SCN10A Antibody, HRP conjugated

What is SCN10A and why is it an important research target?

SCN10A encodes the sodium channel protein type 10 subunit alpha (Nav1.8), a tetrodotoxin-resistant voltage-gated sodium channel that mediates voltage-dependent sodium ion permeability in excitable membranes. This channel assumes opened or closed conformations in response to voltage differences across membranes, creating a sodium-selective pathway through which sodium ions may pass according to their electrochemical gradient . SCN10A plays a crucial role in neuropathic pain mechanisms, making it a significant target for pain research . Recent studies have also revealed its importance in cardiac function through a shortened transcript variant, with genetic variations in and around SCN10A strongly associated with atrial fibrillation, Brugada syndrome, cardiac conduction velocities, and heart rate regulation .

What applications is the SCN10A Antibody, HRP conjugated suitable for?

The HRP-conjugated SCN10A antibody is optimized for enzyme-linked applications. Based on the search results, this conjugated antibody has been validated for:

While the HRP-conjugated version is specifically designed for applications where direct enzymatic detection is advantageous, the unconjugated version of SCN10A antibody has been validated for additional applications:

The HRP conjugation provides direct enzymatic detection capability, eliminating the need for secondary antibody incubation steps in protocols like ELISA and certain immunohistochemical techniques .

What are the optimal storage and handling conditions for SCN10A Antibody, HRP conjugated?

For maximum stability and activity retention of the SCN10A Antibody, HRP conjugated, the following storage and handling guidelines should be followed:

• Store at -20°C when not in use

• The antibody is typically shipped with ice packs; upon receipt, store immediately at the recommended temperature

• For unconjugated forms that ship as lyophilized powder at room temperature, store at -20°C upon arrival

• Reconstituted solutions can be stored at 4°C, protected from light, for up to 1 week

• For longer periods, create small aliquots and store at -20°C

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

• Centrifuge all antibody preparations before use (10,000 x g for 5 minutes)

Proper storage ensures the retention of conjugated HRP enzymatic activity and antibody binding capacity, which are critical for experimental success.

What dilution ranges are recommended for SCN10A Antibody in different applications?

Optimal dilution ranges vary by application and sample type. Based on the search results:

It is strongly recommended to perform titration experiments in each testing system to obtain optimal results, as dilution requirements may be sample-dependent . The optimal antibody concentration balances strong specific signal with minimal background, which must be empirically determined for each experimental context.

How can researchers distinguish SCN10A-short transcript expression from full-length SCN10A when using antibodies?

Recent research has revealed that cardiomyocytes express a short transcript comprising only the last 7 exons of the SCN10A gene (SCN10A-short), while the full-length SCN10A transcript was undetectable in human and mouse heart tissue . This finding has significant implications for cardiac research using SCN10A antibodies.

To distinguish between these transcript variants:

• Epitope mapping: Determine if the antibody's epitope is within the region encoded by the last 7 exons (shared by both variants) or in the N-terminal region (unique to full-length). The SCN10A Antibody (55334-1-AP) is generated against a peptide immunogen , so checking the exact epitope location is crucial.

• Western blot analysis: The calculated molecular weight of full-length SCN10A is 221 kDa , while SCN10A-short would have a significantly lower molecular weight. Running parallel samples from neural tissue (expressing full-length) and cardiac tissue (expressing short variant) could help distinguish the variants based on size.

• RT-PCR with specific primers: Design primers that target either unique regions of the full-length transcript or the junction points specific to SCN10A-short for transcript verification before antibody-based experiments.

• Tissue controls: Include dorsal root ganglion (DRG) samples as positive controls for full-length SCN10A and atrial or ventricular conduction system samples for SCN10A-short .

When interpreting immunolabeling results in cardiac tissue, researchers should consider that signals likely represent SCN10A-short rather than full-length protein, which has important functional implications for data interpretation .

