APA3 Antibody
Description
Definition and Target Specificity
The term "APA3 Antibody" may refer to antibodies targeting:
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Adaptor Protein Complex 3 (AP-3): A vesicle-coat protein involved in intracellular trafficking, particularly in lysosomal targeting and immune cell degranulation .
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Aspergillus Galactomannan Antigen: A fungal cell wall polysaccharide targeted by monoclonal antibodies like AP3 for diagnostic purposes .
AP-3 Complex Antibody (ab85075)
Aspergillus-Specific Antibody AP3
Diagnostic Use in Invasive Aspergillosis
AP3 demonstrates high specificity for Aspergillus galactomannan (GM), reducing cross-reactivity with bacterial polysaccharides .
| Parameter | AP3 Antibody Performance |
|---|---|
| Target Antigen | Aspergillus Gal f residues |
| Diagnostic Utility | Serum GM detection in IA patients |
| Cross-Reactivity | Minimal with bacterial antigens |
Autoimmune Disorders
AP3B2-reactive IgG is linked to autoimmune neurological phenotypes:
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Clinical Presentation: Gait ataxia, sensory neuropathy, and cerebellar dysfunction .
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Mechanism: Targets AP3B2, a synaptic vesicle coat protein, disrupting neuronal signaling .
Therapeutic Potential of Bispecific Antibodies
While not directly APA3, the bispecific antibody AP203 (targeting PD-L1 and CD137) highlights advancements in antibody engineering:
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Mechanism: Blocks PD-1/PD-L1 signaling while activating CD137 on T cells .
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Efficacy: Enhances antitumor activity in xenograft models by increasing CD8+ T-cell infiltration .
Antibody Development and Databases
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Rep-Seq Analysis: Platforms like RAPID integrate antibody repertoire data, aiding in APA3-like antibody characterization .
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Therapeutic Antibodies: Over 500 candidates are cataloged, though APA3 is not yet listed .
Challenges and Future Directions
Product Specs
Composition: 50% Glycerol, 0.01M PBS, pH 7.4
Target Background
Q&A
What is APA3 Antibody and what is its target antigen?
APA3 Antibody is a polyclonal antibody raised in rabbits against recombinant Arabidopsis thaliana APA3 protein. It targets the APA3 protein (UniProt No. Q9XEC4) found in Arabidopsis thaliana, a widely used model organism in plant molecular biology research . This antibody belongs to the IgG isotype and is produced through antigen affinity purification methods to ensure specificity. Unlike antibodies targeting human proteins like ASK3 (which is related to the MAP3K15 gene involved in protein phosphorylation) , APA3 Antibody is specifically designed for plant research applications.
What are the validated applications for APA3 Antibody?
Based on available validation data, APA3 Antibody has been confirmed effective for the following applications:
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ELISA (Enzyme-Linked Immunosorbent Assay): For quantitative detection of APA3 protein in solution
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Western Blot (WB): For identification of APA3 protein in denatured samples
When employing this antibody in research, it's essential to verify its performance for your specific experimental conditions. Validation protocols similar to those used for other research antibodies, such as the Aspergillus-specific mAb AP3, can be adapted for APA3 Antibody validation, including specificity testing through immunoprecipitation and binding assays .
What are the optimal storage conditions for maintaining APA3 Antibody activity?
To preserve antibody function and prevent degradation, store APA3 Antibody at -20°C or -80°C immediately upon receipt. The antibody is supplied in a liquid form containing 50% glycerol, 0.01M PBS (pH 7.4), and 0.03% Proclin 300 as a preservative . This formulation helps maintain stability during freeze-thaw cycles, but repeated freezing and thawing should still be avoided. For short-term storage (1-2 weeks), aliquoting the antibody and storing at 4°C is recommended to prevent unnecessary freeze-thaw cycles.
How should researchers validate APA3 Antibody specificity for experimental use?
Comprehensive validation of APA3 Antibody specificity should include:
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Positive and negative controls: Use of known APA3-expressing and non-expressing tissues
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Cross-reactivity assessment: Testing against related proteins or tissues from other species
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Knockout/knockdown validation: Ideally, testing against APA3 knockout Arabidopsis thaliana samples
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Multiple detection methods: Comparing results across different applications (WB, ELISA)
Modern antibody validation approaches, as described in recent computational modeling studies of antibody specificity, emphasize the identification of different binding modes associated with particular ligands . For plant-specific antibodies like APA3, validation is particularly important as the antibody repertoire for plant research is less extensive than for human targets.
What experimental considerations apply when using APA3 Antibody for Western blotting?
| Parameter | Recommended Condition | Notes |
|---|---|---|
| Sample preparation | Standard plant protein extraction buffers with protease inhibitors | Fresh tissue preferred |
| Protein amount | 20-40 μg total protein per lane | Optimize based on expression level |
| Blocking solution | 5% non-fat dry milk or 3-5% BSA in TBST | BSA may provide lower background |
| Primary antibody dilution | Start with 1:1000 to 1:2000 | Titrate for optimal signal-to-noise ratio |
| Incubation | Overnight at 4°C or 1-2 hours at room temperature | Longer incubation often improves sensitivity |
| Detection method | HRP-conjugated secondary antibody with ECL detection | Fluorescent secondaries also applicable |
These parameters should be optimized for each experimental system. When troubleshooting, consider that plant samples often contain compounds that can interfere with antibody binding, necessitating additional optimization steps compared to mammalian systems.
