OR6C70 Antibody

What is OR6C70 protein and why is it studied?

OR6C70 (Olfactory receptor family 6 subfamily C member 70) is a member of the G-protein coupled receptor 1 family that functions as an odorant receptor. These receptors interact with odorant molecules in the nose to initiate neuronal responses that trigger smell perception. The olfactory receptor proteins share a 7-transmembrane domain structure with many neurotransmitter and hormone receptors and are responsible for the recognition and G protein-mediated transduction of odorant signals . OR6C70 is located on chromosome 12q13.2 in humans, with a molecular weight of approximately 35 kDa .

For optimal performance, follow these research-based guidelines for OR6C70 antibody storage and handling:

• Store at -20°C for long-term storage (up to 1 year of stability)

• For shorter periods (up to one month), some antibodies can be stored at 2-8°C without detectable loss of activity

• Avoid repeated freeze-thaw cycles that can compromise antibody performance

• Most OR6C70 antibodies are supplied in liquid form in PBS containing 50% glycerol, 0.5% BSA and 0.02% sodium azide

• Some are provided lyophilized and need reconstitution with distilled water to achieve a final concentration of 1 mg/mL

• After reconstitution, aliquot and store to minimize freeze-thaw cycles

Experimental Applications and Methodology

When establishing an experimental system using OR6C70 antibodies, proper controls are critical:

• Hep G2 cell lysate has been validated as a positive control for OR6C70 detection by Western blot

• Based on gene expression data, tissue from olfactory epithelium would be appropriate, though specific validation data is limited in the search results

• For knockout/knockdown validation, OR6C70 siRNA (h): sc-95740, OR6C70 shRNA Plasmid (h): sc-95740-SH, and OR6C70 shRNA (h) Lentiviral Particles: sc-95740-V have been reported as suitable tools

• Blocking peptides (e.g., sc-132761 P) can be used in competition studies to confirm antibody specificity

How can I optimize Western blot protocols specifically for OR6C70 detection?

For optimal detection of OR6C70 by Western blot, follow these methodological recommendations compiled from multiple technical sources:

• Sample preparation:

  • Use appropriate lysis buffers that preserve membrane proteins (OR6C70 is a multi-pass membrane protein)

  • Include protease inhibitors to prevent degradation during extraction

  • Avoid excessive heating which may cause aggregation of membrane proteins

• Gel electrophoresis and transfer:

  • Use a gel percentage appropriate for detecting a 35 kDa protein (typically 10-12% SDS-PAGE)

  • Ensure complete transfer of proteins to membrane, especially for hydrophobic membrane proteins

• Blocking and antibody incubation:

  • For primary antibody: Use dilutions in the range of 1:500-1:2000

  • For secondary antibody: For rabbit-host primaries, use anti-rabbit IgG-HRP at 1:2000-1:100,000

  • For goat-host primaries (e.g., sc-132761), use donkey anti-goat IgG-HRP: sc-2020 (dilution range: 1:2000-1:100,000)

  • TBS Blotto A is recommended as a blocking reagent for some OR6C70 antibodies

• Detection:

  • Both chemiluminescent and colorimetric detection methods are suitable

  • Expected molecular weight of OR6C70 is approximately 35 kDa

How do polyclonal and monoclonal OR6C70 antibodies compare in terms of specificity and research applications?

A critical analysis of polyclonal versus monoclonal OR6C70 antibodies reveals important considerations for experimental design:

Polyclonal OR6C70 Antibodies:

• Recognize multiple epitopes on the OR6C70 protein, potentially providing stronger signals through binding multiple sites

• Most commonly raised in rabbits against synthetic peptides derived from human OR6C70 (typically from C-terminal regions)

• Generally recommended for Western blot and ELISA applications across multiple sources

• Higher potential for cross-reactivity with related olfactory receptors due to family sequence homology

• May exhibit batch-to-batch variation in epitope recognition patterns

Monoclonal OR6C70 Antibodies:

• Recognize single epitopes, providing higher specificity but potentially lower sensitivity

• Available monoclonal antibodies (e.g., YP-mAb-13616) are typically mouse-derived

• Offer more consistent results between experiments due to clonal nature

• May be less affected by background interference in complex samples

• Potentially more suitable for quantitative applications requiring high reproducibility

For research requiring high specificity discrimination between closely related olfactory receptors, monoclonal antibodies may be preferable, while polyclonals might offer advantages for detection of low-abundance targets or in applications where conformational changes might affect epitope accessibility.

