DTX43 Antibody

What is DTX43 Antibody and what organism does it target?

DTX43 Antibody (product code CSB-PA874403XA01DOA) is a polyclonal antibody raised in rabbits against recombinant Arabidopsis thaliana DTX43 protein. It specifically recognizes the DTX43 protein from Arabidopsis thaliana (Mouse-ear cress), a model organism widely used in plant molecular biology research . This antibody is designed for research use only and is not intended for diagnostic or therapeutic applications.

What are the validated applications for DTX43 Antibody in plant research?

DTX43 Antibody has been validated for multiple applications in plant research:

• Enzyme-Linked Immunosorbent Assay (ELISA): For quantitative detection of DTX43 protein in plant extracts

• Western Blot (WB): For qualitative identification and semi-quantitative analysis of DTX43 protein in plant tissue extracts
  This versatility allows researchers to investigate protein expression patterns and compare protein levels across different experimental conditions.

How should DTX43 Antibody be stored to maintain optimal activity?

Proper storage is critical for maintaining antibody function:

• Store at -20°C or -80°C upon receipt

• Avoid repeated freeze-thaw cycles by aliquoting into smaller volumes

• The antibody is typically supplied in a storage buffer containing 50% glycerol, 0.01M PBS at pH 7.4, with 0.03% Proclin 300 as a preservative

• When stored properly, the antibody remains stable for approximately one year after shipment

How can DTX43 Antibody be used to investigate plant membrane transport mechanisms?

As DTX43 likely belongs to the MATE (Multidrug And Toxic compound Extrusion) family, the antibody can be used to study transport mechanisms through:

• Subcellular localization studies: Combine immunofluorescence with markers for different membrane compartments to determine precise localization

• Expression analysis under stress conditions: Use Western blot to monitor protein levels under various abiotic stresses, nutrient deficiencies, or toxin exposures

• Co-immunoprecipitation: Employ the antibody to identify protein interaction partners involved in transport complexes

• Comparative analysis across genotypes: Compare expression in wildtype vs. mutant plants to correlate protein levels with transport phenotypes

What controls are essential when using DTX43 Antibody for Western blot analysis?

Rigorous controls ensure reliable and interpretable results:

What methodological approaches can resolve discrepancies between DTX43 protein levels and gene expression data?

When protein and transcript levels don't correlate, consider:

• Technical verification:

  • Confirm antibody specificity using knockout lines

  • Verify primer specificity for transcript detection

  • Examine whether detection methods are within linear range

• Post-transcriptional regulation analysis:

  • Investigate microRNA regulation of DTX43 mRNA

  • Assess transcript stability using actinomycin D treatment

  • Examine alternative splicing that might affect epitope presence

• Protein stability assessment:

  • Test protein half-life using cycloheximide chase experiments

  • Investigate ubiquitination status using proteasome inhibitors

  • Examine post-translational modifications that might affect antibody recognition

• Temporal resolution studies:

  • Implement time-course experiments to detect delays between transcription and translation

  • Sample at multiple timepoints after stimulus application

What is the optimal protocol for using DTX43 Antibody in Western blot analysis of plant tissues?

For maximum sensitivity and specificity:
Sample preparation:

• Grind plant tissue in liquid nitrogen to a fine powder

• Extract proteins in buffer containing appropriate detergents (1% Triton X-100 or 0.5% SDS) to solubilize membrane proteins

• Include protease inhibitors to prevent degradation

• Clear lysates by centrifugation (14,000 × g, 15 min, 4°C)
  SDS-PAGE and transfer:

• Load 20-50 μg total protein per lane

• Separate on 10-12% SDS-PAGE gel

• Transfer to PVDF membrane (optimal for hydrophobic proteins) at 100V for 1 hour or 30V overnight
  Immunoblotting:

• Block with 5% non-fat dry milk in TBST for 1 hour

• Incubate with DTX43 Antibody (1:1000 dilution) overnight at 4°C

• Wash 3 × 10 minutes with TBST

• Incubate with HRP-conjugated anti-rabbit secondary antibody (1:5000) for 1 hour

• Develop using ECL substrate and image

How can DTX43 Antibody specificity be rigorously validated in plant systems?

A comprehensive validation approach includes:

• Genetic validation: Test antibody reactivity in:

  • Wild-type plants (positive signal expected)

  • DTX43 knockout/knockdown lines (absent/reduced signal)

  • DTX43 overexpression lines (enhanced signal)

  • Complementation lines (restored signal)

• Biochemical validation:

  • Peptide competition assays

  • Pre-adsorption with plant extracts lacking DTX43

  • Immunoprecipitation followed by mass spectrometry identification

• Cross-reactivity assessment:

  • Test against recombinant related proteins from the same family

  • Examine signal in species with varying sequence homology

What approaches help troubleshoot non-specific binding when using DTX43 Antibody?

When encountering background or non-specific signals:

• Sample preparation optimization:

  • Test different extraction buffers

  • Add higher concentrations of reducing agents

  • Include additional washing steps during protein extraction

• Blocking optimization:

  • Compare different blocking agents (milk, BSA, commercial blockers)

  • Increase blocking time (2-3 hours or overnight)

  • Add 0.1-0.3% Tween-20 to reduce hydrophobic interactions

• Antibody conditions:

  • Increase antibody dilution (test 2-5× more dilute)

  • Prepare antibody in fresh blocking solution

  • Pre-adsorb with non-specific proteins

• Washing stringency:

  • Increase number of washes (5-6 times instead of 3)

  • Extend washing time (15-20 minutes per wash)

  • Add higher concentrations of detergent (0.1-0.5% Tween-20)

How should quantitative data from DTX43 Antibody Western blots be normalized for meaningful comparisons?

Proper normalization ensures reliable quantitative analysis:

• Loading control normalization:

  • Express results as DTX43/loading control ratio

  • Verify loading control stability under experimental conditions

  • Use multiple loading controls when possible

• Multi-blot normalization:

  • Include common reference sample across all blots

  • Calculate relative expression compared to this reference

  • Consider using fluorescent detection for wider linear range

• Statistical considerations:

  • Ensure analysis is performed within linear detection range

  • Include at least three biological replicates

  • Verify normal distribution before applying parametric statistics

  • Report both fold-change and statistical significance

• Comparative visualization:

  • Present normalized data with appropriate error bars

  • Include representative blot images

  • Show individual data points when possible

What statistical approaches are appropriate for analyzing DTX43 protein expression across different experimental conditions?

The statistical approach should match your experimental design:

• For comparing multiple treatments:

  • One-way ANOVA followed by post-hoc tests (Tukey's HSD, Bonferroni)

  • Non-parametric alternatives (Kruskal-Wallis) if data don't meet normality assumptions

• For comparing treatment × tissue interactions:

  • Two-way ANOVA with interaction term

  • Mixed models if using repeated measures design

  • Planned contrasts for specific hypotheses

• For time-course experiments:

  • Repeated measures ANOVA

  • Area under curve (AUC) analysis

  • Time-to-peak or peak amplitude comparisons

• For correlation with physiological parameters:

  • Pearson or Spearman correlation coefficients

  • Multiple regression for complex relationships

  • Principal component analysis for multivariate data reduction

How can DTX43 protein expression levels be correlated with phenotypic data in Arabidopsis studies?

To establish meaningful correlations between protein levels and phenotypes:

• Experimental design considerations:

  • Measure protein levels and phenotypes in the same experimental units

  • Include multiple independent transgenic or mutant lines

  • Implement time-course analyses to establish causality

• Quantitative phenotyping approaches:

  • Use standardized growth and stress assays

  • Implement high-throughput imaging when possible

  • Consider both morphological and physiological parameters

  • Measure at multiple developmental stages

• Statistical analysis methods:

  • Calculate correlation coefficients between protein levels and phenotypic traits

  • Use regression analysis to establish predictive relationships

  • Apply multivariate methods to identify patterns across multiple phenotypes

  • Consider non-linear relationships when appropriate

How can DTX43 Antibody be integrated with other techniques for comprehensive protein analysis?

A multi-technique approach provides deeper insights:

What methodological approaches can help validate findings from DTX43 Antibody experiments using orthogonal techniques?

• Genetic approaches:

  • CRISPR/Cas9 knockout or knockdown of DTX43

  • Overexpression or complementation lines

  • Promoter-reporter fusions to compare with protein distribution

• Imaging approaches:

  • Fluorescent protein tagging (e.g., GFP-DTX43 fusion)

  • Live-cell imaging to complement fixed-tissue immunolocalization

  • FRET/FLIM to investigate protein-protein interactions

• Functional approaches:

  • Transport assays with isolated membrane vesicles

  • Electrophysiological measurements

  • Metabolite profiling to identify substrate accumulation/depletion

• Computational approaches:

  • Structural modeling of DTX43 to predict functional domains

  • Sequence analysis to identify conserved regions across species

  • Pathway analysis to place DTX43 in broader biological context This FAQ collection provides methodological guidance for researchers at all levels of expertise working with DTX43 Antibody, from basic applications to advanced experimental design considerations. The information presented here reflects current best practices in antibody-based research techniques in plant biology.