FAX7 Antibody

What is FAX7 Antibody and what organism does it target?

FAX7 Antibody (e.g., CSB-PA524640XA01DOA) is a rabbit polyclonal antibody that recognizes the Arabidopsis thaliana FAX7 protein. It is produced through immunization with recombinant Arabidopsis thaliana FAX7 protein and is purified using antigen affinity chromatography. The antibody is intended for research applications in plant biology, particularly for studying fatty acid export processes in Arabidopsis .

What are the validated applications for FAX7 Antibody?

Currently, FAX7 Antibody has been validated for:

• Enzyme-Linked Immunosorbent Assay (ELISA)

• Western Blotting (WB)

These applications enable researchers to detect and quantify FAX7 protein expression in experimental samples. Validation for additional applications such as immunohistochemistry, immunofluorescence, or flow cytometry would require further testing by individual laboratories .

What is the recommended storage condition for FAX7 Antibody?

FAX7 Antibody should be stored at -20°C or -80°C upon receipt. Repeated freeze-thaw cycles should be avoided to maintain antibody integrity and activity. The antibody is typically provided in a storage buffer containing 50% glycerol, 0.01M PBS (pH 7.4), and 0.03% Proclin 300 as a preservative .

How should I validate the specificity of FAX7 Antibody for my research?

To validate FAX7 Antibody specificity:

• Western Blot Validation: Compare signals from wild-type versus FAX7 knockout samples.

• Correlation Analysis: Calculate correlation factors between Western blot signals and array-based signals for the same set of samples (though this may have limitations for proteins with similar expression levels across samples) .

• Positive and Negative Controls: Include FAX7-overexpressing samples and known negative samples.

• Preabsorption Test: Pre-incubate the antibody with purified antigen before immunostaining to confirm specificity.

Similar to validation approaches used for other antibodies, these methods will help establish confidence in your experimental results .

What control samples should be included when using FAX7 Antibody in experiments?

For rigorous experimental design with FAX7 Antibody, include:

• Positive control: Samples known to express FAX7 protein (e.g., specific Arabidopsis thaliana tissues)

• Negative control: Samples known not to express FAX7 (e.g., FAX7 knockout lines)

• Technical controls:

  • No primary antibody control

  • Isotype control (though limitations apply; see FAQ 3.2)

  • For flow cytometry: Fluorescence Minus One (FMO) controls if performing multicolor analysis

• Loading controls: For Western blots, include housekeeping proteins to normalize expression data

These controls help distinguish specific from non-specific signals and enable accurate data interpretation .

How should I determine the optimal antibody concentration for my experiment?

Determining optimal antibody concentration requires careful titration:

• For Western Blot:

  • Test a range of dilutions (e.g., 1:500, 1:1000, 1:2000, 1:5000)

  • Select concentration that gives strongest specific signal with minimal background

  • Consider using gradient SDS-PAGE to improve separation of closely sized proteins

• For ELISA:

  • Prepare antibody dilution series (typically 1:100 to 1:10,000)

  • Plot signal-to-noise ratio against antibody concentration

  • Select concentration at inflection point of curve

Document optimization experiments systematically for reproducibility and protocol refinement .

How can I incorporate FAX7 Antibody into multicolor flow cytometry panels?

Although FAX7 Antibody has not been specifically validated for flow cytometry, principles from other antibody studies apply if developing this application:

• Fluorochrome selection based on antigen density:

  • High density antigens: Use lower brightness index fluorophores

  • Medium density antigens: Use moderate brightness fluorophores

  • Low density antigens: Use high brightness fluorophores (e.g., PE, AF647)

• Panel design considerations:

  • Minimize spectral overlap when possible

  • Use compensation beads for each fluorochrome

  • Implement Fluorescence Minus One (FMO) controls

• Validation protocol:

  • Compare staining between wild-type and knockout samples

  • Confirm specificity with blocking experiments

  • Evaluate sensitivity through titration experiments

This approach follows established methodologies for developing flow cytometry applications with new antibodies .

When should I use or avoid isotype controls with FAX7 Antibody?

The use of isotype controls requires careful consideration:

When to use:

• Initial validation of a new antibody lot

• When assessing potential Fc receptor-mediated binding

• When developing new applications or protocols

When to avoid:

• Once specificity has been established through proper controls (knockout/knockdown)

• When more informative controls are available (e.g., FMO in flow cytometry)

• When the isotype control lacks matching characteristics (concentration, F/P ratio, storage time)

The scientific community increasingly recognizes that knockout/knockdown controls provide more definitive evidence of specificity than isotype controls alone .

What advanced imaging techniques can be applied with FAX7 Antibody for subcellular localization studies?