What are the challenges in detecting endogenous OR6C70 expression and how can they be addressed?

Detecting endogenous OR6C70 presents several methodological challenges:

• Low expression levels: Olfactory receptors often show tissue-specific expression patterns with relatively low abundance

  • Solution: Consider signal amplification methods such as tyramide signal amplification for immunohistochemistry/immunofluorescence applications

  • Use sensitive detection reagents and optimize exposure times for Western blots

• Membrane protein detection issues: As a 7-transmembrane protein, OR6C70 can be difficult to extract and maintain in its native conformation

  • Solution: Optimize lysis conditions using detergents suitable for membrane proteins (e.g., RIPA or specialized membrane protein extraction buffers)

  • Consider native PAGE techniques for applications requiring conformational epitopes

• Cross-reactivity with related receptors: The olfactory receptor family is the largest gene family in the genome

  • Solution: Validate antibody specificity using blocking peptides

  • Consider complementary detection methods like RT-PCR to confirm expression at the mRNA level

  • Where possible, use negative controls from tissues known not to express OR6C70

• Limited validation data: The search results indicate relatively limited published validation for most commercial OR6C70 antibodies

  • Solution: Conduct thorough validation experiments including positive and negative controls

  • Consider using recombinant OR6C70 expression systems for initial antibody characterization

What experimental approaches can be used to study OR6C70 functionality in olfactory signaling pathways?

For researchers investigating the functional role of OR6C70 in olfactory signaling, consider these methodological approaches:

• Calcium imaging assays:

  • Transfect cells with OR6C70 expression constructs and calcium indicators

  • Challenge with potential odorant ligands to detect receptor activation

  • Compare response profiles to related olfactory receptors

• Receptor trafficking and localization studies:

  • Use OR6C70 antibodies for immunofluorescence to track receptor localization

  • Create fluorescently tagged OR6C70 constructs for live-cell imaging

  • Examine co-localization with trafficking machinery components

• Knockdown/knockout approaches:

  • Utilize siRNA (e.g., sc-95740) or shRNA constructs (e.g., sc-95740-SH) to reduce OR6C70 expression

  • CRISPR-Cas9 gene editing to generate OR6C70 knockout models

  • Assess olfactory function in models with reduced OR6C70 expression

• Ligand binding studies:

  • Perform competitive binding assays using labeled putative ligands

  • Molecular docking simulations to predict odorant binding sites

  • Site-directed mutagenesis of predicted binding residues followed by functional characterization

• Downstream signaling analysis:

  • Examine G-protein activation and cAMP production upon OR6C70 stimulation

  • Investigate protein-protein interactions with signaling components

  • Compare signaling kinetics with other characterized olfactory receptors

How can I address non-specific binding or high background when using OR6C70 antibodies?

When encountering non-specific binding or high background with OR6C70 antibodies, consider these research-based troubleshooting approaches:

• For Western blot applications:

  • Increase blocking time/concentration (using TBS Blotto A or similar reagents)

  • Try alternative blocking agents (BSA, non-fat milk, commercial blockers)

  • Optimize primary antibody concentration (test dilutions from 1:500-1:2000)

  • Include competitive blocking with the immunizing peptide where available

  • Increase washing duration and number of washes between antibody incubations

  • Consider using more specific detection methods or higher stringency conditions

• For immunofluorescence/immunocytochemistry:

  • Include appropriate negative controls (secondary antibody alone, non-expressing tissues)

  • Pre-adsorb antibodies with blocking peptides to reduce non-specific binding

  • Use proper antigen retrieval methods if appropriate

  • Optimize antibody concentration (starting with dilutions of 1:50-1:500)

  • Increase washing steps and duration

  • Use specialized blocking reagents to reduce background

• General approaches:

  • Validate antibody specificity using known positive and negative controls

  • Consider using alternative antibodies targeting different epitopes of OR6C70

  • Compare results across different detection methods (e.g., WB vs. IF)

  • Use freshly prepared reagents and avoid antibody contamination

What are the key considerations when interpreting OR6C70 expression data across different experimental systems?