What methodological approaches can optimize signal specificity when using SCN10A Antibody, HRP conjugated?

Achieving high signal specificity with HRP-conjugated antibodies requires careful experimental design and rigorous controls:

How do genetic variants in SCN10A affect antibody binding and experimental interpretation?

Genetic variants in and around SCN10A have been associated with cardiac abnormalities including atrial fibrillation, Brugada syndrome, and altered cardiac conduction . These variants can potentially affect antibody recognition and experimental results in several ways:

To address these challenges:

What are the critical differences between neuronal and cardiac SCN10A detection that researchers should consider?

The discovery of differential SCN10A expression patterns between neuronal and cardiac tissues presents important methodological considerations:

• Transcript variant targeting:

  • Neuronal tissues express full-length SCN10A transcript

  • Cardiac tissues express primarily SCN10A-short transcript comprising only the last 7 exons of the gene

  • Antibodies targeting N-terminal regions may fail to detect cardiac SCN10A-short

• Cell type specificity in cardiac tissue:

  • SCN10A-short is expressed in cardiomyocytes of the atria, sinoatrial node, and ventricular conduction system

  • Expression patterns are heterogeneous, requiring careful selection of cardiac regions for analysis

• Functional interactions:

  • In neurons, SCN10A functions as an independent sodium channel

  • In cardiac tissue, SCN10A-short appears to interact with and enhance Nav1.5-mediated sodium current

  • Experimental designs should account for this interaction, potentially including co-immunoprecipitation or proximity ligation assays

• Appropriate controls by tissue type:

  Tissue Type	Positive Control	Negative Control
  Neuronal	Dorsal root ganglia	Central nervous system regions with low SCN10A expression
  Cardiac	Atrial tissue, sinoatrial node	Working ventricular myocardium (low expression)

• Protocol optimizations by tissue type:

  Parameter	Neuronal Tissue	Cardiac Tissue
  Fixation	Brief fixation (4-6 hours)	Standard fixation (12-24 hours)
  Antigen Retrieval	Mild (citrate buffer)	Strong (EDTA buffer, pH 9.0)
  Primary Antibody Incubation	Short (overnight at 4°C)	Extended (36-48 hours at 4°C)
  Detection System	Direct visualization often sufficient	May require amplification for SCN10A-short

Understanding these tissue-specific differences is crucial for proper experimental design and interpretation of results when studying SCN10A in different physiological contexts.

What strategies can address common troubleshooting issues with HRP-conjugated SCN10A antibodies?

When working with HRP-conjugated SCN10A antibodies, researchers may encounter several technical challenges. Here are evidence-based troubleshooting approaches:

• High background signal

  • Increase blocking duration (2-3 hours) and concentration (5% BSA or serum)

  • Add 0.1-0.3% Triton X-100 to wash buffers to reduce non-specific binding

  • Optimize HRP quenching with 0.3-3% H₂O₂ for 10-30 minutes before antibody application

  • Reduce antibody concentration; for HRP-conjugated SCN10A antibody, try serial dilutions beyond the recommended range

• Weak or no signal

  • Confirm sample expression: SCN10A has tissue-specific expression, with validated detection in SH-SY5Y and HeLa cells for the unconjugated version

  • Optimize antigen retrieval: Compare heat-mediated (citrate or EDTA buffers) and enzymatic methods

  • Check HRP activity: Include a positive control for HRP activity separate from the antibody specificity control

  • Extend primary antibody incubation time (24-48 hours at 4°C)

  • Assess impact of storage conditions: HRP activity can diminish over time, particularly with repeated freeze-thaw cycles

• Non-specific or unexpected banding patterns in Western blot

  • The calculated molecular weight of full-length SCN10A is 221 kDa , while SCN10A-short would appear at a lower molecular weight

  • Run reduced and non-reduced samples in parallel to identify potential disulfide-linked complexes

  • Include cardiac tissue (for SCN10A-short) and neuronal tissue (for full-length) controls