For advanced subcellular localization using FAX7 Antibody:

• Super-resolution microscopy:

  • STED (Stimulated Emission Depletion) microscopy

  • PALM (Photoactivated Localization Microscopy)

  • STORM (Stochastic Optical Reconstruction Microscopy)

• Co-localization studies:

  • Use organelle-specific markers alongside FAX7 Antibody

  • Implement Pearson's correlation coefficient analysis

  • Consider proximity ligation assays for protein-protein interactions

• Live cell imaging adaptations:

  • Develop Fab fragments for reduced interference

  • Consider single-chain variable fragments (scFv) approaches

These techniques require additional validation but can provide valuable insights into protein function and localization .

What are common causes of high background when using FAX7 Antibody in immunoassays?

High background with FAX7 Antibody may result from:

• Antibody-related factors:

  • Excessive antibody concentration (solution: optimize through titration)

  • Non-specific binding (solution: increase blocking time/concentration)

  • Cross-reactivity with similar epitopes (solution: pre-absorb with related proteins)

• Sample preparation issues:

  • Incomplete blocking (solution: test alternative blocking agents)

  • Over-fixation affecting epitope accessibility (solution: optimize fixation protocol)

  • Endogenous peroxidase activity (solution: add quenching step)

• Detection system problems:

  • Excessive incubation with detection reagents (solution: optimize incubation time)

  • Light exposure of light-sensitive detection reagents (solution: protect from light)

Systematic evaluation of each factor can help identify and address the specific source of background .

How can I troubleshoot weak or absent signals when using FAX7 Antibody?

When experiencing weak or absent signals:

• Epitope accessibility issues:

  • Test different antigen retrieval methods

  • Try alternative fixation protocols

  • Consider native vs. denaturing conditions

• Technical considerations:

  Problem	Possible Solution
  Protein degradation	Add protease inhibitors
  Low transfer efficiency	Optimize transfer parameters
  Insufficient incubation	Increase incubation time
  Low antibody concentration	Increase antibody concentration
  Detection sensitivity	Switch to more sensitive detection system

• Biological factors:

  • Verify expression timing/conditions

  • Consider post-translational modifications affecting epitope

  • Evaluate target protein half-life

A methodical approach to troubleshooting will help identify the specific limitation in your experimental system .

How can I adapt protocols for FAX7 Antibody use in specialized applications like chromatin immunoprecipitation (ChIP)?

Adapting FAX7 Antibody for specialized applications requires careful optimization:

• For ChIP applications:

  • Test different crosslinking conditions (formaldehyde concentration, time)

  • Optimize sonication/fragmentation parameters

  • Determine optimal antibody concentration through titration

  • Validate enrichment with known targets

  • Include appropriate negative controls (IgG, non-target regions)

• For co-immunoprecipitation (Co-IP):

  • Test various lysis conditions to maintain protein interactions

  • Optimize antibody-to-bead ratios

  • Consider crosslinking antibody to beads to prevent contamination

  • Include appropriate controls (isotype antibody, non-expressing cells)

Each adaptation requires validation specific to the application and experimental system .

What statistical methods are appropriate for analyzing FAX7 Antibody-derived quantitative data?

For robust statistical analysis of FAX7 Antibody data:

• Western blot quantification:

  • Normalize target band to loading control

  • Use triplicate biological replicates minimum

  • Apply appropriate statistical tests (t-test, ANOVA, etc.)

  • Consider non-parametric alternatives if normality assumptions aren't met

• ELISA data analysis:

  • Generate standard curves using appropriate regression methods

  • Calculate coefficient of variation between replicates (aim for <10%)

  • Determine assay detection limits (LOD, LLOQ, ULOQ)

  • Apply dilution linearity tests to confirm accuracy

• Image-based analysis:

  • Use consistent thresholding methods

  • Implement blind analysis when possible

  • Consider machine learning approaches for complex pattern recognition

Each analytical approach should be validated and documented thoroughly for reproducibility .

How should I validate FAX7 Antibody results with complementary techniques?

Robust research requires orthogonal validation:

• Complementary protein detection methods:

  • Mass spectrometry-based proteomics

  • Proximity ligation assays

  • In situ hybridization (for mRNA correlation)

• Genetic validation approaches:

  • CRISPR/Cas9 knockout controls

  • RNAi knockdown experiments

  • Overexpression systems

• Functional validation methods:

  • Phenotypic analyses in FAX7-deficient systems

  • Rescue experiments with wild-type protein

  • Structure-function relationship studies

What considerations should be made when comparing data between different lots of FAX7 Antibody?