When analyzing and interpreting OR6C70 expression data, researchers should consider these critical factors:

• Expression level variation:

  • Olfactory receptors typically show regulated expression patterns that may vary between tissues, developmental stages, and physiological conditions

  • Compare expression levels to appropriate reference genes when performing quantitative analysis

  • Consider the sensitivity limitations of the detection method used

• Protein vs. mRNA expression correlation:

  • Validate protein expression findings with mRNA data when possible, as post-transcriptional regulation may affect protein abundance

  • Be aware that antibody-based detection measures protein levels while techniques like RT-PCR detect mRNA

• Cross-reactivity considerations:

  • The high sequence homology within olfactory receptor families creates potential for cross-reactivity

  • Interpret single-antibody results cautiously, especially with polyclonal antibodies

  • Consider orthogonal validation methods to confirm specificity

• Subcellular localization interpretation:

  • As a membrane protein, OR6C70 should primarily localize to the plasma membrane and potentially to intracellular membrane compartments during trafficking

  • Unexpected localization patterns may indicate experimental artifacts or reveal novel biology

  • Validate unusual localization patterns using multiple antibodies or tagged expression constructs

• Model system considerations:

  • Expression patterns in cell lines may differ from native tissues

  • Species differences in OR6C70 sequence and expression should be considered when working with non-human models

  • Heterologous expression systems may not recapitulate all aspects of native receptor function and regulation

How can I validate the specificity of OR6C70 antibody staining in tissue sections or cultured cells?

To rigorously validate OR6C70 antibody specificity in experimental systems, implement these methodological approaches:

• Peptide competition assays:

  • Pre-incubate the OR6C70 antibody with the immunizing peptide (e.g., sc-132761 P)

  • Compare staining patterns with and without peptide competition

  • Specific staining should be significantly reduced or eliminated by peptide competition

• Genetic validation approaches:

  • Use siRNA knockdown (e.g., OR6C70 siRNA (h): sc-95740) to reduce expression

  • Compare staining in wild-type vs. knockdown samples

  • Generate knockout models using CRISPR-Cas9 as definitive negative controls

• Multi-antibody validation:

  • Compare staining patterns using antibodies targeting different epitopes of OR6C70

  • Consistent patterns across multiple antibodies increase confidence in specificity

  • Use both polyclonal and monoclonal antibodies when available

• Correlation with other detection methods:

  • Validate protein expression with mRNA detection methods

  • Use in situ hybridization to confirm tissue expression patterns

  • Correlate Western blot data with immunohistochemistry findings

• Positive and negative control tissues:

  • Include tissues known to express OR6C70 (e.g., olfactory epithelium) as positive controls

  • Include tissues known not to express OR6C70 as negative controls

  • For cell lines, Hep G2 has been validated as expressing OR6C70

How are OR6C70 antibodies being used in current olfactory research?

While the search results don't provide specific examples of OR6C70 antibodies in current olfactory research, based on general principles in the field, these antibodies would typically be employed in:

• Receptor expression mapping:

  • Characterizing OR6C70 expression patterns across different regions of olfactory epithelium

  • Examining developmental regulation of OR6C70 expression

  • Investigating species differences in OR6C70 distribution

• Olfactory signal transduction studies:

  • Investigating the coupling of OR6C70 to specific G-proteins

  • Examining receptor internalization and recycling following odorant exposure

  • Studying adaptation mechanisms in olfactory signaling

• Odorant specificity research:

  • Identifying specific odorant ligands that activate OR6C70

  • Characterizing structure-activity relationships for OR6C70 ligands

  • Comparing selectivity profiles with related olfactory receptors

• Cell biology of olfactory neurons:

  • Studying the trafficking of OR6C70 to olfactory cilia

  • Investigating the clustering of olfactory receptors in membrane microdomains

  • Examining receptor turnover and lifetime in olfactory neurons

What methodological advances are improving the specificity and sensitivity of OR6C70 detection?

Recent methodological advances relevant to OR6C70 detection include:

• Antibody technology improvements:

  • Development of monoclonal antibodies with higher specificity

  • Recombinant antibody approaches for improved consistency

  • Enhanced purification methods using epitope-specific immunogens

• Signal amplification techniques:

  • Tyramide signal amplification for immunohistochemistry

  • Proximity ligation assays for protein interaction studies

  • Super-resolution microscopy for precise localization studies

• Complementary validation approaches:

  • Integration with transcriptomic data from resources like the Observed Antibody Space database

  • Mass spectrometry validation of antibody targets

  • CRISPR-based tagging of endogenous proteins for antibody-independent detection

• Computational approaches:

  • Improved epitope prediction for antibody design

  • Cross-reactivity prediction algorithms

  • Structure-based design of highly specific antibodies

How might OR6C70 antibodies contribute to understanding broader olfactory receptor biology?

OR6C70 antibodies can provide valuable insights into broader olfactory receptor biology through: