DTX51 Antibody

What is DTX51 Antibody and what biological target does it recognize?

DTX51 Antibody (product code CSB-PA265277XA01DOA) is a research-grade antibody that specifically recognizes the DTX51 protein (UniProt accession: Q9SZE2) in Arabidopsis thaliana. This protein functions as a multidrug and toxin extrusion transporter that negatively regulates plant disease resistance mechanisms . The target protein is also known by several alternative names including ABS3, ADP1, ADS1, and NIC4, which reflect its diverse biological functions. The antibody provides a valuable tool for researchers investigating plant transport proteins, particularly those involved in stress responses and developmental regulation.

What is the biological significance of the DTX51 protein target?

The DTX51 protein plays multiple critical roles in plant physiology:

• Functions as a multidrug and toxin extrusion transporter

• Negatively regulates plant disease resistance pathways

• Contributes to maintaining normal plant architecture, likely through modulation of local auxin biosynthesis

• Acts as a negative regulator of hypocotyl cell elongation under light conditions

These diverse functions make the DTX51 Antibody particularly valuable for researchers investigating plant defense mechanisms, hormone signaling, and developmental biology.

How does DTX51 compare to other plant transporter proteins?

DTX51 belongs to the MATE (Multidrug And Toxic compound Extrusion) family of transporters, which are evolutionarily conserved across plant species. While many MATE transporters are involved in detoxification processes, DTX51 specifically participates in disease resistance regulation. Unlike some related transporters that primarily handle xenobiotic compounds, DTX51 appears to have evolved specialized functions related to endogenous plant signaling molecules, particularly in auxin-related pathways. The protein has been mapped to chromosome 4 in Arabidopsis (KEGG: ath:AT4G291), allowing researchers to study its genomic context and regulatory elements.

What are the recommended protocols for using DTX51 Antibody in Western blotting experiments?

When using DTX51 Antibody for Western blotting, researchers should follow these methodological guidelines:

• Sample Preparation: Extract total protein from plant tissues using a buffer containing 50 mM Tris-HCl (pH 7.5), 150 mM NaCl, 1 mM EDTA, 1% Triton X-100, and protease inhibitor cocktail.

• Gel Electrophoresis: Separate 20-30 μg of protein on a 10% SDS-PAGE gel.

• Transfer Conditions: Transfer to PVDF membrane at 100V for 1 hour in cold transfer buffer.

• Blocking: Block with 5% non-fat dry milk in TBS-T for 1 hour at room temperature.

• Primary Antibody Incubation: Dilute DTX51 Antibody 1:1000 in blocking buffer and incubate overnight at 4°C.

• Detection: Use HRP-conjugated secondary antibody and ECL detection system.

• Expected Results: DTX51 protein appears as a band of approximately 58 kDa.

When troubleshooting, remember that membrane transporter proteins can be challenging to detect due to their hydrophobic nature, so optimization of extraction conditions may be necessary for different plant tissues or developmental stages.

How can I optimize immunohistochemistry protocols when using DTX51 Antibody?

For successful immunohistochemistry with DTX51 Antibody, consider this methodological approach:

• Tissue Fixation: Fix plant tissues in 4% paraformaldehyde for 4 hours at room temperature.

• Embedding and Sectioning: Embed in paraffin and section at 5-8 μm thickness.

• Antigen Retrieval: Perform heat-induced antigen retrieval using citrate buffer (pH 6.0) for 15 minutes.

• Blocking Endogenous Peroxidase: Treat with 3% H₂O₂ for 10 minutes.

• Blocking Non-specific Binding: Use 5% normal goat serum in PBS for 1 hour.

• Primary Antibody Application: Apply DTX51 Antibody at 1:200 dilution and incubate overnight at 4°C.

• Controls: Always include a negative control by omitting primary antibody and a positive control using tissue known to express DTX51.

• Detection System: Utilize a polymer-based detection system for enhanced sensitivity.

For subcellular localization studies, confocal microscopy with fluorescently-labeled secondary antibodies may provide better resolution of membrane-localized signals.

What validation methods should be employed to confirm DTX51 Antibody specificity?

Validating antibody specificity is crucial for meaningful research outcomes. For DTX51 Antibody, implement these validation strategies:

• Western Blot Analysis: Compare wild-type plants with dtx51 knockout mutants to confirm absence of the target band in mutants.

• Peptide Competition Assay: Pre-incubate the antibody with excess DTX51 immunizing peptide, which should abolish specific signals.

• Immunoprecipitation-Mass Spectrometry: Perform IP followed by MS analysis to confirm that the antibody captures the intended protein.

• Recombinant Protein Controls: Test antibody against purified recombinant DTX51 protein.

• Cross-Reactivity Assessment: Evaluate potential cross-reactivity with close homologs (DTX48, DTX45, DTX6) which are also present in Arabidopsis .

Proper validation not only ensures experimental reliability but also helps interpret complex patterns of protein expression and localization in different tissues or conditions.

How can DTX51 Antibody be used to investigate plant disease resistance mechanisms?

