OR13C8 Antibody, HRP conjugated

What is OR13C8 and what functional role does it play in human biology?

OR13C8 (Olfactory Receptor Family 13, Subfamily C, Member 8) is a member of the olfactory receptor protein family. These G-protein coupled receptors are primarily expressed in olfactory sensory neurons and play crucial roles in odor detection and signal transduction. OR13C8 specifically binds to odorant molecules and initiates a signaling cascade that ultimately results in odor perception. Research on these receptors provides insights into olfactory system function, neuronal signaling mechanisms, and sensory processing. The antibody against OR13C8 is valuable for detecting this protein in experimental settings to understand its expression patterns and functions .

What are the key specifications of commercially available OR13C8 Antibody, HRP conjugated?

The commercially available OR13C8 Antibody with HRP conjugation is a polyclonal antibody raised in rabbits against amino acids 260-274 of human OR13C8. It has been purified using Protein G, with purity exceeding 95%. The antibody primarily demonstrates reactivity with human samples. It is specifically designed for ELISA applications, allowing researchers to detect OR13C8 expression levels in various experimental contexts. The HRP (horseradish peroxidase) conjugation enables direct detection through colorimetric or chemiluminescent substrates without requiring secondary antibodies .

What is the cross-reactivity profile of OR13C8 Antibody?

OR13C8 antibodies show confirmed reactivity with human samples across multiple suppliers. Some variants also demonstrate cross-reactivity with mouse samples, making them valuable for comparative studies between human and murine models. The specific HRP-conjugated variant (targeting aa 260-274) has been primarily validated with human samples. When working with other species or variant epitopes, researchers should conduct preliminary specificity testing to confirm cross-reactivity. The peptide-specific nature of these antibodies means their specificity is directly related to sequence conservation across species .

What different conjugates are available for OR13C8 antibodies?

OR13C8 antibodies are available in multiple conjugated forms beyond HRP, including:

Custom conjugation services are also available for OR13C8 antibodies, offering a wide range of potential labels including additional fluorophores (AF350, AF488, AF555, AF594, AF647, etc.), proteins (Alkaline Phosphatase, Streptavidin), and tandem dyes (APC, PE, PE/Cy5, etc.). The selection of conjugate should be determined by the specific detection method and experimental design requirements .

How should appropriate controls be designed when using OR13C8 Antibody, HRP conjugated?

When designing experiments with OR13C8 Antibody, HRP conjugated, include the following controls:

• Positive Control: Utilize samples known to express OR13C8 (human olfactory tissue or confirmed OR13C8-expressing cell lines)

• Negative Control: Include samples known not to express OR13C8 or samples where the protein has been knocked down/out

• Peptide Competition Control: Pre-incubate the antibody with the immunizing peptide (aa 260-274) to block specific binding, as demonstrated in Western blot analyses where blocking with immunizing peptide eliminates the specific signal

• Isotype Control: Include rabbit IgG at equivalent concentration to assess non-specific binding

• Secondary Antibody Control: For non-direct detection methods, include a control omitting the primary antibody

These controls allow proper evaluation of assay specificity and sensitivity, helping distinguish true signals from background or non-specific interactions .

What strategies should be employed for optimizing OR13C8 detection in ELISA?

For optimizing OR13C8 detection in ELISA using HRP-conjugated antibodies:

• Antibody Titration: Test a range of antibody dilutions (e.g., 1:500, 1:1000, 1:2000) to determine optimal signal-to-noise ratio

• Sample Preparation: For cell/tissue lysates, use extraction buffers containing protease inhibitors to prevent degradation

• Blocking Optimization: Test different blocking agents (BSA, non-fat milk, commercial blockers) at various concentrations (3-5%)

• Incubation Parameters: Optimize incubation time (1-2 hours at room temperature vs. overnight at 4°C) and temperature

• Substrate Selection: Compare different HRP substrates (TMB, ABTS, OPD) for optimal sensitivity and signal development time

• Wash Protocol: Perform sufficient wash steps (typically 3-5 washes) with appropriate buffers (PBS-T or TBS-T)

Systematic optimization of these parameters helps achieve maximum sensitivity while maintaining specificity in ELISA applications .

