COR410 Antibody

What is COR410 protein and why is it significant in plant research?

COR410 (also known as Dehydrin COR410) is a cold-regulated dehydrin protein expressed in Triticum aestivum (wheat) that plays a critical role in plant stress responses, particularly to cold and drought conditions . As a member of the dehydrin family (LEA D-11 family), it functions as a stress-protective protein that helps maintain cellular integrity during environmental stress by preventing protein aggregation and membrane destabilization.

The protein has a UniProt accession number of P46524 and is particularly important in agriculture research focused on enhancing crop resilience to abiotic stressors . Study of COR410 contributes to understanding mechanisms of plant stress tolerance and potential genetic targets for improving crop durability in changing climate conditions.

What are the validated applications for COR410 antibody?

Based on technical validation data, COR410 antibody (product code CSB-PA342413ZA01TQN) has been validated for the following applications:

The antibody has been specifically tested against plant samples, with confirmed reactivity to Triticum aestivum targets . Unlike many antibodies in the research market that claim applications without rigorous validation, the COR410 antibody has undergone application-specific testing to ensure performance reliability in these methods .

What are the storage and handling recommendations for optimal antibody performance?

For maximum stability and performance of COR410 antibody:

• Store at -20°C or -80°C upon receipt

• Avoid repeated freeze-thaw cycles which can compromise antibody binding capacity

• The antibody is supplied in a storage buffer containing preservative (0.03% Proclin 300) and constituents (50% Glycerol, 0.01M PBS, pH 7.4)

• For long-term storage, aliquoting the antibody into smaller volumes is recommended to minimize freeze-thaw cycles

Research shows that proper antibody storage significantly impacts experimental reproducibility. In one study evaluating antibody degradation, samples subjected to multiple freeze-thaw cycles showed up to 30% reduction in binding efficacy compared to properly aliquoted samples .

What is the recommended protocol for using COR410 antibody in Western blotting?

Based on standard protocols for plant antibodies with similar characteristics:

• Sample preparation:

  • Extract total protein from plant tissue using a buffer containing 50 mM Tris-HCl (pH 7.5), 150 mM NaCl, 1% Triton X-100, and protease inhibitor cocktail

  • For drought or cold-stressed samples, immediate flash freezing in liquid nitrogen before extraction is critical to preserve protein modification states

• Western blot parameters:

  • Recommended dilution: Start with 1:1000 and optimize based on signal strength

  • Primary antibody incubation: Overnight at 4°C or 2 hours at room temperature

  • Secondary antibody: Anti-rabbit IgG conjugated with HRP (1:5000 dilution)

  • Blocking solution: 5% non-fat dry milk or 3% BSA in TBST

• Controls:

  • Positive control: Recombinant Triticum aestivum COR410 protein

  • Negative control: Pre-immune serum (supplied with antibody)

  • Loading control: Anti-actin or anti-tubulin antibody for normalization

The purification method (antigen affinity) ensures higher specificity compared to protein A/G purified antibodies, resulting in cleaner Western blot results with minimal non-specific bands .

How should researchers optimize ELISA protocols for COR410 detection?

For optimal ELISA performance with COR410 antibody:

• Plate coating:

  • For direct ELISA: Coat wells with plant extract (5-10 μg/ml) in carbonate buffer (pH 9.6)

  • For sandwich ELISA: Use a capture antibody against a different epitope of COR410

• Antibody application:

  • Starting dilution: 1:2000 in blocking buffer

  • Incubation: 1-2 hours at room temperature or overnight at 4°C

  • Detection: HRP-conjugated anti-rabbit secondary antibody (1:5000)

• Optimization parameters:

  • Perform checkerboard titration to determine optimal antibody concentration

  • Test different blocking agents (1-3% BSA, 5% non-fat milk)

  • Compare overnight vs. short incubation times for sensitivity differences

• Quantification:

  • Use purified recombinant COR410 protein to generate standard curve

  • Analyze data using four-parameter logistic regression for most accurate results

Research indicates that optimized ELISA protocols with polyclonal antibodies like COR410 can achieve detection limits in the low ng/ml range when properly optimized .

How is the specificity of COR410 antibody verified?

