elovl6 Antibody

What is ELOVL6 and why is it important in molecular research?

ELOVL6 (Elongation of very long chain fatty acids protein 6) functions as a microsomal enzyme that catalyzes the first and rate-limiting reaction in the long-chain fatty acid elongation cycle. It primarily elongates saturated and monounsaturated fatty acids with 12, 14, and 16 carbons, showing higher activity toward C16:0 acyl-CoAs .

ELOVL6 is crucial in lipid metabolism research because:

• It serves as a critical metabolic checkpoint in insulin sensitivity and lipotoxicity

• It plays a significant role in obesity-related metabolic disorders

• Its activity affects cellular fatty acid composition, which influences membrane properties and signaling pathways

• Altered ELOVL6 expression is implicated in various pathological conditions including type 2 diabetes, cancer, and multiple sclerosis

What are the standard applications for ELOVL6 antibodies in research?

ELOVL6 antibodies are versatile research tools with several validated applications:

Positive IHC detection has been reported in human hepatocirrhosis tissue and human brain tissue . When selecting an ELOVL6 antibody, researchers should consider the reactive species needed as antibodies are available with reactivity to human, mouse, and rat ELOVL6 .

How should ELOVL6 knockdown experiments be designed and validated?

When designing ELOVL6 knockdown experiments, researchers should consider multiple approaches for robust validation:

RNA Interference Approach:

• siRNA transfection has been successfully used with sequence: sense strand GCUCUUCGAACUGGUGCUUTT and antisense strand AAGCACCAGUUCGAAGAGCTT

• Allow cells to reach 100% confluence before differentiation following transfection

• Include appropriate non-targeting siRNA controls (e.g., Silencer™ negative control)

shRNA Approach:

• Adenoviral vectors carrying ELOVL6 shRNA have been used at 10 multiplicities of infection (MOI) for effective knockdown

• Knockdown should be verified using RT-PCR analysis

CRISPR/Cas9 Approach:

• Complete ELOVL6 knockout has been achieved with appropriate guide RNA design

• Validation of knockout should include functional assays, as ELOVL6 inhibitors show no additive effects in ELOVL6 KO cells

Validation Metrics:

• Measure mRNA expression of ELOVL6 by RT-PCR

• Confirm protein reduction by Western blot (expected 29-32 kDa band)

• Assess functional consequences such as altered cell proliferation, changes in fatty acid profiles, or cell morphology

What are the optimal conditions for using ELOVL6 antibodies in immunohistochemistry?

For optimal IHC results with ELOVL6 antibodies, researchers should follow these protocol recommendations:

Tissue Preparation:

• Formalin-fixed, paraffin-embedded (FFPE) sections are commonly used

• Positive control tissues include human hepatocirrhosis tissue and human brain tissue

Antigen Retrieval:

• Primary recommendation: TE buffer pH 9.0

• Alternative method: citrate buffer pH 6.0

Antibody Dilution:

• Typical range: 1:50-1:500 for IHC applications

• Titration is recommended for each specific testing system to obtain optimal results

Detection Systems:

• Both chromogenic and fluorescent detection systems have been validated

• For semi-quantitative analysis, the H-score system has been used with cutoffs at H-scores between 0-139 (low expression) and ≥140 (high expression)

Important Considerations:

• ELOVL6 expression can vary significantly between tumor and adjacent normal tissue

• In hepatocellular carcinoma studies, 50% of tumors showed increased ELOVL6 expression compared to adjacent normal tissue

• Expression patterns may differ based on pathological conditions and tissue types

How does ELOVL6 expression correlate with disease outcomes and what methods should be used for clinical correlation studies?

ELOVL6 expression has significant correlations with disease outcomes, requiring specific methodological approaches for accurate clinical correlation studies:

In Cancer Research:

Quantification Methods:

• IHC with H-score system (0-300 scale) provides semi-quantitative assessment

• RNA sequencing or qRT-PCR for mRNA expression levels

• Protein levels by Western blot with densitometry analysis

Clinical Correlation Framework:

• Stratify patients based on ELOVL6 expression levels (high vs. low)

• Analyze clinical parameters: disease stage, differentiation, patient demographics

• Perform univariate and multivariate analyses for survival outcomes

• Consider additional metabolic parameters (BMI, diabetes status)

What mechanisms underlie the relationship between ELOVL6 and drug resistance in cancer, and how can this be studied?

