At2g45070 Antibody
Description
Molecular Target and Function
The At2g45070 gene encodes SEC61β1, part of the SEC61 translocon that facilitates co-translational protein translocation into the ER . Structural features:
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Protein structure: SEC61β1 is a tail-anchored (TA) membrane protein .
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Complex role: Partners with SEC61γ and SEC61α to form the core translocon .
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Functional homolog: Shares evolutionary conservation with yeast and mammalian SEC61β subunits .
Protein Translocation Studies
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Membrane fractionation: Used to quantify SEC61β1 in ER-enriched fractions, revealing reduced SYP123 (a SNARE protein) levels in Atget1 and Atget3a mutants .
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Interaction mapping: Identified via co-immunoprecipitation with SEC61γ (At5g50460) and other translocon components .
Genetic Knockout Analysis
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Phenotypic correlation: Arabidopsis lines lacking functional SEC61β1 show compromised root hair development, suggesting disrupted protein trafficking .
Validation and Specificity Concerns
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Cross-reactivity risks: Studies on analogous antibodies (e.g., angiotensin II AT2 receptor antibodies) highlight widespread nonspecific binding in Western blots and immunocytochemistry .
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Critical validation steps:
Key Research Findings
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GET pathway interaction: SEC61β1 abundance in membranes is reduced in Atget1 mutants, linking GET-mediated TA protein targeting to translocon stability .
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Transcriptional regulation: SEC61β1 mRNA levels decrease in stress conditions, suggesting feedback mechanisms .
Technical Considerations
Limitations and Alternatives
Product Specs
Constituents: 50% Glycerol, 0.01M Phosphate Buffered Saline (PBS), pH 7.4
Target Background
Q&A
Here’s a structured collection of FAQs tailored for researchers working with the At2g45070 antibody, synthesized from peer-reviewed studies and technical guidelines:
What experimental controls are essential when using the At2g45070 antibody for immunolocalization?
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Methodological answer: Always include:
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Knockout/knockdown controls: Validate specificity by comparing wild-type and At2g45070-deficient samples (e.g., CRISPR mutants) .
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Secondary antibody-only controls: Rule out nonspecific binding.
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Tissue/cell-type specificity controls: Test antibody reactivity in tissues with known At2g45070 expression (e.g., Arabidopsis endosperm) .
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How do I optimize Western blot conditions for At2g45070 antibody?
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Methodological answer:
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Gel electrophoresis: Use 10–12% SDS-PAGE to resolve the target protein (~50–60 kDa, depending on post-translational modifications) .
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Blocking: 5% nonfat milk in TBST for 1 hr to reduce background.
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Antibody dilution: Start with 1:1,000 dilution in blocking buffer; adjust based on signal-to-noise ratios .
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How to resolve discrepancies in At22g45070 antibody reactivity across assays?
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Analysis framework:
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Assay comparison: Compare results across platforms (e.g., ELISA vs. Western blot) to identify platform-specific biases .
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Temporal dynamics: Account for time-dependent antibody decay (e.g., pseudovirus neutralization assays show shorter half-lives than binding assays) .
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Quantitative normalization: Use internal reference proteins (e.g., actin) to standardize signal intensity .
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What computational tools improve quantification of At2g45070 antibody-based assays?
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Methodology:
How to address nonspecific binding of At2g45070 antibodies in heterologous systems?
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Validation strategy:
What factors influence longitudinal detection of At2g45070 antibodies?
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Critical variables:
Factor Impact on Detectability Mitigation Strategy Assay platform Roche N-assay vs. Ortho S-assay Use spike-targeting assays Disease severity Higher titers in severe infection Stratify cohorts by severity Time post-infection Antibody decay rate (~96–925 days) Optimize sampling intervals
How to reconcile conflicting immunolocalization and Western blot data?
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Troubleshooting workflow:
