent studies have revealed that SLC25A35 plays a crucial role in reprogramming mitochondrial metabolism in cancer cells . Researchers can utilize SLC25A35 antibodies to investigate this process through:

• Co-localization studies: Combine SLC25A35 antibodies with mitochondrial markers to confirm localization and distribution patterns in cancer versus normal cells.

• Metabolic pathway analysis: Use SLC25A35 antibodies in combination with antibodies against fatty acid oxidation (FAO) components to study pathway interactions.

• Bioenergetic profiling: Correlate SLC25A35 expression levels (determined by immunoblotting) with oxygen consumption rate (OCR) and ATP production measurements.

• Post-translational modification analysis: Investigate how acetylation affects SLC25A35 function using specific antibodies against acetylated proteins.

• Mitochondrial biogenesis assessment: Quantify relationship between SLC25A35 expression and mitochondrial mass/DNA content using immunofluorescence and qPCR.

Research has demonstrated that SLC25A35 enhances mitochondrial function characterized by increased oxygen consumption rate, ATP production, and decreased ROS levels through fatty acid oxidation enhancement , making these investigations particularly valuable.

What approaches can be used to study the relationship between SLC25A35 and PGC-1α in regulating mitochondrial function?

SLC25A35 has been shown to facilitate fatty acid oxidation and mitochondrial biogenesis through regulation of PGC-1α . To investigate this relationship, researchers can:

• Co-immunoprecipitation: Use SLC25A35 antibodies to pull down protein complexes and probe for PGC-1α interactions.

• Acetylation status analysis: Employ acetyl-CoA and acetylated-PGC-1α antibodies in conjunction with SLC25A35 antibodies to explore SLC25A35's role in PGC-1α acetylation.

• Chromatin immunoprecipitation (ChIP): Investigate how SLC25A35-mediated changes in PGC-1α affect binding to target gene promoters.

• Proximity ligation assay (PLA): Visualize and quantify interactions between SLC25A35 and PGC-1α in situ.

• Transcriptional reporter assays: Measure PGC-1α transcriptional activity in the presence of varying SLC25A35 levels.

This methodological approach can help elucidate the mechanistic pathway through which SLC25A35 upregulates PGC-1α via increased acetyl-CoA-mediated acetylation, ultimately enhancing mitochondrial function .

How can researchers resolve conflicting data when studying SLC25A35 across different cancer types?

When facing contradictory results regarding SLC25A35 function or expression across different cancer types, researchers should consider:

• Tissue-specific expression profiling: Use SLC25A35 antibodies for immunohistochemical analysis across diverse cancer tissue microarrays to establish tissue-specific expression patterns.

• Isoform-specific analysis: Employ antibodies targeting different epitopes to identify potential isoform-specific functions.

• Context-dependent signaling analysis: Investigate SLC25A35 in conjunction with tissue-specific transcription factors and signaling pathways.

• Multi-omics integration: Correlate protein expression data (obtained using antibodies) with transcriptomics and metabolomics data.

• Patient stratification: Analyze SLC25A35 expression in patient cohorts stratified by clinical parameters, genetic alterations, or metabolic profiles.

While SLC25A35 has been identified as an oncogene in hepatocellular carcinoma , research suggests that the broader SLC25 family may have varied prognostic value across different cancer types, including colon cancer , necessitating careful comparative studies.

What is the significance of SLC25A35 expression in hepatocellular carcinoma (HCC) and its potential as a therapeutic target?

SLC25A35 has emerged as a critical oncogene in hepatocellular carcinoma with significant therapeutic implications:

• Elevated expression: SLC25A35 is highly expressed in HCC tissues compared to adjacent non-tumor tissues, as demonstrated in a comprehensive study of 238 pairs of HCC and adjacent non-tumor tissues .

• Clinical correlation: High SLC25A35 expression correlates with adverse patient survival outcomes, suggesting its value as a prognostic marker .

• Functional significance: SLC25A35 promotes:

  • Proliferation of HCC cells in vitro and in vivo

  • Metastasis of HCC cells

  • Carcinogenesis in DEN-induced HCC mouse models

• Molecular mechanism: SLC25A35 reprograms mitochondrial metabolism by enhancing fatty acid oxidation and mitochondrial biogenesis through PGC-1α upregulation .

• Regulatory insights: SLC25A35 upregulation in HCC is partly caused by decreased miR-663a expression .

These findings position SLC25A35 as a promising therapeutic target for HCC treatment, where antibodies could serve as valuable tools for patient stratification and response monitoring .

How can SLC25A35 antibodies be applied in patient-derived xenograft (PDX) models for personalized cancer therapy research?

Researchers working with patient-derived xenograft models can utilize SLC25A35 antibodies to develop personalized approaches to cancer therapy:

• Expression profiling in PDX tissue: Use immunohistochemistry with SLC25A35 antibodies to characterize expression patterns in individual patient tumors grown in PDX models.

• Therapy response correlation:

  • Quantify SLC25A35 expression before and after treatment to identify expression changes

  • Correlate expression levels with response to metabolic-targeting therapies

• Heterogeneity mapping: Apply multicolor immunofluorescence with SLC25A35 and other markers to map intratumoral heterogeneity.