  • Use gradient gels (4-12%) to improve resolution of high molecular weight proteins

• Signal variability between experiments

  • Standardize cell/tissue processing: fixation duration, buffer composition, and pH

  • Prepare master mixes of antibody dilutions for technical replicates

  • Monitor the temperature during all incubation steps

  • For samples with variable endogenous biotin or peroxidase, implement appropriate blocking steps

• Cross-reactivity concerns

  • The HRP-conjugated SCN10A antibody is validated for human reactivity

  • For studies in other species, preliminary validation is essential as cross-reactivity is not guaranteed

  • Consider epitope sequence conservation analysis across species before application

How can SCN10A Antibody, HRP conjugated be used to investigate cardiac channelopathies?

Recent research has established that genetic variants in SCN10A modulate enhancer function and expression of a cardiac-specific SCN10A-short transcript, impacting Nav1.5-mediated sodium current and heart rhythm . This connection to cardiac function opens important research applications for SCN10A antibodies in studying channelopathies:

• Expression analysis in patient samples:

  • Compare SCN10A-short expression levels in atrial tissue samples from patients with and without arrhythmias

  • Correlate expression patterns with specific genetic variants, particularly those in linkage disequilibrium with rs6801957 in the intronic enhancer

  • Develop IHC scoring systems for SCN10A positivity in cardiac conduction system tissues

• Co-localization studies:

  • Use dual-labeling approaches to examine SCN10A-short and Nav1.5 co-localization in cardiac tissues

  • Implement proximity ligation assays to confirm physical interaction between these channels

  • Compare distribution patterns in normal vs. diseased cardiac tissue

• Genotype-phenotype correlations:

  • Stratify immunohistochemical analyses based on patient genotypes at SCN10A loci

  • Develop tissue microarrays from genotyped cardiac samples to enable high-throughput analysis

  • Correlate antibody staining patterns with clinical parameters such as PR interval, QRS duration, and arrhythmia susceptibility

• Functional validation:

  • Combine immunolabeling with patch-clamp electrophysiology in the same samples

  • Use SCN10A antibodies to confirm expression in systems where pharmacological modulation of the channel is being tested

  • Develop in vitro models that recapitulate the genetic variants affecting SCN10A expression

When designing such studies, researchers should consider that the HRP-conjugated format offers advantages for quantitative applications like ELISA and chromogenic IHC, while fluorescently-labeled secondary antibodies might be preferable for co-localization studies.

What considerations are important when using SCN10A Antibody, HRP conjugated in multiplex detection systems?

Multiplex detection involving HRP-conjugated SCN10A antibody requires careful planning to prevent signal interference and ensure accurate results:

• HRP signal separation in chromogenic multiplex IHC:

  • When using multiple HRP-conjugated antibodies, sequential detection with complete HRP inactivation between rounds is necessary

  • Implement microwave treatment (10 minutes in citrate buffer) or 2% sodium azide treatment to inactivate HRP before introducing the next HRP-conjugated antibody

  • Use spectrally distinct chromogens (DAB, Vector® VIP, Vector® SG) for different targets

  • Consider tyramide signal amplification (TSA) with different fluorophores that can later be unmixed spectrally

• Antibody panel design:

  • When studying SCN10A in cardiac tissue, consider including markers for:

    • Cardiomyocyte identification (cardiac troponin T)

    • Conduction system components (HCN4 for sinoatrial node, Cx40 for fast-conducting pathways)

    • Other sodium channels, particularly Nav1.5 (SCN5A)

  • For neuronal studies, include markers for:

    • Sensory neuron subtypes (TRPV1, CGRP)

    • Myelination status (MBP)

    • Other pain-associated channels (TRPA1)

• Signal order optimization:

  • In sequential multiplex protocols, detect the least abundant target (potentially SCN10A-short in some tissues) first

