KCND2 Antibody

What is KCND2 and what cellular functions does it mediate?

KCND2, also known as Kv4.2, is a shal-related voltage-gated potassium channel encoded by the KCND2 gene in humans. It plays crucial roles in neuronal excitability and functions downstream of the metabotropic glutamate receptor GRM5, mediating nociception through GRM5 activation. At the molecular level, Kv4.2 can form functional heterotetrameric channels with Kv4.3, creating diversity in potassium channel complexes .

From a neurophysiological perspective, Kv4.2 channels regulate several aspects of neuronal firing patterns, including:

• Prolonging the latency before the first spike in action potential series

• Regulating the frequency of repetitive action potential firing

• Shortening the duration of action potentials

• Controlling the back-propagation of action potentials from the neuronal cell body to the dendrites

These functions position KCND2 as a critical regulator of neuronal excitability and signaling strength in the central nervous system.

What are the common applications for KCND2 antibodies in research?

KCND2 antibodies are employed in multiple experimental techniques, each offering distinct advantages for specific research questions:

• Immunofluorescence labeling: Recommended dilution of 1:100 for visualizing spatial distribution of Kv4.2 in tissue sections or cultured cells

• Immunohistochemistry (IHC): Used for detection of Kv4.2 in fixed tissue specimens to understand expression patterns across different cell types

• Western blotting: Typically used at 1:500 dilution to detect Kv4.2 protein (predicted band size of 71 kDa) in cell or tissue lysates

• Flow cytometry: Applied at 1:200 dilution for quantitative assessment of Kv4.2 expression in cell populations

The selection of a specific application should be guided by the experimental question, with immunofluorescence providing spatial context, western blotting confirming molecular weight and expression levels, and flow cytometry enabling quantitative single-cell analysis.

How do I select the appropriate KCND2 antibody for my specific experiment?

When selecting a KCND2 antibody, consider the following parameters based on your experimental needs:

• Host species and antibody class: Both rabbit polyclonal (offering multiple epitope recognition) and mouse monoclonal (providing high specificity) antibodies are available

• Target species reactivity: Confirm cross-reactivity with your model system. Available antibodies show reactivity with human, mouse, and rat samples

• Validated applications: Ensure the antibody has been validated for your specific application (IF, IHC, WB, or flow cytometry)

• Immunogen information: For epitope-specific studies, check the immunogen details, such as synthetic peptides derived from specific regions (e.g., C-terminal region) or recombinant fragment proteins (e.g., within aa 1-200)

• Concentration and formulation: Available concentrations range from 0.25 mg/ml purified IgG, with storage recommendations typically at -20°C in PBS (pH 7.2) with 0.1% sodium azide

Selecting antibodies validated specifically for your application and species of interest will significantly improve experimental reproducibility and data reliability.

How can KCND2 antibodies be used to investigate channel mutations associated with neurological disorders?

KCND2 variants have been identified in individuals with early-onset global developmental delay, impaired motor, speech and cognitive development, and developmental epileptic encephalopathy. Using antibodies in mutation research requires sophisticated experimental approaches:

• Mutation-specific detection: For studying specific mutations like p.(Glu323Lys) (E323K), p.(Pro403Ala) (P403A), p.(Val404Leu) (V404L), and p.(Val404Met) (V404M) that affect critical channel gating sites, researchers can use antibodies in combination with expression systems

• Functional correlation studies: Combining immunostaining with electrophysiology to correlate expression levels with functional alterations in channel properties

• Structural analysis: Using antibodies in conjunction with structural biology techniques to understand how specific mutations (particularly those affecting conserved residues) alter protein conformation and interactional properties

• Protein trafficking assessment: Employing antibodies in live-cell imaging experiments to determine whether mutations affect normal trafficking of Kv4.2 to the cell membrane

For investigating the p.(Val404Met) variant previously identified in monozygotic twins with severe, intractable seizures and autism, combined approaches using antibodies and functional assays provide comprehensive insights into pathological mechanisms .

What is the role of KCND2 in cancer progression and how can antibodies help elucidate these mechanisms?