How can researchers address cross-reactivity concerns with APA3 Antibody?
Cross-reactivity testing is vital, especially when working with polyclonal antibodies like APA3 Antibody. Researchers should:
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Test antibody reactivity against proteins from related species to determine species specificity beyond the confirmed Arabidopsis thaliana reactivity
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Perform pre-adsorption tests using purified antigens to confirm binding specificity
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Include appropriate controls in all experiments to distinguish specific from non-specific binding
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Consider epitope mapping to identify the specific binding regions, similar to the approach used for the AP3 monoclonal antibody where glycoarray analysis revealed specific oligosaccharide binding patterns
Understanding that polyclonal antibodies recognize multiple epitopes on the target antigen is essential when interpreting potential cross-reactivity results.
What are common causes of inconsistent results when using APA3 Antibody?
Inconsistent results can stem from several factors:
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Antibody degradation: Improper storage or excessive freeze-thaw cycles
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Variable antigen accessibility: Differences in sample preparation affecting epitope exposure
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Post-translational modifications: Changes in the target protein affecting antibody recognition
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Buffer incompatibility: Components in extraction or assay buffers interfering with binding
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Sample degradation: Protein degradation due to inadequate protease inhibition
A systematic approach to troubleshooting involves changing only one variable at a time and documenting all experimental conditions. The principles used in Rep-Seq dataset analysis platforms for antibody characterization can be adapted to systematically analyze variables affecting APA3 Antibody performance .
How can APA3 Antibody performance be enhanced for low-abundance targets?
For detecting low-abundance APA3 protein:
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Sample enrichment: Use immunoprecipitation to concentrate the target before analysis
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Signal amplification: Employ tyramide signal amplification or other enhancement methods
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Optimized extraction: Develop tissue-specific extraction protocols to maximize target protein yield
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Reduced background: Increase washing stringency and optimize blocking conditions
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Enhanced detection systems: Use highly sensitive chemiluminescent or fluorescent detection systems
The sensitivity can be further improved by combining multiple detection methods, similar to approaches used in autoantibody research where multiple detection parameters are evaluated simultaneously .
How does APA3 Antibody compare to other plant-specific antibodies in research applications?
When comparing APA3 Antibody to other plant antibodies:
| Antibody Type | Target Specificity | Applications | Advantages | Limitations |
|---|---|---|---|---|
| APA3 Antibody (Polyclonal) | Arabidopsis thaliana APA3 protein | ELISA, WB | Wide epitope recognition, strong signal | Batch-to-batch variation, potential cross-reactivity |
| Plant-specific monoclonal antibodies | Single epitope on target protein | Multiple applications including IHC, IF | High specificity, reproducibility | Limited epitope recognition, potentially lower signal |
| Anti-model protein antibodies (e.g., GFP, HA) | Tagged recombinant proteins | Multiple applications | High specificity regardless of target | Requires genetic modification of target organism |
The choice between these options depends on specific research requirements, with APA3 Antibody being particularly valuable for studies focused on native APA3 protein in Arabidopsis thaliana.
What research questions about plant biology can be addressed using APA3 Antibody?
APA3 Antibody can facilitate research into:
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Protein expression patterns: Examining APA3 expression across different tissues and developmental stages
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Response to environmental stimuli: Investigating changes in APA3 levels under various stress conditions
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Protein-protein interactions: Identifying binding partners through co-immunoprecipitation
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Post-translational modifications: Analyzing how modifications affect APA3 function
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Subcellular localization: Determining where APA3 functions within plant cells
These applications contribute to the broader understanding of plant cellular processes, similar to how anti-CD3 antibodies have advanced our understanding of T-cell biology in immune research .
How might emerging antibody technologies improve APA3 detection and analysis?
Future advancements that could enhance APA3 Antibody applications include:
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Computational antibody design: Using approaches like those described for customized specificity profiles to create antibodies with enhanced specificity for APA3
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Single-domain antibodies: Developing smaller antibody formats with improved tissue penetration
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Multi-specific antibodies: Creating antibodies that recognize both APA3 and other proteins of interest
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Antibody engineering: Modifying existing antibodies to improve affinity or reduce background
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Integration with high-throughput platforms: Incorporating APA3 detection into broader -omics approaches
These innovations could parallel developments seen in therapeutic antibody research, where modifications to antibody structure have dramatically improved performance characteristics .
What methodological considerations apply when using APA3 Antibody in complex plant samples?
Working with complex plant samples requires additional considerations:
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Sample preparation optimization: Different tissues may require specific extraction protocols
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Removal of interfering compounds: Plants contain various secondary metabolites that can interfere with antibody binding
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Tissue-specific protocols: Modification of standard protocols based on tissue type (leaves, roots, reproductive structures)
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Developmental timing: Consideration of protein expression changes during plant development
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Environmental variables: Accounting for how growth conditions affect target protein expression
Researchers should develop systematic approaches to address these variables, similar to how researchers studying autoantibodies have developed standardized protocols for analyzing complex human samples .