  • For targets with overlapping distribution, use spectrally distinct detection systems

  • When using both HRP and alkaline phosphatase (AP) systems together, typically apply the HRP system first

• Controls for multiplex systems:

  • Single-stain controls for each antibody in the panel

  • Minus-one controls (omitting one antibody at a time)

  • Absorption controls for each antibody

  • Isotype controls matched to each primary antibody

• Data analysis approaches:

  • For quantitative colocalization, utilize Manders' or Pearson's correlation coefficients

  • Consider supervised machine learning approaches for pattern recognition in complex multiplex images

  • Implement tissue cytometry for single-cell quantification of multiple markers

How does the HRP conjugation affect the performance of SCN10A antibody compared to unconjugated versions?

The HRP conjugation of SCN10A antibody introduces specific performance characteristics that differ from unconjugated versions:

• Sensitivity and detection threshold:

  • HRP conjugation provides enzymatic amplification, potentially increasing sensitivity for low-abundance targets

  • Signal-to-noise ratio may be enhanced through optimized substrate reaction times

  • The direct conjugation eliminates signal loss that can occur with secondary antibody binding steps

• Methodological advantages and limitations:

  Parameter	HRP-Conjugated	Unconjugated
  Protocol Length	Shorter (no secondary incubation)	Longer (requires secondary antibody)
  Signal Amplification	Fixed by conjugation ratio	Adjustable through secondary antibody concentration
  Multiplexing Flexibility	Limited by enzyme	Higher (through secondary antibody species/isotypes)
  Background Potential	May be higher due to direct detection	Can be lower with optimized secondaries
  Storage Stability	Lower (HRP activity can diminish)	Higher (unconjugated antibodies more stable)

• Application-specific considerations:

  • ELISA: HRP-conjugated antibodies allow for direct detection systems, reducing protocol steps and improving reproducibility

  • Flow cytometry: Direct HRP conjugation may limit fluorescent multiplexing options compared to unconjugated versions

  • IHC/ICC: While both formats work, unconjugated versions offer more flexibility in detection system selection

• Optimization parameters that differ between formats:

  • HRP-conjugated versions require optimization of substrate development time rather than secondary antibody dilution

  • Quenching of endogenous peroxidases becomes more critical with HRP-conjugated primary antibodies

  • Amplification systems (like tyramide) interact differently with direct HRP conjugates versus HRP-labeled secondary antibodies

• Batch-to-batch variations:

  • HRP-conjugated antibodies may show more variability related to the degree of labeling and HRP activity

  • Critical to validate each new lot with consistent positive controls

  • May require adjustment of dilution factors between lots to maintain signal consistency

What are the most effective complementary techniques to validate findings obtained with SCN10A Antibody, HRP conjugated?

• Molecular validation approaches:

  • RT-qPCR to quantify SCN10A transcript levels (with isoform-specific primers for SCN10A-short)

  • RNAscope® in situ hybridization to visualize transcript localization

  • Western blotting using antibodies targeting different epitopes of SCN10A

  • CRISPR/Cas9 knockout validation, particularly in cell line models

• Functional validation techniques:

  • Patch-clamp electrophysiology to measure sodium currents

  • Multi-electrode array (MEA) recordings for cardiac tissue preparations

  • Calcium imaging as an indirect measure of excitability

  • Pharmacological manipulation using specific Nav1.8 inhibitors (e.g., A-803467, PF-01247324)

• Alternative antibody approaches:

  • Use antibodies from different host species targeting different epitopes

  • Compare polyclonal vs. monoclonal antibody staining patterns

  • Test different detection systems (chromogenic vs. fluorescent)

  • Implement proximity ligation assays (PLA) to confirm protein interactions

• Genetic correlation:

  • Genotype samples for known SCN10A variants, particularly those in the intronic enhancer (e.g., rs6801957)

  • Compare antibody staining patterns across different genotypes

  • Correlate findings with expression quantitative trait loci (eQTL) data

• Integration of computational approaches:

  • Utilize protein structure prediction to understand epitope accessibility

  • Implement machine learning for unbiased pattern recognition in imaging data

  • Develop systems biology models that integrate SCN10A expression with functional outcomes

How can SCN10A Antibody, HRP conjugated be utilized in studying the link between cardiac and neuronal sodium channelopathies?