Inter-lot comparisons require careful consideration:

• Lot-to-lot validation protocol:

  • Test both antibody lots side-by-side on identical samples

  • Calculate correlation coefficients between results

  • Determine adjustment factors if necessary

• Documentation practices:

  • Record lot numbers in all experimental notes

  • Document any observed differences in specificity/sensitivity

  • Include lot information in publications/reports

• Experimental design adaptations:

  • When possible, complete experimental series with single lot

  • Include internal reference samples across experiments

  • Consider critical experiments that would require revalidation with new lot

Proactive approach to lot-to-lot variation helps maintain data consistency and integrity over time .

How can FAX7 Antibody be used to study protein-protein interactions in plant systems?

For protein interaction studies with FAX7 Antibody:

• Co-immunoprecipitation (Co-IP):

  • Optimize gentle lysis conditions to maintain protein complexes

  • Use chemical crosslinking to stabilize transient interactions

  • Consider proximity-dependent biotinylation (BioID) for weak/transient interactions

  • Validate interactions through reciprocal Co-IP

• Proximity Ligation Assay (PLA):

  • Combine FAX7 Antibody with antibodies against potential interaction partners

  • Quantify interaction signals through fluorescent spot analysis

  • Include appropriate controls (single primary antibodies)

• FRET-based approaches:

  • Use fluorescently labeled secondary antibodies within FRET distance

  • Perform acceptor photobleaching to confirm energy transfer

  • Calculate FRET efficiency to estimate proximity

These methods provide complementary approaches to map FAX7 protein interaction networks .

What are the considerations for using FAX7 Antibody in high-throughput screening applications?

For high-throughput applications:

• Reverse Phase Protein Array (RPPA) considerations:

  • Validate antibody linearity across concentration range

  • Implement appropriate normalization methods

  • Use Variable Slope (VS) normalization for slide-specific effects

  • Apply "SuperCurve" or similar algorithms for quantification

• Automated immunostaining platforms:

  • Optimize antibody concentration for automation

  • Include position controls to account for staining gradients

  • Validate reproducibility across multiple runs

  • Implement quality control metrics

• Data analysis and integration:

  • Develop robust signal quantification pipelines

  • Implement stringent statistical thresholds

  • Consider machine learning approaches for pattern recognition

  • Integrate with other -omics datasets

These approaches enhance throughput while maintaining data quality and reproducibility .

How can computational modeling help predict FAX7 Antibody binding characteristics?

Computational approaches for antibody binding prediction:

• Epitope mapping prediction:

  • Use protein structure prediction tools (e.g., AlphaFold2)

  • Apply B-cell epitope prediction algorithms

  • Consider surface accessibility and hydrophilicity

• Cross-reactivity assessment:

  • Perform sequence alignment against related proteins

  • Calculate epitope conservation scores

  • Predict potential off-target binding

• Binding affinity estimation:

  • Molecular dynamics simulations of antibody-antigen complexes

  • Free energy calculations for binding interactions

  • Comparison with experimentally determined structures of similar antibodies

These computational approaches can guide experimental design and interpretation, particularly for antibodies with limited characterization .

How might next-generation antibody technologies enhance FAX7 research?

Emerging technologies with potential applications for FAX7 research:

• Synthetic antibody alternatives:

  • Single-domain antibodies (nanobodies)

  • Synthetic binding proteins (DARPins, Affimers)

  • Aptamer-based detection systems

• Engineered antibody formats:

  • Bispecific antibodies for co-localization studies

  • pH-sensitive antibodies for trafficking studies

  • Split-antibody complementation systems

• Integration with emerging technologies:

  • CRISPR-based tagging for endogenous protein tracking

  • Optogenetic coupling for light-controlled protein manipulation

  • Multiplexed imaging with DNA-barcoded antibodies

These approaches may overcome current limitations and enable new research applications .

What considerations should be made when developing custom FAX7 Antibodies using phage display or other recombinant technologies?

For custom FAX7 antibody development:

• Antigen design considerations:

  • Use unique, surface-exposed regions

  • Consider multiple constructs (full protein vs. domains)

  • Evaluate post-translational modifications

• Selection strategy optimization:

  • Implement negative selection against related proteins

  • Consider CysDisplay for high-affinity clone retention

  • Use alternating selection strategies to avoid plastic binders

• Validation requirements:

  • Comprehensive cross-reactivity testing

  • Epitope binning to identify distinct binding sites

  • Comparison to existing antibodies when available

• Production considerations:

  • Format selection (scFv, Fab, IgG)

  • Expression system optimization

  • Stability engineering if needed

These approaches can yield renewable, sequence-defined antibody reagents with enhanced specificity .