Recent research has revealed significant roles for KCND2 in cancer, particularly gastric cancer. KCND2 antibodies enable researchers to investigate these mechanisms through several experimental approaches:

• Expression correlation studies: KCND2 is markedly elevated in gastric cancer, with expression correlating with different grades, T stages, and N stages

• Prognostic marker validation: KCND2 serves as an independent predictor of prognosis, with elevated levels in TCGA database revealing a more unfavorable prognosis for gastric cancer patients

• Functional pathway analysis: Antibodies can be used in co-immunoprecipitation experiments to demonstrate that KCND2 stimulates NF-κB pathways in both cellular and animal models, thereby enhancing cancer cell proliferation

• Immune interaction studies: Immunofluorescence and flow cytometry can reveal how KCND2 regulates the immune microenvironment by promoting M2 macrophage infiltration through NF-κB activation

For studying the HER2 status-independent predictive value of KCND2 as a biomarker, researchers can apply KCND2 antibodies in tissue microarrays to validate expression patterns across large patient cohorts .

How do post-translational modifications affect KCND2 function and how can antibodies detect these changes?

Post-translational modifications (PTMs) of KCND2 significantly impact channel function, and modification-specific antibodies serve as critical tools for investigating these regulatory mechanisms:

• Phosphorylation detection: Specific antibodies that recognize phosphorylated forms of Kv4.2 allow researchers to study activation-dependent changes in channel properties

• Glycosylation analysis: Antibodies targeting glycosylated epitopes help researchers understand how this modification affects channel trafficking and membrane insertion

• Ubiquitination studies: Antibodies that recognize ubiquitinated forms of Kv4.2 provide insights into protein degradation pathways that regulate channel turnover

• Interaction partner identification: Co-immunoprecipitation with KCND2 antibodies followed by mass spectrometry can reveal interaction partners that mediate or are affected by PTMs

When using PTM-specific antibodies, researchers should include appropriate controls to validate specificity, including dephosphorylation treatments, mutation of key residues, or pharmacological inhibition of relevant modifying enzymes.

What are the optimal protocols for KCND2 antibody validation in neuronal tissues?

Rigorous validation of KCND2 antibodies is essential for reliable research outcomes. For neuronal tissues, consider this comprehensive validation protocol:

• Knockout/knockdown verification: Test antibodies on tissues from KCND2-knockout animals or neurons treated with KCND2-specific siRNA to confirm specificity

• Cross-reactivity assessment: Evaluate potential cross-reactivity with closely related channels (particularly Kv4.3, which can form heterotetrameric channels with Kv4.2)

• Multi-antibody confirmation: Use multiple antibodies targeting different epitopes of Kv4.2 and confirm consistent staining patterns

• Species-specific validation: Despite high conservation across species, validate antibody performance specifically in your model system (human, mouse, rat)

• Signal-to-noise optimization: For immunofluorescence applications (typical dilution 1:100), optimize blocking protocols to minimize background staining while preserving specific signal

Proper validation significantly enhances data reliability, particularly for studies examining subtle changes in channel expression or localization in complex neuronal tissues.

How should researchers optimize western blotting protocols for KCND2 detection?

Western blotting for KCND2 requires careful optimization to obtain clear, specific bands at the expected molecular weight:

• Sample preparation: For membrane proteins like KCND2, use detergent-based lysis buffers (containing NP-40 or Triton X-100) that efficiently solubilize membrane proteins while preserving epitope integrity

• Expected molecular weight: While the predicted molecular weight of KCND2 is 71 kDa, post-translational modifications may result in higher observed weights. Expected MWt is typically around 42.3 kDa for the core protein

• Antibody dilution: Start with the manufacturer's recommended dilution (often 1:500) and optimize based on signal strength and background

• Positive controls: Include recombinant KCND2 protein (such as aa27-184) as a positive control to confirm antibody functionality

• Blocking optimization: For membrane proteins, optimization of blocking conditions (BSA vs. milk, blocking duration) is particularly important to reduce non-specific binding while maintaining specific signal

• Signal development optimization: Adjust exposure times to capture specific bands while minimizing background, using appropriate positive controls such as Kv4.2/KCND2 (aa27-184)-hIgGFc transfected HEK293 cell lysate

These optimizations ensure reliable detection of KCND2 in various sample types, from cell lines to complex tissue lysates.

What considerations are important for immunofluorescence applications with KCND2 antibodies?