The specificity of COR410 antibody has been verified through multiple complementary approaches, following best practices in antibody validation :

• Immunogen verification:

  • The antibody was raised against a purified recombinant Triticum aestivum COR410 protein

  • The immunogen sequence was carefully selected to minimize cross-reactivity with other dehydrin family members

• Pre-immune serum comparison:

  • The antibody kit includes pre-immune serum as a negative control to establish baseline reactivity

  • This control allows researchers to distinguish specific from non-specific binding

• Validation controls:

  • The manufacturer provides positive control antigen (200μg) for validation experiments

  • This allows researchers to confirm antibody performance in their specific experimental conditions

While genetic knockout validation (considered the gold standard per IWGAV recommendations ) has not been specifically documented for this antibody, the orthogonal validation methods employed provide substantial evidence for specificity within its intended applications.

What approaches should researchers use to validate COR410 antibody in their own experimental systems?

Researchers should implement a systematic validation strategy:

• Positive and negative controls:

  • Use the supplied recombinant COR410 protein as positive control

  • Include samples from non-stressed plants (low COR410 expression) as biological negative controls

  • The pre-immune serum serves as technical negative control

• Expression correlation:

  • Verify that COR410 protein levels (detected by the antibody) correlate with mRNA expression (measured by qRT-PCR)

  • This orthogonal validation approach confirms target detection

• Stress induction:

  • Compare COR410 detection in normal vs. cold/drought-stressed plants

  • COR410 is significantly upregulated under stress conditions, so signal should increase accordingly

• Competition assay:

  • Pre-incubate antibody with excess recombinant COR410 protein

  • This should abolish specific binding in subsequent applications

  • Any remaining signal indicates non-specific binding

The scientific literature emphasizes that using multiple validation strategies significantly enhances confidence in antibody specificity, with at least two orthogonal methods recommended for robust validation .

How can COR410 antibody be used to investigate protein-protein interactions involving dehydrins?

Investigating protein-protein interactions with COR410 antibody requires specialized approaches:

• Co-immunoprecipitation (Co-IP):

  • Lyse plant tissue in non-denaturing buffer (50 mM Tris-HCl pH 7.5, 150 mM NaCl, 0.5% NP-40, protease inhibitors)

  • Pre-clear lysate with Protein A/G beads

  • Incubate with COR410 antibody (5-10 μg per 1 mg protein lysate)

  • Capture with fresh Protein A/G beads, wash extensively

  • Elute and analyze interacting proteins by mass spectrometry

• Proximity ligation assay (PLA):

  • Fix plant tissue sections with 4% paraformaldehyde

  • Incubate with COR410 antibody and antibody against suspected interaction partner

  • Use species-specific PLA probes and detection reagents

  • Analyze interaction signals by fluorescence microscopy

• Pull-down verification:

  • Express recombinant COR410 with affinity tag (His, GST)

  • Perform pull-down with plant extract

  • Use COR410 antibody to verify pull-down efficiency

  • Identify interaction partners by mass spectrometry

Research on similar dehydrin proteins has shown that these approaches can identify both structural interactions and transient stress-induced associations, providing insight into dehydrin function during stress responses .

What considerations are important when using COR410 antibody for comparative studies across different wheat varieties or related species?

When conducting cross-variety or cross-species studies:

• Sequence homology assessment:

  • Perform sequence alignment of COR410 across target species/varieties

  • The antibody targets a specific epitope; significant sequence divergence may affect recognition

  • Consider generating a phylogenetic tree of dehydrin proteins to predict cross-reactivity

• Validation for each species:

  • Perform Western blot with samples from each species/variety

  • Look for differences in band intensity, molecular weight, or multiple bands

  • Consider titration experiments to determine optimal antibody concentration for each species

• Data normalization strategy:

  • Use multiple reference proteins as loading controls

  • Consider absolute quantification with recombinant protein standards

  • Document any species-specific differences in extraction efficiency

• Controls for specificity:

  • Include pre-absorbed antibody controls for each species

  • Consider using RNAi or CRISPR plants (if available) as negative controls

  • Correlate protein detection with mRNA levels for each species

Studies using antibodies across species have shown that while many antibodies maintain reactivity with orthologs showing >70% sequence identity, validation for each specific target organism is essential for reliable results .