• Ex vivo drug sensitivity testing: Use SLC25A35 antibodies in combination with viability assays to correlate expression with drug sensitivity in PDX-derived organoids or slice cultures.

• Combinatorial therapy development: Identify potential synergistic targets that interact with SLC25A35-mediated metabolic pathways.

This approach could help identify patient subgroups likely to benefit from metabolism-targeting therapies, particularly those that interfere with fatty acid oxidation or mitochondrial biogenesis that are enhanced by SLC25A35 .

What is the metabolic impact of SLC25A35 inhibition or knockdown in cancer cells?

Understanding the metabolic consequences of SLC25A35 inhibition is essential for evaluating its potential as a therapeutic target:

These metabolic alterations following SLC25A35 inhibition provide a mechanistic basis for the observed reduction in cancer cell proliferation and metastasis, supporting the therapeutic potential of targeting this protein .

How can high-throughput screening approaches utilize SLC25A35 antibodies to identify novel inhibitors?

Researchers can implement SLC25A35 antibody-based high-throughput screening strategies to identify potential therapeutic compounds:

• Cell-based immunofluorescence assays: Develop automated imaging platforms using SLC25A35 antibodies to screen compound libraries for molecules that alter SLC25A35 expression, localization, or post-translational modifications.

• AlphaLISA/HTRF assays: Design homogeneous antibody-based assays to detect changes in SLC25A35 protein interactions or modifications following compound treatment.

• Reporter systems: Generate cells with fluorescent tags linked to SLC25A35 regulatory elements and validate with antibodies to screen for transcriptional modulators.

• Functional metabolic screening: Combine SLC25A35 expression analysis (using antibodies) with Seahorse-based metabolic profiling to identify compounds that specifically disrupt SLC25A35-mediated metabolic reprogramming.

• Phenotypic correlation: Correlate compound-induced changes in SLC25A35 expression/function (detected by antibodies) with cancer cell proliferation, migration, and invasion.

This approach could yield novel therapeutic candidates that specifically target the oncogenic functions of SLC25A35 in mitochondrial metabolism reprogramming .

What are the emerging techniques for studying SLC25A35 interaction with the mitochondrial proteome?

Advanced techniques for investigating SLC25A35's interactions within the mitochondrial environment include:

• Proximity-dependent biotin identification (BioID): Fuse BioID to SLC25A35 and use streptavidin pull-down followed by mass spectrometry to identify proximal proteins, validating key interactions with SLC25A35 antibodies.

• APEX2-based proximity labeling: Apply APEX2-tagged SLC25A35 for temporal mapping of protein neighborhoods within mitochondria.

• Cross-linking mass spectrometry (XL-MS): Use chemical crosslinkers to capture transient interactions involving SLC25A35, followed by mass spectrometry identification and antibody validation.

• Super-resolution microscopy: Combine SLC25A35 antibodies with super-resolution techniques like STORM or PALM to visualize nanoscale organization within mitochondria.

• Mitochondrial interactome mapping: Apply affinity purification with SLC25A35 antibodies coupled with quantitative proteomics to map comprehensive interaction networks.

These emerging techniques could reveal novel SLC25A35 interaction partners and mechanisms involved in mitochondrial metabolism regulation and cancer progression .

What are the key considerations for researchers new to working with SLC25A35 antibodies?

Researchers beginning work with SLC25A35 antibodies should consider these essential points:

• Antibody selection: Choose antibodies targeting specific epitopes based on your research question; middle region antibodies (AA 120-168) have been well-validated for Western blot applications .

• Validation strategy: Implement multiple validation approaches including positive and negative controls, particularly using SLC25A35 knockdown or knockout models .

• Application-specific optimization: Optimize conditions for each application (WB, IHC, IF) as dilution ranges vary from 1:500-1:2000 depending on the application and specific antibody .

• Species considerations: Verify cross-reactivity for your model organism, though most antibodies show good conservation across mammals .

• Storage and handling: Aliquot antibodies to avoid freeze-thaw cycles and store at -20°C for maximum stability .

Following these guidelines will help ensure reliable and reproducible results when studying SLC25A35 in various research contexts.

How might the field of SLC25A35 research evolve in the coming years?

The field of SLC25A35 research is poised for significant advances in several directions:

• Expanded cancer applications: Beyond HCC, investigation of SLC25A35's role in other cancer types will likely emerge, building on findings of SLC25 family members' prognostic value in colon cancer .

• Therapeutic development: Development of specific inhibitors targeting SLC25A35 or its downstream effectors may yield novel cancer therapeutics, particularly for metabolically active cancers .

• Biomarker potential: Validation of SLC25A35 as a prognostic or predictive biomarker for specific cancer subtypes could improve patient stratification and treatment selection.

• Systems biology integration: Integration of SLC25A35 into broader mitochondrial metabolic networks will enhance understanding of its regulatory role in health and disease.

• Structure-function studies: Determination of SLC25A35's 3D structure and transport mechanism will facilitate rational drug design approaches.

As research progresses, SLC25A35 antibodies will remain essential tools for characterizing expression patterns, protein interactions, and functional consequences of this emerging mitochondrial metabolism regulator .