tsp-15 Antibody
Product Specs
Constituents: 50% Glycerol, 0.01M PBS, pH 7.4
Target Background
- TSP-15 is essential for maintaining epithelial cell integrity. PMID: 15454573
Q&A
Here’s a structured collection of FAQs tailored for academic researchers working with TSP-15 antibodies, organized by complexity and methodological focus. Data from the provided sources are synthesized into actionable insights.
How to validate TSP-15 antibody specificity in model organisms like C. elegans?
Methodological Answer:
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Knockout/RNAi Controls: Use tsp-15 loss-of-function mutants or RNA interference (RNAi) to confirm antibody specificity. Loss of signal in mutants (e.g., sv15 hypomorph) validates specificity .
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Western Blot: Compare band patterns between wild-type and mutant lysates. TSP-15 should show reduced/no expression in mutants .
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Epitope Mapping: Align immunogen sequences (e.g., RVEDIIMEHSVTDGLLGPGAKPSVEAAGTGCCLCYPN ) with target species homologs to assess cross-reactivity.
Key Data:
| Validation Method | Expected Outcome (Wild-Type vs. Mutant) | Source |
|---|---|---|
| RNAi knockdown | ↓ Hypodermal membrane integrity | |
| Western blot | Absence of ~25 kDa band in mutants |
What criteria should guide TSP-15 antibody selection for protein interaction studies?
Methodological Answer:
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Epitope Localization: Prioritize antibodies targeting extracellular domains (e.g., residues 115–235 in human TSPAN15 ) for cell-surface interaction studies.
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Functional Validation: Use co-immunoprecipitation (Co-IP) assays to confirm interactions (e.g., TSP-15/BLI-3/DOXA-1 complexes in C. elegans ).
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Application Compatibility: Verify antibody performance in required assays (e.g., IF, ELISA ).
Antibody Comparison:
| Feature | PA5-139906 | CSB-PA025152LA01HU |
|---|---|---|
| Immunogen Region | RVEDIIMEHSVTDGLLGPGAK | 115–235AA |
| Applications Validated | WB, IF, IHC | ELISA, IF |
| Species Reactivity | Human | Human |
How to resolve contradictions in TSP-15’s dual roles in ROS generation and structural maintenance?
Methodological Answer:
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Context-Specific Assays:
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Genetic Rescue: Overexpress bli-3 and doxa-1 in tsp-15 mutants to dissect qualitative vs. quantitative roles .
Key Findings:
| Role | Experimental Evidence | Source |
|---|---|---|
| ROS generation | TSP-15 enables BLI-3 activation at the plasma membrane via rapid complex assembly | |
| Structural integrity | Loss of TSP-15 disrupts hypodermal barrier function |
How to optimize protocols for studying TSP-15/BLI-3/DOXA-1 interactions?
Methodological Answer:
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Co-IP Optimization:
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Cell Fusion Assays: Fuse TSP-15-expressing cells with BLI-3/DOXA-1-expressing cells (e.g., HVJ-mediated fusion ). Monitor H<sub>2</sub>O<sub>2</sub> production within 15 minutes post-fusion .
Critical Parameters:
| Parameter | Recommendation | Source |
|---|---|---|
| Protein expression | Stable TSP-15 + transient BLI-3/DOXA-1 | |
| Assay timing | Measure H<sub>2</sub>O<sub>2</sub> within 15–30 min post-fusion |
How to address cross-reactivity challenges in non-model organisms?
Methodological Answer:
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Phylogenetic Analysis: Compare TSP-15 sequences across species (e.g., human O95858 vs. C. elegans TSP-15 ).
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Functional Complementation: Express human TSPAN15 in C. elegans mutants to test cross-species activity .
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Epitope Mapping: Use antibodies targeting conserved regions (e.g., tetraspanin transmembrane domains ).