Recent discoveries about SCN10A's dual role in both pain pathways and cardiac function open intriguing research directions using SCN10A antibodies:

• Comparative tissue analysis:

  • Implement parallel staining of cardiac and peripheral nervous system tissues from the same individuals

  • Compare SCN10A expression patterns in patients with combined phenotypes (e.g., neuropathic pain and cardiac arrhythmias)

  • Develop quantitative image analysis protocols to standardize comparisons across tissue types

• Transgenic model characterization:

  • Analyze expression patterns in models with genetic modification of the intronic enhancer, as described in recent research

  • Compare cardiac and neural SCN10A distribution in these models

  • Correlate antibody staining with electrophysiological phenotypes in both tissue types

• Clinical correlations in patient cohorts:

  • Develop tissue biobanks from patients with channelopathies affecting both systems

  • Implement standardized immunohistochemical scoring systems for SCN10A positivity

  • Correlate antibody staining patterns with:

    • Cardiac phenotypes (conduction parameters, arrhythmia susceptibility)

    • Neurological phenotypes (pain sensitivity, small fiber neuropathy markers)

• Pharmacological studies:

  • Use SCN10A antibodies to confirm target engagement in drug development pipelines

  • Compare effects of Nav1.8 modulators on cardiac versus neuronal tissue

  • Develop predictive biomarkers for treatment response based on expression patterns

• Single-cell resolution approaches:

  • Implement multiplex immunofluorescence with digital spatial profiling

  • Correlate SCN10A protein expression with single-cell transcriptomics

  • Develop spatial transcriptomics approaches to map full-length versus SCN10A-short expression patterns

This research direction has significant therapeutic implications, as it could identify shared mechanisms underlying seemingly distinct neurological and cardiac conditions, potentially leading to unified treatment approaches.

What novel methodological approaches can enhance the specificity and sensitivity of SCN10A detection in complex tissue samples?

As research into SCN10A advances, several cutting-edge methodological approaches can be implemented to enhance detection specificity and sensitivity:

• Super-resolution microscopy approaches:

  • Implement STORM or PALM microscopy to visualize SCN10A distribution at nanoscale resolution

  • Use expansion microscopy to physically enlarge specimens for improved visualization of channel clustering

  • Apply stimulated emission depletion (STED) microscopy for live-cell imaging of SCN10A dynamics

• Proximity-based detection methods:

  • Develop proximity ligation assays (PLA) to confirm SCN10A-short interaction with Nav1.5 in cardiac tissue

  • Implement FRET-based approaches to study dynamic interactions between sodium channel subunits

  • Apply BioID or APEX2 proximity labeling to identify the complete interactome of SCN10A in different contexts

• Tissue clearing and 3D imaging:

  • Apply CLARITY, iDISCO, or CUBIC clearing methods to visualize SCN10A distribution in intact tissue volumes

  • Develop computational approaches to quantify spatial relationships with other channel proteins in 3D

  • Implement light-sheet microscopy for rapid volumetric imaging of cleared tissues

• Artificial intelligence-enhanced analysis:

  • Develop deep learning algorithms for automated quantification of SCN10A staining patterns

  • Implement segment anything models (SAM) for automated identification of positive cells

  • Create digital pathology workflows for standardized scoring across laboratories

• Multi-omic integration approaches:

  • Combine SCN10A immunolabeling with in situ RNA detection (e.g., MERFISH, seqFISH)

  • Integrate spatial transcriptomics data with protein-level detection

  • Develop computational frameworks to correlate genotype, expression patterns, and functional readouts

These advanced methodologies can help resolve long-standing questions about SCN10A localization and function, particularly in tissues with complex cellular compositions like the cardiac conduction system and sensory ganglia.