How does epitope selection (aa 260-274) affect OR13C8 antibody specificity?

The epitope selection of amino acids 260-274 in OR13C8 has significant implications for antibody performance:

• Domain Location: This region represents part of the extracellular/transmembrane domain of the G-protein coupled receptor

• Sequence Uniqueness: The specificity is determined by the uniqueness of this sequence compared to other olfactory receptors

• Structural Accessibility: This epitope appears accessible in native and denatured forms, enabling detection in multiple applications

• Cross-Reactivity Implications: Antibodies targeting this region show reactivity with human OR13C8 specifically

• Alternative Epitopes: Other available antibodies target different regions (e.g., aa 271-320), which may affect detection in different experimental contexts

Researchers should consider these factors when selecting between different epitope-specific antibodies for OR13C8 detection. The 260-274 epitope appears to provide good specificity while maintaining application versatility .

What are the recommended storage and handling procedures for OR13C8 Antibody, HRP conjugated?

To maintain optimal activity of OR13C8 Antibody, HRP conjugated:

• Storage Temperature: Store at -20°C for long-term storage or at 4°C for short-term use (typically up to 2 weeks)

• Aliquoting: Upon receipt, divide into small working aliquots to avoid repeated freeze-thaw cycles

• Buffer Composition: The antibody is supplied in a liquid format, typically in a stabilizing buffer

• Freeze-Thaw Cycles: Minimize freeze-thaw cycles (preferably <5) to prevent denaturation and aggregation

• Working Dilution Stability: Diluted antibody solutions are generally stable for up to 24 hours at 4°C

• Light Sensitivity: Protect from prolonged exposure to light, particularly if using fluorescent detection substrates

• Expiration: Most antibodies maintain activity for at least 12 months when properly stored

Following these storage and handling guidelines will help preserve antibody performance throughout the research project timeline .

What is the recommended protocol for using OR13C8 Antibody, HRP conjugated in ELISA?

Recommended ELISA Protocol for OR13C8 Antibody, HRP conjugated:

• Plate Coating:

  • Coat 96-well plate with target capture antibody or antigen

  • Incubate overnight at 4°C

• Blocking:

  • Block with 3-5% BSA or non-fat milk in PBST for 1-2 hours at room temperature

• Sample Addition:

  • Add samples and standards diluted in blocking buffer

  • Incubate for 2 hours at room temperature

• Primary Antibody:

  • Add OR13C8 Antibody, HRP conjugated at 1:1000 dilution

  • Incubate for 1-2 hours at room temperature

• Washing:

  • Wash 5 times with PBST (PBS + 0.05% Tween-20)

• Detection:

  • Add TMB substrate solution

  • Incubate for 15-30 minutes at room temperature in the dark

• Stop Reaction:

  • Add stop solution (usually 2N H₂SO₄)

• Measurement:

  • Read absorbance at 450nm with 570nm reference

This protocol should be optimized for specific experimental conditions, sample types, and desired sensitivity requirements .

How can researchers troubleshoot high background when using OR13C8 Antibody, HRP conjugated?

When encountering high background with OR13C8 Antibody, HRP conjugated, consider these troubleshooting approaches:

• Antibody Concentration: Dilute the antibody further (e.g., 1:2000 instead of 1:1000)

• Blocking Optimization:

  • Increase blocking agent concentration (5% instead of 3%)

  • Try alternative blocking agents (e.g., switch from BSA to casein)

  • Extend blocking time (2-3 hours instead of 1 hour)

• Washing Improvements:

  • Increase wash buffer stringency (add 0.1% SDS or increase Tween-20 to 0.1%)

  • Increase number of wash cycles (6-8 instead of 3-5)

  • Extend wash durations (1-2 minutes per wash)

• Buffer Additives:

  • Add 0.1-0.5% non-ionic detergent to antibody dilution buffer

  • Include 1-5% normal serum from the same species as samples

• Substrate Handling:

  • Reduce substrate incubation time

  • Prepare fresh substrate solution

  • Store substrate protected from light

• Sample Preparation:

  • Pre-clear lysates by centrifugation at higher speeds

  • Pre-absorb antibody with non-specific proteins

Systematic evaluation of these factors typically resolves background issues while maintaining specific signal detection .

