At3g45770 Antibody
Product Specs
Constituents: 50% Glycerol, 0.01M PBS, pH 7.4
Target Background
Q&A
FAQs for Researchers on At3g45770 Antibody Applications in Academic Research
Advanced Research Questions
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How can structural insights improve At3g45770 antibody engineering for enhanced affinity?
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Use generative deep learning models (e.g., IgDesign) to redesign complementarity-determining regions (CDRs) while retaining framework stability. Validate designs via surface plasmon resonance (SPR) to measure binding kinetics (KD, kon/koff) .
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Conduct alanine scanning mutagenesis on CDR residues to identify critical binding hotspots. For example, a single HCDR3 mutation (e.g., T57H) improved affinity 2.6-fold in a GPC3-targeting antibody .
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What strategies resolve contradictions between in vitro binding data and in vivo functional efficacy?
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Comparative structural analysis: Resolve discrepancies by solving antibody-antigen complexes via X-ray crystallography. For example, antibodies targeting Aβ3–7 showed identical in vitro binding but divergent in vivo efficacy due to HCDR3 conformational differences .
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Functional assays: Test antibodies in transgenic plant models (e.g., Arabidopsis overexpressing At3g45770) to assess target engagement and phenotypic rescue .
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Methodological Comparisons
Table 1: Antibody Validation Techniques
| Method | Strengths | Limitations | Use Case |
|---|---|---|---|
| Western Blot | High specificity for linear epitopes | Limited to denatured proteins | Initial specificity screening |
| SPR | Real-time kinetic data (KD, kon/koff) | Requires purified antigen | Affinity optimization |
| IHC | Spatial resolution in tissues | Epitope masking risks | Localization studies |
Table 2: Mutagenesis Approaches for Affinity Maturation
Troubleshooting Data Conflicts
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Why do At3g45770 antibodies show variable performance across plant species?
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Analyze sequence divergence in epitope regions using multiple sequence alignment. For example, residues 12–18 of At3g45770 may differ in orthologs, requiring species-specific antibody validation .
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Test cross-reactivity using protein microarrays with homologs from related species (e.g., Brassica napus) .
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How to prioritize antibody clones for in vivo studies?
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Screen clones using ex vivo synaptic plasticity assays (e.g., hippocampal slice electrophysiology) to link target engagement to functional outcomes, as demonstrated for Aβ-targeting antibodies .
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For plant studies, use tissue-specific promoters (e.g., vascular-driven expression) to assess antibody penetration and biodistribution .
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