Successful immunofluorescence staining for KCND2 requires attention to several technical aspects:

• Fixation protocol optimization: For membrane proteins like KCND2, mild fixation (2-4% paraformaldehyde for 10-15 minutes) often preserves antigenicity better than stronger fixatives or longer fixation times

• Permeabilization adjustment: Fine-tune permeabilization conditions to allow antibody access to intracellular epitopes while maintaining membrane structure integrity

• Antibody concentration: Start with validated dilutions (typically 1:100 for immunofluorescence) and adjust based on signal-to-noise ratio

• Incubation conditions: For low-abundance proteins, consider overnight primary antibody incubation at 4°C to maximize binding while minimizing non-specific interactions

• Appropriate controls: Include both positive controls (tissues known to express KCND2) and negative controls (primary antibody omission and ideally KCND2-knockout tissues)

• Multi-channel imaging: When colocalizing KCND2 with other markers, carefully select fluorophore combinations to minimize spectral overlap and ensure proper compensation during image acquisition

These considerations enable precise visualization of KCND2 localization patterns in neurons and other cell types, facilitating studies of trafficking, polarized distribution, and colocalization with interaction partners.

How can KCND2 antibodies be used to study neurodevelopmental disorders?

KCND2 variants are associated with global developmental delay and epileptic encephalopathy. Antibody-based approaches offer valuable insights into disease mechanisms:

• Expression pattern analysis: Compare KCND2 expression and localization patterns between normal and pathological tissues using immunohistochemistry or immunofluorescence

• Mutation-specific characterization: For identified variants like p.(Glu323Lys), p.(Pro403Ala), p.(Val404Leu), and p.(Val404Met), use antibodies to assess whether mutations affect protein stability, localization, or interaction patterns

• Animal model validation: Apply antibodies to validate KCND2 expression in animal models of neurodevelopmental disorders, confirming that the model recapitulates key aspects of human pathology

• Therapeutic response monitoring: Use antibodies to track changes in KCND2 expression or localization in response to experimental therapeutics targeting channel function or expression

For studies involving the p.(Val404Met) variant associated with severe seizures and autism in monozygotic twins, antibodies provide critical tools for understanding how genetic variants translate to protein-level alterations with functional consequences .

What is the role of KCND2 in gastric cancer and how should researchers approach its study?

Recent discoveries have established KCND2 as a significant player in gastric cancer progression. A comprehensive research approach includes:

• Expression correlation analysis: Use KCND2 antibodies for immunohistochemistry on tissue microarrays to correlate expression with clinical parameters including tumor grade, T stage, and N stage

• Prognostic marker validation: Immunohistochemical staining with KCND2 antibodies can validate its potential as a prognostic marker, particularly in well-differentiated, intestinal type, and Stage III patients, independent of HER2 status

• Functional pathway investigation: Combine KCND2 antibodies with those targeting NF-κB pathway components to elucidate the mechanistic link between KCND2 overexpression and enhanced cancer cell proliferation

• Immune microenvironment characterization: Multiplex immunofluorescence with KCND2 and macrophage markers can reveal how KCND2 influences M2 macrophage infiltration, potentially through NF-κB activation

• Therapeutic target evaluation: Use antibodies to track changes in KCND2 expression following experimental interventions targeting this channel as a potential therapeutic approach

These approaches support the emerging potential of KCND2 as both a biomarker and therapeutic target in gastric cancer, with KCND2 antibodies serving as essential tools for translational research in this field .

How do heteromeric channel complexes involving KCND2 impact experimental design and data interpretation?

KCND2 (Kv4.2) can form functional heterotetrameric channels with Kv4.3, creating complexity for researchers studying these channels . This heterogeneity necessitates specialized experimental approaches:

• Subunit-specific detection: Use antibodies with confirmed specificity for KCND2 over related channel subunits to accurately attribute observed signals to KCND2 rather than heteromeric complexes

• Co-immunoprecipitation studies: Apply KCND2 antibodies in co-IP experiments followed by detection of potential partner subunits to identify heteromeric complexes in specific tissues or under different conditions

• Proximity ligation assays: Combine antibodies against KCND2 and potential partner subunits in proximity ligation assays to visualize and quantify specific heteromeric interactions with spatial resolution

• Subunit manipulation controls: When manipulating KCND2 expression (through knockdown or overexpression), monitor effects on partner subunits, as changes in KCND2 may indirectly affect channel function through altered heteromeric complex formation

• Electrophysiological correlation: Correlate immunostaining results with electrophysiological recordings to understand the functional implications of detected heteromeric complexes

This comprehensive approach enables researchers to distinguish the specific contributions of KCND2 from those of heteromeric channel complexes, providing more accurate insights into channel function in both normal physiology and disease states.