What are the optimal substrate options for OR13C8 Antibody, HRP conjugated?

For OR13C8 Antibody with HRP conjugation, several substrate options are available with varying sensitivity and detection characteristics:

TMB (3,3',5,5'-Tetramethylbenzidine) is most commonly used for standard ELISA applications due to its good sensitivity and ease of use. For particularly low abundance targets, chemiluminescent substrates offer enhanced sensitivity. Selection should be based on required sensitivity, available detection instruments, and throughput needs .

What considerations are important when designing quantitative analyses with OR13C8 Antibody?

For quantitative analysis using OR13C8 Antibody, HRP conjugated:

• Standard Curve Generation:

  • Use recombinant OR13C8 protein as standard

  • Prepare 7-8 point standard curve with 2-fold serial dilutions

  • Include zero standard (blank)

• Sample Preparation Consistency:

  • Standardize cell/tissue lysis procedures

  • Normalize protein concentrations across samples (use BCA or Bradford assay)

  • Prepare samples in identical buffer conditions

• Assay Validation:

  • Determine limit of detection (LOD) and quantification (LOQ)

  • Assess linearity of dilution for actual samples

  • Evaluate intra-assay and inter-assay variability (<15% CV)

• Controls:

  • Include internal controls with known OR13C8 concentration

  • Run peptide competition controls to confirm specificity

• Data Analysis:

  • Use 4-parameter logistic regression for standard curve fitting

  • Include quality control criteria (R² > 0.98, recovery 80-120%)

  • Apply appropriate statistical analysis (ANOVA, t-test) for comparing groups

These methodological considerations ensure reliable quantitative results when measuring OR13C8 expression levels across different experimental conditions .

How can OR13C8 Antibody be effectively used in co-immunoprecipitation studies?

For co-immunoprecipitation (Co-IP) studies with OR13C8 Antibody:

• Antibody Selection: Use the unconjugated version of the OR13C8 antibody rather than HRP-conjugated

• Sample Preparation:

  • Lyse cells in non-denaturing buffer (e.g., NP-40 or CHAPS-based)

  • Include protease/phosphatase inhibitors

  • Pre-clear lysate with Protein A/G beads

• Immunoprecipitation Protocol:

  • Incubate 1-5 μg antibody with 500-1000 μg protein lysate overnight at 4°C

  • Add Protein G beads (OR13C8 antibody is rabbit IgG)

  • Wash extensively (4-5 times) with wash buffer of decreasing stringency

• Controls:

  • Include IgG control from same species (rabbit)

  • Include input sample (5-10% of lysate used for IP)

  • Consider reverse Co-IP to confirm interaction

• Detection:

  • Western blot using antibodies against suspected interaction partners

  • Consider mass spectrometry for unbiased interaction discovery

This approach can identify proteins that interact with OR13C8, providing insights into its signaling complexes and molecular functions .

What approaches are recommended for studying OR13C8 expression in tissue samples?

For analyzing OR13C8 expression in tissue samples:

• Immunohistochemistry (IHC):

  • Use unconjugated OR13C8 antibody with appropriate secondary detection

  • Optimize antigen retrieval methods (citrate or EDTA-based)

  • Test multiple fixation protocols (formalin, paraformaldehyde)

  • Include olfactory epithelium as positive control tissue

• Multiplex Immunofluorescence:

  • Combine OR13C8 detection with neuronal markers (TUJ1, MAP2)

  • Use appropriate fluorophore-conjugated secondary antibodies

  • Include DAPI for nuclear counterstaining

  • Apply spectral unmixing for multi-color analysis

• RNA Analysis Correlation:

  • Perform RNA-seq or qRT-PCR on adjacent tissue sections

  • Correlate protein expression with mRNA levels

  • Consider single-cell approaches for heterogeneous tissues

• Quantification Methods:

  • Use digital pathology software for quantitative analysis

  • Measure intensity, distribution, and co-localization parameters

  • Apply appropriate statistical analyses for group comparisons

These complementary approaches provide comprehensive understanding of OR13C8 expression patterns in different tissue contexts and physiological/pathological states .

How should researchers analyze the specificity of OR13C8 Antibody across different experimental conditions?