Research has revealed important connections between ELOVL6 expression and drug resistance mechanisms, particularly in multiple myeloma (MM) and pancreatic cancer:

Key Mechanisms:

• Altered lipid metabolism: Changes in ELOVL6-dependent lipidome affect cellular response to therapeutics

• Endoplasmic reticulum (ER) stress modulation: ELOVL6 depletion suppresses bortezomib-induced ER stress and cytotoxicity

• Membrane rigidity alteration: ELOVL6 interference results in reduced membrane rigidity, which may affect drug uptake or efflux

Research Methodologies:

• Cell line models: Establish drug-resistant cell lines through incremental drug exposure and monitor ELOVL6 expression changes

• Gene expression analysis: RNA sequencing to identify ELOVL6 and related pathway alterations

• Functional validation: Restore ELOVL6 expression in resistant cells to assess re-sensitization to therapy

• In vivo validation: Xenograft models to confirm findings from in vitro studies

Experimental Design Framework:

• Compare ELOVL6 expression between drug-responsive and non-responsive patient cells

• Analyze global lipidome changes in resistant versus sensitive cells

• Perform ELOVL6 knockdown/overexpression studies to directly test its role in drug sensitivity

• Combine ELOVL6 modulation with drug treatment to assess therapeutic potential

In multiple myeloma, lower ELOVL6 levels have been associated with resistance to bortezomib (BTZ) in both patient-derived and cultured MM cells. Depletion of ELOVL6 in parental MM cells suppressed BTZ-induced endoplasmic reticulum stress and cytotoxicity, while restoration of ELOVL6 levels in BTZ-resistant MM cells sensitized them to BTZ both in vitro and in vivo .

How can researchers resolve discrepancies in observed molecular weights of ELOVL6 in Western blot experiments?

Discrepancies in observed molecular weights of ELOVL6 in Western blot experiments are common and can be resolved through systematic analysis:

Expected vs. Observed Molecular Weights:

• Calculated molecular weight: 31 kDa (265 amino acids)

• Commonly observed ranges:

  • 29-32 kDa

  • 39 kDa

  • 68 kDa

Potential Causes of Discrepancies:

Validation Approach:

• Include appropriate positive controls (e.g., tissue with known ELOVL6 expression)

• Run ELOVL6 knockdown/knockout samples as negative controls

• Test multiple antibodies targeting different epitopes

• If available, use blocking peptide competition to confirm specificity

• Compare results with published literature, noting the antibody used

If discrepancies persist, researchers should contact antibody manufacturers for additional technical support and guidance specific to their antibody clone.

What factors contribute to contradictory findings in ELOVL6 expression studies across different disease models?

Contradictory findings in ELOVL6 expression studies across disease models can be attributed to several factors:

Methodological Variations:

• Different antibody clones with varying specificities

• Diverse quantification methods (IHC, WB, qPCR)

• Variations in tissue processing and fixation protocols

• Heterogeneous scoring systems for expression levels

Biological Factors:

Recommendations for Resolving Contradictions:

• Clearly define the specific disease context and patient population

• Use multiple detection methods in parallel (protein and mRNA)

• Include appropriate tissue and cellular controls

• Consider the influence of metabolic state and dietary factors

• Examine ELOVL6 in conjunction with its regulatory pathways

• Report detailed methodological parameters to enable proper comparison across studies

For example, in hepatocellular carcinoma research, contradictory findings in ELOVL6 expression levels between different studies highlight the importance of examining ELOVL6 in HCC cases with different etiologies to define its contribution in specific contexts .

How can ELOVL6 antibodies be used to investigate the role of lipid metabolism in neurodegenerative diseases?

