ADR2 Antibody
Description
Target Protein: Adr2 in Rickettsia Species
Adr2 is an outer membrane protein expressed by Rickettsia conorii (spotted fever group) and Rickettsia prowazekii (typhus group). It plays a critical role in:
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Serum resistance: Mediating evasion of complement-mediated killing .
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Vitronectin binding: Facilitating interaction with the host complement regulator vitronectin (Vn) to inhibit membrane attack complex formation .
Key structural features:
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Predicted extracellular loops (Loop 1 and Loop 2) are immunogenic and targeted by polyclonal antibodies .
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Highly conserved among pathogenic Rickettsiae (97.8% homology in SFG species; 88.1% in TG species) .
Serum Resistance Mechanism
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Expression in E. coli: Adr2-expressing E. coli showed 60–80% survival in human serum, compared to <20% for controls (P < 0.001) .
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Dependency on vitronectin: Adr2-bound Vn was confirmed via co-sedimentation assays and Western blot (75 kDa band detected with anti-Vn antibodies) .
Table 1: Functional Comparison of Adr2 Homologs
| Species | Homology to R. conorii Adr2 | Serum Resistance (Survival %) | Vn Binding Confirmed? |
|---|---|---|---|
| R. conorii (SFG) | 100% | 75–85% | Yes |
| R. prowazekii (TG) | 88.1% | 70–80% | Yes |
| Vector control | N/A | <20% | No |
Antibody Applications
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Flow cytometry: Demonstrated surface localization of Adr2 in paraformaldehyde-fixed R. conorii (shift in fluorescence intensity: ΔMFI = 1,200) .
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Western blot: Detected Adr2 at ~25 kDa in R. conorii and R. rickettsii lysates .
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Functional assays: Validated Adr2’s role in serum resistance using CFU-based survival assays .
Antibody Validation and Specificity
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Epitope recognition: Targets extracellular loops 1 and 2 of Adr2 .
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Cross-reactivity: Confirmed with R. prowazekii Adr2 homologs .
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Negative controls: No binding observed in E. coli with empty vectors or secondary antibody-only treatments .
Implications for Therapeutic Development
Adr2’s interaction with vitronectin highlights its potential as a:
Product Specs
Target Background
- ADR2-mediated resistance operates via an Enhanced Disease Susceptibility (EDS) and salicylic acid (SA)-dependent defense signaling network, functioning independently of COI1 or ETR1. [activated disease resistance 2] [ADR2] PMID: 19549129
- WRR4, a related protein, offers a potential novel source of white rust resistance in oilseed and vegetable brassica crops. PMID: 20447277
- WRR4 (*At1g56510*) encodes a cytoplasmic toll-interleukin receptor-like nucleotide-binding leucine-rich repeat receptor-like protein conferring dominant, broad-spectrum white rust resistance (WRR). PMID: 18624640
Q&A
Basic Research Questions
How do I validate ADR2 antibody specificity for Western blot applications?
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Perform parallel testing with knockout cell lines or siRNA-mediated target protein knockdown .
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Use peptide blocking controls (pre-incubate antibody with immunizing peptide) to confirm signal specificity .
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Validate across multiple species if applicable (human/mouse/rat reactivity confirmed in ADD2 studies) .
Experimental validation table
| Method | Protocol | Controls Required |
|---|---|---|
| Western Blot | 1:500-1:1000 dilution, 80kDa band | Knockout lysates, peptide block |
| Immunofluorescence | 1:100-1:500, nuclear/cytoplasmic localization | Isotype controls, secondary-only |
What experimental factors influence ADR2 antibody performance in ELISA?
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Optimize coating concentration (0.5-2 μg/ml range based on ADD2 protocols)
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Address matrix effects by testing serum/plasma spike-recovery experiments
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Validate linear detection range using recombinant protein standards
Advanced Research Challenges
How to resolve conflicting data between ADR2 detection methods?
Case example: Discrepant WB vs. IF results may indicate:
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Post-translational modifications affecting epitope accessibility
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Subcellular localization differences (membrane-bound vs. cytosolic forms)
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Solution: Combine mass spectrometry validation with antibody-based assays
What strategies mitigate cross-reactivity in complex biological samples?
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Implement negative selection chromatography to pre-clear sera
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Use tandem antibody approaches (e.g., IP-Western with distinct epitope tags)
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Employ computational epitope mapping against protein databases
Clinically significant antibody assessment framework
Mechanistic Research Considerations
How to evaluate ADR2's role in drug resistance pathways?
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Establish isogenic cell lines with modulated ADR2 expression
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Monitor real-time trafficking using pH-sensitive tags (e.g., pHrodo-ADR2)
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Combine transcriptomic profiling (RNA-seq) with protein interaction studies
Key findings from combination therapy studies
| Condition | ADR2 Modulation Effect | Citation |
|---|---|---|
| Hyperglycemia | ↑GLUT1 → Chemoresistance | PMC5108333 |
| Zinc supplementation | Restores drug sensitivity | PMC5108333 |