How should researchers interpret differential expression patterns of SCN10A across various cardiac regions and pathological states?

The interpretation of SCN10A expression patterns in cardiac tissue requires nuanced analysis, particularly given recent discoveries about SCN10A-short and its regional variation:

• Regional expression patterns in normal heart:

  • SCN10A-short is preferentially expressed in the atria, sinoatrial node, and ventricular conduction system

  • Expression is heterogeneous, requiring sampling across multiple regions

  • Standardized mapping approaches should document expression relative to anatomical landmarks

• Interpretational framework for pathological states:

  Expression Change	Potential Interpretation	Validation Approach
  Increased atrial expression	May contribute to atrial arrhythmogenesis	Correlate with electrophysiological parameters
  Altered distribution in conduction system	May affect impulse propagation	Map activation sequences with optical mapping
  Redistribution to working myocardium	May indicate pathological remodeling	Correlate with structural heart disease markers
  Changes in subcellular localization	May reflect trafficking abnormalities	Co-stain with membrane markers and assess colocalization

• Quantification approaches:

  • Develop region-normalized scoring systems that account for baseline differences

  • Implement digital image analysis with machine learning segmentation

  • Consider dual-marker strategies (e.g., SCN10A + connexin isoforms) to identify specific cardiac compartments

• Genotype correlation considerations:

  • Interpret expression changes in the context of patient-specific SCN10A variants

  • Consider that intronic enhancer variants (e.g., rs6801957) modulate SCN10A-short expression

  • Stratify analyses based on known expression quantitative trait loci (eQTL)

• Functional correlation framework:

  • Changes in expression should be interpreted alongside:

    • Action potential characteristics (upstroke velocity, duration)

    • Conduction velocity measurements

    • Arrhythmia susceptibility metrics

    • Response to sodium channel blockers

This interpretive framework acknowledges that SCN10A expression changes may be adaptive or maladaptive depending on context, and integrates genetic, molecular, and functional data for comprehensive understanding.

What future directions might emerge for SCN10A antibody applications in personalized medicine?

The intersection of SCN10A research with personalized medicine presents several promising future directions:

• Pharmacogenomic biomarker development:

  • Establish whether SCN10A expression patterns predict response to sodium channel blockers

  • Develop immunohistochemical scoring systems that correlate with drug efficacy

  • Create patient-derived organoid models for personalized drug screening, validated with SCN10A antibodies

• Risk stratification applications:

  • Determine whether SCN10A expression patterns in accessible samples correlate with arrhythmia risk

  • Develop standardized reporting systems for SCN10A immunohistochemistry in cardiac biopsies

  • Integrate SCN10A expression data with genetic risk scores for enhanced prediction

• Therapeutic monitoring approaches:

  • Use SCN10A antibodies to track changes in expression during disease progression

  • Monitor effects of interventions on channel expression and distribution

  • Develop in vivo imaging approaches using labeled antibody fragments

• Precision targeting of therapy:

  • Identify patient subgroups with specific SCN10A expression patterns that might benefit from tailored interventions

  • Develop therapeutic antibodies or antibody-drug conjugates targeting specific conformations or variants of SCN10A

  • Create nanotechnology-based delivery systems for selective targeting of tissues with high SCN10A expression

• Integration with digital health platforms:

  • Develop image analysis algorithms for standardized quantification of SCN10A staining

  • Create cloud-based platforms for multi-institutional sharing and analysis of immunohistochemical data

  • Implement machine learning to identify novel patterns correlating expression with clinical outcomes

These approaches could transform SCN10A from a research target to a clinically actionable biomarker, particularly in the fields of electrophysiology, pain management, and channelopathy diagnosis and treatment.