At1g65352 Antibody

What is the At1g65352 Antibody and what gene product does it target?

At1g65352 Antibody (Product Code: CSB-PA651206XA01DOA) targets the protein product encoded by the At1g65352 gene in Arabidopsis thaliana, corresponding to UniProt accession number Q2V4F0. This antibody is designed specifically for detecting the target protein in experimental applications, providing researchers with a tool for studying gene expression and protein localization in this model plant organism .

What are the standard applications for At1g65352 Antibody in plant molecular biology?

The At1g65352 Antibody is primarily utilized in immunological detection methods including Western blotting, immunoprecipitation, immunohistochemistry, and ELISA when studying Arabidopsis thaliana. Each application provides different insights: Western blotting confirms protein expression and molecular weight, immunoprecipitation isolates protein complexes, immunohistochemistry reveals spatial localization within tissues, and ELISA enables quantitative protein measurement. These applications form the foundation of protein-level studies in plant molecular biology research.

What expression systems can be used to validate At1g65352 Antibody specificity?

Mammalian cell expression systems like Expi293 cells have been demonstrated as effective for antibody production and validation, as evidenced in related research protocols. For plant-specific antibodies like At1g65352, researchers typically validate specificity using Arabidopsis wild-type tissues compared to knockout mutants, or through heterologous expression in systems like E. coli, yeast, or insect cells. Validation involves demonstrating signal presence in samples containing the target protein and absence in negative controls .

How should researchers store and handle At1g65352 Antibody to maintain reactivity?

Proper storage and handling of At1g65352 Antibody is critical for maintaining its functional properties. The antibody is typically available in 0.1ml or 2ml sizes and should be stored at -20°C for long-term preservation . For routine use, aliquoting is recommended to avoid repeated freeze-thaw cycles that can compromise antibody function. When working with the antibody, maintaining cold chain conditions (4°C) during experimental procedures helps preserve binding capacity, and appropriate buffer conditions (typically PBS with stabilizers) should be used for dilution.

How should researchers design experiments to distinguish between specific and non-specific binding of At1g65352 Antibody?

Designing experiments to evaluate antibody specificity requires multiple controls. Researchers should include:

• Arabidopsis knockout/knockdown mutants lacking the At1g65352 gene product

• Pre-absorption controls where the antibody is pre-incubated with purified target protein

• Secondary antibody-only controls to assess background

• Cross-reactivity assessment using closely related Arabidopsis proteins

The inclusion of these controls helps differentiate genuine signal from artifacts, particularly when working with plant tissues that may contain complex profiles of related proteins or compounds that could interfere with antibody binding.

What are the optimal sample preparation methods for detecting At1g65352 protein in different plant tissues?

Optimal sample preparation varies by tissue type and developmental stage. For protein extraction from Arabidopsis tissues, researchers should consider:

Tissue-specific optimization of extraction conditions ensures maximum recovery of the target protein while minimizing degradation or modification that could affect antibody recognition.

How can At1g65352 Antibody be utilized in co-immunoprecipitation studies to identify protein interaction partners?

For co-immunoprecipitation (Co-IP) studies, At1g65352 Antibody should be conjugated to a solid support (typically Protein A/G beads) via direct chemical coupling or using secondary antibody bridges. The experimental workflow should include:

• Preparation of plant lysates under non-denaturing conditions

• Pre-clearing of lysates with blank beads to reduce non-specific binding

• Incubation with antibody-conjugated beads

• Stringent washing to remove non-specific interactions

• Elution of bound complexes for downstream analysis by mass spectrometry

This approach enables identification of proteins that physically interact with the At1g65352 gene product in vivo, providing insights into its biological function and regulatory networks in Arabidopsis.

What considerations should be made when using At1g65352 Antibody in chromatin immunoprecipitation (ChIP) experiments?

When adapting At1g65352 Antibody for ChIP applications, researchers should consider:

• Crosslinking optimization (typically 1-2% formaldehyde for 10-15 minutes)

• Sonication parameters to generate appropriate DNA fragment sizes (200-600 bp)

• Antibody specificity validation through epitope controls

• Input normalization and negative region controls

• Sequential ChIP approaches if investigating protein complexes

If the At1g65352 protein functions in transcriptional regulation or chromatin modification, ChIP experiments can reveal its genomic binding sites and contribute to understanding its role in gene expression control.

How does post-translational modification affect At1g65352 protein detection by antibodies?

Post-translational modifications (PTMs) can significantly impact antibody recognition of the At1g65352 protein. Common plant protein PTMs include phosphorylation, ubiquitination, SUMOylation, and glycosylation. Researchers should consider:

• Epitope mapping to determine if the antibody recognition site includes known or predicted modification sites

• Comparison of detection in samples treated with phosphatases or deglycosylation enzymes

• Use of modification-specific antibodies if studying particular PTM states

• Western blot analysis to identify potential mobility shifts associated with modifications

Understanding these factors helps interpret variation in detection efficiency across different experimental conditions or developmental stages.

What are common causes of false negative results when using At1g65352 Antibody, and how can they be addressed?

False negative results may arise from several factors:

Systematic troubleshooting using this framework can help identify and overcome technical challenges in At1g65352 protein detection.

How can researchers distinguish between contradictory data when comparing At1g65352 Antibody results across different experimental platforms?

When faced with contradictory results across platforms, researchers should:

• Evaluate antibody validation data for each experimental method

• Consider epitope accessibility differences between applications (native vs. denatured conditions)

• Assess potential cross-reactivity with related proteins

• Implement orthogonal detection methods (e.g., mass spectrometry, reporter gene fusion)

• Examine the literature for known technical challenges with the specific protein

What statistical approaches are most appropriate for analyzing immunological data generated with At1g65352 Antibody?

Statistical analysis of immunological data should be tailored to the specific experimental design. For quantitative Western blot or ELISA data, researchers should:

• Perform at least three biological replicates with technical replicates

• Apply appropriate normalization to loading controls or reference genes

• Test for normal distribution before selecting parametric or non-parametric tests

• Use paired tests when comparing treatments within the same biological sample

• Apply multiple testing correction for experiments analyzing multiple conditions

For spatial data (immunohistochemistry), quantification should include representative sampling across tissues and statistical comparison of signal intensity or pattern distribution.

How might emerging antibody engineering technologies improve At1g65352 Antibody performance?

Recent advancements in antibody engineering, like those described in the DyAb platform, could enhance At1g65352 Antibody performance. This sequence-based antibody design approach uses machine learning to predict and optimize properties like binding affinity . Applied to plant antibodies, these technologies could:

• Improve specificity through complementarity-determining region (CDR) optimization

• Enhance affinity through mutational scanning and combinatorial design

• Increase stability for challenging experimental conditions

• Develop cross-species reactivity for comparative studies across plant models

These engineered antibodies could provide more consistent results and expand the range of applicable techniques for At1g65352 research.

What are the considerations for developing multiplexed immunoassays including At1g65352 Antibody?

Developing multiplexed assays requires careful antibody selection and validation. Researchers should:

• Verify compatible fixation and antigen retrieval conditions across all targets

• Select primary antibodies from different host species to enable distinct detection

• Test for cross-reactivity between secondary detection systems

• Optimize signal amplification methods to balance detection of low and high abundance targets

• Validate spectral separation when using fluorescent detection systems

Multiplexed approaches allow simultaneous analysis of At1g65352 with interacting partners or pathway components, providing more comprehensive biological insights.