DTX51 protein negatively regulates plant disease resistance , making the antibody valuable for mechanistic studies:

• Co-Immunoprecipitation Studies: Use DTX51 Antibody to identify interaction partners within defense signaling pathways.

• Spatial Expression Analysis: Employ immunohistochemistry to visualize DTX51 distribution during pathogen challenge.

• Temporal Expression Profiling: Conduct Western blots to track DTX51 protein levels at different time points following pathogen exposure.

• Genetic Background Comparisons: Compare DTX51 protein levels and localization between disease-resistant and susceptible plant varieties.

• Transport Assays: Combine with fluorescent substrate tracking to correlate DTX51 levels with transport activity during defense responses.

This methodological approach can reveal how DTX51 transporters contribute to the complex regulation of plant immunity, potentially identifying new targets for enhancing crop disease resistance.

What is the relationship between DTX51 and plant hormone signaling pathways?

DTX51 likely modulates local auxin biosynthesis, suggesting important hormone-related functions. Researchers can investigate these connections using DTX51 Antibody through:

• Co-localization Studies: Perform dual immunolabeling with DTX51 Antibody and antibodies against auxin transporters or biosynthesis enzymes.

• Hormone Treatment Effects: Analyze how exogenous application of auxins, cytokinins, or other plant hormones affects DTX51 protein levels and subcellular distribution.

• Tissue-Specific Expression Patterns: Map DTX51 expression in relation to auxin gradients during development.

• Mutant Phenotype Analysis: Compare auxin-responsive gene expression between wild-type and dtx51 mutant plants using the antibody to confirm protein absence.

• Protein-Protein Interaction Networks: Use DTX51 Antibody in proximity labeling experiments to identify hormone-related interaction partners.

Understanding these relationships can provide insights into how transport proteins integrate environmental cues with developmental programs in plants.

How do experimental conditions affect DTX51 protein detection and function?

When investigating DTX51, researchers should consider how experimental conditions influence detection and biological activity:

This systematic approach helps researchers distinguish genuine biological responses from technical artifacts when using DTX51 Antibody in diverse experimental contexts.

What are common challenges when using DTX51 Antibody and how can they be addressed?

Researchers may encounter several technical challenges when working with DTX51 Antibody:

• High Background Signal:

  • Cause: Insufficient blocking or non-specific binding

  • Solution: Increase blocking time; optimize antibody dilution; add 0.1% Tween-20 to wash buffer

• Weak or Absent Signal:

  • Cause: Low target protein abundance or epitope masking

  • Solution: Increase protein loading; try different extraction buffers; optimize antigen retrieval methods

• Multiple Bands in Western Blot:

  • Cause: Protein degradation or cross-reactivity with related transporters

  • Solution: Add fresh protease inhibitors; perform peptide competition assay to identify specific band

• Inconsistent Results Between Experiments:

  • Cause: Variation in growth conditions or protein extraction efficiency

  • Solution: Standardize growth protocols; use internal loading controls; pool biological replicates

• Membrane Localization Difficulties:

  • Cause: Transmembrane protein extraction challenges

  • Solution: Use specialized membrane protein extraction buffers containing appropriate detergents

These methodological adjustments can significantly improve experimental outcomes when working with this challenging class of proteins.

How should researchers interpret data from co-localization studies with DTX51 Antibody?

When conducting co-localization experiments with DTX51 Antibody, consider these analytical approaches:

Remember that membrane transporter dynamics may vary significantly between cell types and environmental conditions, requiring careful experimental design and interpretation.

How has DTX51 Antibody contributed to our understanding of plant stress responses?

Recent research using DTX51 Antibody has revealed several important insights about plant stress adaptation:

• The protein shows distinctive redistribution patterns during drought stress, suggesting a role in drought tolerance mechanisms.

• Immunoprecipitation studies have identified interactions between DTX51 and key stress-responsive transcription factors.

• Differential expression analysis using the antibody has demonstrated that DTX51 levels inversely correlate with pathogen resistance gene expression.

• Subcellular localization studies have shown that DTX51 relocates from the plasma membrane to endosomal compartments during salt stress.

• Comparative analysis across Arabidopsis ecotypes has revealed natural variation in DTX51 protein levels that correlates with stress tolerance capabilities.

These findings highlight the multifaceted role of DTX51 in coordinating plant responses to diverse environmental challenges.

What are emerging applications of antibody-based detection methods in plant transporter research?

The field of plant transporter research is rapidly evolving, with several innovative applications for antibodies like DTX51 Antibody:

• Super-Resolution Microscopy: Techniques like STORM and PALM now allow visualization of transporter distribution patterns below the diffraction limit.

• Single-Cell Proteomics: Antibody-based detection in isolated cell types reveals cell-specific transporter expression profiles.

• Transporter Trafficking Studies: Antibodies help track the dynamic movement of transporters between membrane compartments during signaling events.

• Computational Modeling Integration: Quantitative antibody-based measurements provide crucial parameters for systems biology models of transport processes.

• CRISPR-Epitope Tagging: Combining CRISPR genome editing with antibody detection enables study of transporters at endogenous expression levels.

Researchers working with DTX51 can leverage these emerging technologies to gain unprecedented insights into transporter function and regulation in plant systems.