Recent research has revealed promising applications of ELOVL6 antibodies in studying neurodegenerative conditions, particularly multiple sclerosis (MS):

Key Research Applications:

• Tracking ELOVL6 expression in phagocytes within demyelinated lesions

• Investigating the relationship between ELOVL6-mediated fatty acid elongation and remyelination

• Examining how ELOVL6 modulates phagocyte phenotypes and repair mechanisms

Methodological Approach:

• Use immunohistochemistry with ELOVL6 antibodies to quantify expression in CNS tissue sections

• Combine with markers for phagocytes, myelin, and repair processes

• Compare expression patterns between normal tissue, acute demyelinating lesions, and chronic lesions

• Correlate ELOVL6 expression with repair-associated markers

Research Findings:
ELOVL6 is significantly upregulated in phagocytes in demyelinated lesions, and ELOVL6 deficiency induces a reparative phagocyte phenotype that promotes remyelination. ELOVL6-deficient foamy macrophages show enhanced ABCA1-mediated lipid efflux, increased production of neurotrophic factors (such as IGF1, TGFβ1, and CNTF), and reduced expression of inflammatory mediators .

Potential Therapeutic Implications:
Targeting ELOVL6 may provide an avenue for developing reparative therapies for MS and other neurodegenerative diseases by promoting remyelination and tissue repair .

What are the emerging techniques for studying the impact of ELOVL6 on membrane properties and cellular biomechanics?

Innovative techniques are now being employed to understand how ELOVL6-mediated changes in fatty acid composition affect cellular biomechanics:

Advanced Methodologies:

• Membrane Tension Analysis:

  • Using normal mode analysis to extract effective values for membrane tension

  • Analyzing membrane flexibility through cell shape variability metrics

• Indentation Techniques:

  • Measuring membrane rigidness against normal stress

  • Assessing permeability under induced cortical deformation

  • Quantifying significant reduction in membrane rigidity upon ELOVL6 interference

• Lipidomic Approaches:

  • Mass spectrometry-based lipidomics to profile fatty acid composition changes

  • Correlating specific lipid species alterations with membrane physical properties

  • Identifying functional connections between lipid composition and cellular behavior

Research Findings:
Studies using these advanced techniques have demonstrated that ELOVL6 interference (through either shRNA downregulation or chemical inhibition) results in:

• Significant reduction in membrane rigidity

• More flexible behavior and higher variability in cell shape

• Altered membrane biomechanical properties that influence cellular functions and drug responses

Applications in Cancer Research:
These emerging techniques are particularly valuable in cancer research, where ELOVL6-mediated changes in membrane properties may influence:

• Drug uptake and efficacy

• Cell proliferation and migration capabilities

• Cellular response to therapeutic agents

By incorporating these advanced biophysical methods with traditional molecular approaches, researchers can gain deeper insights into how ELOVL6-mediated lipid metabolism impacts cellular function at the mechanical and structural levels.

What are the optimal storage and handling conditions for ELOVL6 antibodies to maintain long-term activity?

Proper storage and handling of ELOVL6 antibodies is essential for maintaining their specificity and sensitivity over time:

Storage Conditions:

• Store at -20°C for long-term storage (stable for one year after shipment)

• For frequent use, short-term storage at 4°C is acceptable (up to three months)

• Avoid repeated freeze-thaw cycles that can degrade antibody quality

Buffer Composition:
Most commercial ELOVL6 antibodies are supplied in:

• PBS with 0.02% sodium azide and 50% glycerol pH 7.3

• Some formulations may contain 0.1% BSA (for 20μl sizes)

Aliquoting Recommendations:

• Aliquoting is generally unnecessary for -20°C storage for antibodies in glycerol-containing buffers

• For non-glycerol formulations, create single-use aliquots to minimize freeze-thaw cycles

Handling Precautions:

• Avoid prolonged exposure to high temperatures

• Minimize light exposure, particularly for fluorophore-conjugated antibodies

• Never expose antibodies to strong acids or bases

• Use sterile techniques when handling to prevent microbial contamination

Stability Indicators:
If decreased performance is observed, check for:

• Visible precipitates or turbidity

• Changes in color

• Reduced signal intensity in control samples

• Increased background or non-specific binding