To thoroughly validate OR13C8 Antibody specificity:

• Peptide Competition Assay:

  • Pre-incubate antibody with excess immunizing peptide (aa 260-274)

  • Run treated and untreated antibody in parallel

  • Specific signals should be abolished in peptide-blocked samples

  • This approach has been demonstrated successfully in Western blot analyses of OR13C8

• Genetic Knockdown/Knockout Validation:

  • Use siRNA, shRNA, or CRISPR/Cas9 to reduce OR13C8 expression

  • Compare antibody signal in control vs. knockdown/knockout samples

  • Specific signal should decrease proportionally to knockdown efficiency

• Recombinant Protein Controls:

  • Test antibody against recombinant OR13C8 protein

  • Include related olfactory receptors to assess cross-reactivity

  • Determine antibody sensitivity and dynamic range

• Cross-Application Validation:

  • Confirm specificity across multiple techniques (ELISA, WB, IF)

  • Compare results obtained with antibodies targeting different epitopes

  • Correlate with mRNA expression data where possible

• Mass Spectrometry Validation:

  • Immunoprecipitate OR13C8 and analyze by mass spectrometry

  • Confirm presence of OR13C8 peptides in the immunoprecipitated material

This comprehensive validation ensures that experimental results truly reflect OR13C8 biology rather than non-specific interactions .

What are the considerations for using OR13C8 Antibody in olfactory receptor trafficking studies?

When investigating OR13C8 trafficking:

• Subcellular Fractionation:

  • Separate membrane, cytosolic, and nuclear fractions

  • Use Western blotting with OR13C8 antibody to detect distribution

  • Include appropriate markers for each fraction (Na⁺/K⁺-ATPase for membrane, GAPDH for cytosol)

• Live Cell Imaging:

  • Consider using OR13C8 antibody with cell-permeable fluorescent tags

  • Alternatively, create OR13C8-fluorescent protein fusions

  • Employ time-lapse microscopy to track dynamic trafficking

• Pulse-Chase Experiments:

  • Label surface proteins with biotin or specific tags

  • Track internalization and recycling kinetics of OR13C8

  • Quantify surface vs. internalized receptor pools

• Co-localization Studies:

  • Combine OR13C8 antibody with markers for different cellular compartments:

    • Endoplasmic reticulum (calnexin)

    • Golgi apparatus (GM130)

    • Endosomes (Rab5, Rab7, Rab11)

    • Lysosomes (LAMP1)

  • Calculate co-localization coefficients (Pearson's, Mander's)

• Stimulus-Dependent Trafficking:

  • Monitor OR13C8 localization changes upon odorant exposure

  • Assess receptor internalization and desensitization kinetics

  • Quantify recycling vs. degradation pathways

These approaches provide comprehensive understanding of OR13C8 trafficking in response to stimuli and during receptor lifecycle .

How can researchers differentiate between specific and non-specific binding in OR13C8 immunoassays?

To distinguish specific from non-specific binding:

• Titration Analysis:

  • Perform antibody titration series (1:100 to 1:10,000)

  • Plot signal-to-noise ratio against antibody concentration

  • Identify optimal concentration where specific signal is maximized while background remains low

• Peptide Competition Controls:

  • Pre-incubate antibody with excess immunizing peptide (aa 260-274)

  • Compare signal intensity with and without peptide competition

  • Specific signals should be significantly reduced or eliminated

  • This approach has been demonstrated successfully in Western blot analyses

• Cross-Absorption Techniques:

  • Pre-absorb antibody with tissues/cells lacking OR13C8 expression

  • Remove antibodies binding to non-specific epitopes

  • Compare results with non-absorbed antibody

• Statistical Analysis:

  • Apply methods to differentiate signal from background (z-score calculation)

  • Establish clear criteria for positive vs. negative results

  • Use consistent thresholds across experiments

• Alternative Detection Methods:

  • Compare results across different detection systems

  • Validate findings with orthogonal techniques (PCR, mass spectrometry)

  • Use multiple antibodies targeting different OR13C8 epitopes

These methodological approaches ensure robust discrimination between specific OR13C8 detection and experimental artifacts, enhancing data reliability and interpretation .