What are common issues encountered with COR410 antibody and their solutions?

These troubleshooting approaches are based on extensive experience with plant antibodies and follow best practices established in the antibody validation literature .

How can researchers enhance detection sensitivity when working with low-abundance COR410 protein?

For enhanced sensitivity in detecting low-abundance COR410:

• Sample preparation optimization:

  • Use specialized extraction buffers containing 8M urea to maximize dehydrin solubilization

  • Consider subcellular fractionation to concentrate COR410 from relevant compartments

  • Implement protein precipitation (TCA/acetone) to concentrate samples before analysis

• Signal amplification techniques:

  • For Western blots: Use high-sensitivity ECL substrates with extended exposure times

  • For ELISA: Implement biotin-streptavidin amplification systems

  • Consider tyramide signal amplification for immunohistochemistry applications

• Antibody enhancement strategies:

  • Use concentrated antibody solutions (1:500 instead of standard 1:1000)

  • Extend primary antibody incubation time to overnight at 4°C

  • Consider using protein A/G-conjugated magnetic beads for immunoprecipitation enrichment before detection

• Instrument optimization:

  • Use highly sensitive digital imaging systems with cooling capabilities for Western blots

  • For ELISA, employ readers with enhanced sensitivity in the appropriate wavelength range

  • Consider fluorescently-labeled secondary antibodies for quantitative Western blot analysis

Studies on low-abundance plant proteins have demonstrated that combining these approaches can increase detection sensitivity by 5-10 fold compared to standard protocols .

How can COR410 antibody be used to study the dynamics of dehydrin accumulation during stress responses?

COR410 antibody enables several approaches to study stress-induced dehydrin dynamics:

• Time-course experiments:

  • Subject plants to cold/drought stress for varying durations (0, 3, 6, 12, 24, 48 hours)

  • Extract proteins and quantify COR410 levels by Western blot or ELISA

  • Correlate protein accumulation with physiological stress markers

• Tissue-specific expression analysis:

  • Perform protein extraction from different tissues (roots, leaves, stems)

  • Compare COR410 accumulation patterns across tissues during stress

  • Complement with immunohistochemistry to visualize cellular localization

• Recovery dynamics:

  • Monitor COR410 levels during stress application and subsequent recovery

  • Track protein degradation/turnover rates using cycloheximide treatment

  • Correlate protein persistence with stress recovery metrics

• Stress signaling integration:

  • Combine COR410 detection with analysis of upstream regulators (DREB/CBF transcription factors)

  • Use pharmacological inhibitors of stress signaling pathways to determine regulatory mechanisms

  • Implement genetic approaches (mutants/transgenics) to establish signaling relationships

Research on similar dehydrin proteins shows that such approaches can reveal distinct accumulation patterns that correlate with stress tolerance thresholds in different plant varieties .

What are advanced approaches for studying COR410 post-translational modifications using this antibody?

To investigate post-translational modifications (PTMs) of COR410:

• Phosphorylation analysis:

  • Treat protein extracts with lambda phosphatase before Western blotting

  • Compare migration patterns with and without phosphatase treatment

  • Use Phos-tag™ acrylamide gels to enhance separation of phosphorylated forms

  • Consider generating phospho-specific antibodies for key regulatory sites

• 2D gel electrophoresis approach:

  • Separate proteins by isoelectric focusing followed by SDS-PAGE

  • Detect COR410 isoforms using the antibody

  • Identify spots with different pI values indicating PTMs

  • Excise spots for mass spectrometry analysis of modifications

• Mass spectrometry integration:

  • Immunoprecipitate COR410 using the antibody

  • Subject purified protein to tryptic digestion

  • Analyze peptides by LC-MS/MS for PTM identification

  • Quantify modification stoichiometry under different stress conditions

• Site-directed mutagenesis validation:

  • Express wild-type and mutated COR410 (at putative modification sites)

  • Compare antibody recognition patterns

  • Correlate modifications with functional properties (protein interactions, subcellular localization)

Studies on dehydrins have revealed that phosphorylation significantly affects their functional properties, including membrane binding and cryoprotective activities .