DCHS1 Antibody, HRP conjugated

What is DCHS1 and why are antibodies against it important for research?

DCHS1 (dachsous cadherin-related 1) is a large transmembrane protein belonging to the cadherin superfamily. In humans, the canonical protein has 3298 amino acid residues with a molecular mass of 346.2 kDa and localizes to the cell membrane . DCHS1 functions primarily in cell adhesion and is expressed in fibroblasts but not in melanocytes or keratinocytes .

Antibodies against DCHS1 are crucial research tools because:

• DCHS1 has been implicated in Van Maldergem syndrome and mitral valve prolapse

• DCHS1 forms a ligand-receptor pair with FAT4 during development

• The protein participates in a complex with LIX1L and SEPT9 (the DLS complex) during valve morphogenesis

Research with these antibodies enables visualization and quantification of DCHS1 in experimental systems, advancing our understanding of its role in development and disease.

What are the key characteristics of DCHS1 Antibody, HRP conjugated?

DCHS1 Antibody, HRP conjugated (e.g., CSB-PA850320OB01HU) has the following properties:

The antibody targets a specific region of the DCHS1 protein, providing selective detection capabilities when coupled with the enzymatic activity of HRP for signal amplification .

How does HRP conjugation affect antibody performance in DCHS1 detection?

HRP conjugation provides several methodological advantages for DCHS1 detection:

• Enhanced sensitivity: The enzymatic amplification by HRP enables detection of low abundance DCHS1 proteins

• Direct detection: Eliminates the need for secondary antibodies, reducing background and non-specific binding

• Versatile applications: Compatible with various substrates (TMB, DAB, luminol) for colorimetric, chemiluminescent, or fluorescent detection

• Stable signal generation: Produces consistent results when proper storage conditions are maintained

What applications are suitable for DCHS1 Antibody, HRP conjugated?

DCHS1 Antibody, HRP conjugated is primarily validated for ELISA applications , but depending on the specific product, may be suitable for:

• ELISA: Direct detection of DCHS1 in biological samples

• Immunohistochemistry: Some DCHS1 antibodies can be used for IHC applications

• Immunocytochemistry: Detection of DCHS1 in cultured cells

When considering application suitability, researchers should note that DCHS1 antibodies from different suppliers may have different application validations. For instance, while the HRP-conjugated version is validated for ELISA, other DCHS1 antibodies have been validated for Western blot (WB), immunofluorescence (IF), immunohistochemistry (IHC), and immunocytochemistry (ICC) .

What are the molecular interactions of DCHS1 and how can antibodies help study these interactions?

DCHS1 engages in several critical molecular interactions that can be studied using antibodies:

DCHS1-FAT4 Interaction:
DCHS1 forms a heterotypic interaction with FAT4 (FAT Atypical Cadherin 4) that affects planar cell polarity . This ligand-receptor pairing is essential during murine development and required in multiple organs .

DCHS1-LIX1L Interaction:
A yeast two-hybrid screen revealed that the cytoplasmic portion of DCHS1 (amino acids 2962-3191) interacts with the cytoplasmic protein LIX1L (specifically within amino acids 87-311 of LIX1L). The interaction domain was narrowed to amino acids 3130-3191 of DCHS1 .

DCHS1-LIX1L-SEPT9 (DLS) Complex:
DCHS1, LIX1L, and SEPT9 form a complex during valve morphogenesis. LIX1L is necessary for stabilizing this complex, as expression of DCHS1 in HEK293T cells is only detectable when co-expressed with LIX1L .

Antibodies can help study these interactions through:

• Co-immunoprecipitation assays

• Proximity ligation assays

• Immunofluorescence co-localization studies

• Protein complex isolation followed by mass spectrometry

Researchers should select antibodies targeting epitopes that don't interfere with the interaction domains of interest.

What are the optimal conditions for using DCHS1 Antibody, HRP conjugated in ELISA?

For optimal ELISA performance with DCHS1 Antibody, HRP conjugated:

Buffer Considerations:

• Avoid buffers containing sodium azide, as it inhibits HRP activity

• Use phosphate-buffered saline (PBS) at pH 7.4 for dilution

• Ensure buffers don't contain nucleophilic components, primary amines, or thiols

Protocol Optimization:

• Coating: Use 1-10 μg/ml of capture antigen or antibody in carbonate/bicarbonate buffer (pH 9.6)

• Blocking: 1-2% BSA or 5% non-fat dry milk in PBS for 1-2 hours at room temperature

• Antibody dilution: Start with manufacturer's recommended dilution (typically 1:500-1:2000) and optimize through titration

• Incubation: 1-2 hours at room temperature or overnight at 4°C

• Substrate: TMB is commonly used; develop for 5-30 minutes and stop with 2N H₂SO₄

• Controls: Include negative controls (no primary antibody) and positive controls (known DCHS1-positive samples)

Signal Enhancement Strategies:

• Extended substrate incubation (monitor to prevent overdevelopment)

• Optimized antibody concentration

• Enhanced blocking to reduce background

How can I validate the specificity of DCHS1 Antibody, HRP conjugated in my experimental system?

Comprehensive validation of DCHS1 Antibody, HRP conjugated should include:

Epitope Validation:

• Competitive inhibition with immunizing peptide (amino acids 2964-2981 of human DCHS1)

• Testing reactivity against recombinant DCHS1 protein

Cross-Reactivity Assessment:

• Testing in DCHS1 knockout/knockdown systems

• Comparing signals across species (noting that reactivity may vary)

• Testing in tissues known to express or lack DCHS1 (positive in fibroblasts, negative in melanocytes and keratinocytes)

Specificity Controls:

• Pre-absorption controls using the immunizing peptide

• Comparison with alternative DCHS1 antibodies targeting different epitopes

• Western blot to confirm detection at the expected molecular weight (~350 kDa)

Functional Validation:

• Immunoprecipitation followed by mass spectrometry

• Detection of expected DCHS1 interactions with FAT4 and LIX1L

What troubleshooting approaches are recommended for inconsistent results with DCHS1 Antibody, HRP conjugated?

When facing inconsistent results with DCHS1 Antibody, HRP conjugated:

High Background Issues:

• Increase blocking concentration (3-5% BSA or 5-10% non-fat dry milk)

• Optimize antibody dilution (try serial dilutions)

• Reduce incubation time or temperature

• Ensure thorough washing between steps (5-6 washes, 3-5 minutes each)

• Check for buffer contamination

Weak or No Signal:

• Verify DCHS1 expression in your samples (DCHS1 is primarily expressed in fibroblasts)

• Check HRP activity using a direct enzyme assay

• Increase antibody concentration

• Extend incubation times

• Ensure proper storage conditions were maintained (avoid repeated freeze-thaw cycles)

• Consider sample preparation methods that preserve epitope integrity

Inconsistent Results Between Experiments:

• Standardize protocols rigorously

• Use consistent lot numbers of antibody

• Prepare fresh working dilutions for each experiment

• Implement positive and negative controls in each experiment

• Account for DCHS1 complex formation requirements (co-expression of LIX1L may be necessary for stability)

How can DCHS1 Antibody, HRP conjugated be used to study mitral valve prolapse mechanisms?

DCHS1 has been identified as one of two known pathogenic genes associated with mitral valve prolapse (MVP) . To study this connection using DCHS1 Antibody, HRP conjugated:

Methodological Approaches:

• Comparative Expression Analysis:

  • Quantify DCHS1 protein levels in normal versus MVP patient samples

  • Correlate DCHS1 expression with disease severity

  • Compare expression patterns with clinical phenotypes

• Developmental Studies:

  • Track DCHS1 expression during valve morphogenesis

  • Examine co-localization of DCHS1, LIX1L, and SEPT9 in the developing valve

  • Study DCHS1 expression in valve endothelial cells and mesenchymal cells within the atrialis and valve tips

• Mutation Effect Assessment:

  • Compare wild-type versus mutant DCHS1 protein expression and localization

  • Evaluate the impact of mutations on DCHS1-FAT4 interactions

  • Determine if mutations affect the formation of the DCHS1-LIX1L-SEPT9 complex

• Tissue-Specific Analysis:

  • DCHS1 is expressed throughout the valve from embryonic day 15.5 to postnatal day zero

  • Expression is primarily restricted to endothelial cells and mesenchymal cells

  • Compare DCHS1 distribution in different regions of the valve

This targeted approach can provide insights into the molecular mechanisms by which DCHS1 mutations contribute to MVP, potentially revealing therapeutic targets.

What are the advantages and limitations of using polyclonal DCHS1 Antibody, HRP conjugated compared to monoclonal alternatives?

Advantages of Polyclonal DCHS1 Antibody, HRP conjugated:

• Multiple epitope recognition: Recognizes several epitopes on the DCHS1 protein, increasing detection sensitivity

• Robust to protein denaturation: More likely to recognize denatured proteins in applications like Western blotting

• Cost-effective production: Generally less expensive than monoclonal antibodies

• Rapid availability: Shorter production time compared to developing monoclonal antibodies

Limitations of Polyclonal DCHS1 Antibody, HRP conjugated:

• Batch-to-batch variation: Consistency between different lots may vary

• Cross-reactivity potential: May recognize similar epitopes on other proteins

• Less specificity for conformational studies: May not distinguish between different protein conformations

• Limited supply: Eventual exhaustion of the original antiserum

Comparative Analysis for Research Applications: